International Archives of Otorhinolaryngology

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Editor-in-ChiefGeraldo Pereira Jotz

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ISSN 1809-9777Issue 2 Volume 19 April - May - June 2015

OTORHINOLARYNGOLOGYOffi cial Publication of the Otorhinolaryngology Foundation and Societas Oto-Rhino-Laryngologia Latina

INTERNATIONAL ARCHIVES OF

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International Archives of Otorhinolaryngology

ISSN 1809-9777

Editor

Geraldo Pereira JotzUFRGS, Porto Alegre, Brazil

Co-Editor

Aline Gomes BittencourtUSP, São Paulo, Brazil

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João Ferreira de Mello JuniorUSP, São Paulo, Brazil

Luiz Paulo KowalskiH. AC Camargo, São Paulo, Brazil

Marcelo M. HuebUFTM, Uberaba, Brazil

Marcos MocelinUFPR, Curitiba, Brazil

Marcus Miranda LessaUFBA, Salvador, Brazil

Michiel W. M. Van den BrekelNetherlands Cancer Institute, Amsterdam, Netherlands

Priscila Bogar RapoportFMABC, Santo André, Brazil

Ricardo Ferreira BentoUSP, São Paulo, Brazil

Ricardo L. CarrauOhio State University, OH, USA

Richard VoegelsUSP, São Paulo, Brazil

Robert T. Sataloff Drexel University College of Medicine, Philadelphia, USA

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Adriana Brondani da Rocha Conselho de Informações sobre Biotecnologia, São Paulo, Brazil

Adriane TeixeiraUFRGS, Porto Alegre, Brazil

Agrício CrespoUNICAMP, Campinas, Brazil

Agustin del CanizoUniversidad de Salamanca, Salamanca,Spain

Alberto Alencar NuldelmannPUC, Porto Alegre, Brazil

Alejandro RivasVanderbilt University Medical Center,Tennessee, USA

Alexandre Felippu NetoInstituto Felippu, São Paulo, Brazil

Alfi o FerlitoUdine School of Medicine, Unide, Italy

Ana Cristina H. HoshinoUSP, São Paulo, Brazil

André Luiz Lopes SampaioUNB, Brasília, Brazil

Antonio Celso Nassif FilhoPUC, Curitiba, Brazil

Badr Eldin MostafaAin-Shams University, Cairo, Egypt

Bernard FraysseHôpital PURPAN, Toulouse, France

Carlos Augusto Pires de OliveiraUNB, Brasília, Brazil

Carlos CuretUniversidad Nacional de Córdoba, Córdoba, Argentina

Carlos Diógenes Pinheiro NetoAlbany Medical College, New York, USA

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Eduardo Crema

UFTM, Uberaba, Brazil

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USP, São Paulo, Brazil

Elisabete Carrara de Angelis

Hospital AC Camargo, São Paulo, Brazil

Fabrizio Ricci Romano


Fayez Bahmad Junior

UNB, Brasília, Brazil

Filipe Matuba

Agostinho Neto University, Luanda, Angola

Fernando Luis Dias

INCA, Rio de Janeiro, Brazil

Francini Grecco de Melo Pádua

UNIFESP, São Paulo, Brazil

Francisco Verissimo de Mello Filho

USP-RP, Ribeirão Preto, Brazil

Gerson Schulz Maahs

UFRGS, Porto Alegre, Brazil

Giovanni Danesi

Ospedali Riuniti di Bergamo, Bergamo, Italy

Héctor Rondón Cardoso

Universidad Nacional de San Agustín, Arequipa, Perú

Heinz Stammberger

Graz University, Graz, Austria

Jacques Magnan

Université dAix-Marseille, Marseille, France

Jair Cortez Mantovani

UNESP, Botucatu, Brazil

Jeferson S. D’Avila

UFSE, Aracajú, Brazil

Jesús Algaba Guimera

Hospital Donostia de San Sebastián, San Sebastián, Spain

José Faibes Lubianca Neto

UFCSPA, Porto Alegre, Brazil

Jose N. FayadKeck School of Medicine, USC, California USA

Karine SchwarzUFRGS, Porto Alegre, Brazil

Lídio Granato

FCMSCSP, São Paulo, Brazil

Lilian Muniz

Universidade Federal de Recife, Recife, Brazil

Luiz Antonio Guerra Bernd

UFRS, Porto Alegre, Brazil

Luiz Lavinsky


Luiz Ubirajara Sennes


Maira Rozenfeld Olchik


Manuel Manrique Rodríguez

Universidad de Navarra, Navarra, España

Marcelo Lazzaron Lamers


Marcelo Ribeiro de Toledo Piza

Associação Paparella, Ribeirão Preto, Brazil

Márcio Abrahão

UNIFESP, São Paulo, Brazil

Márcio Nakanishi

UNB, Brasília, Brazil

Marcos Vial Goycoolea

Clinic of Las Condes, Santiago, Chile

Maria Valéria Schimidt Goffi Gómez


Mario AndréaLisboa University, Lisboa, Portugal

Mario SvirskyNew York University, New York, USA

Maurizio BarbaraSapienza University, SantAndrea Hospital, Rome, Italy

Minoru HiranoKurume University, Kurume, Japan

Nédio Steff enPUC, Porto Alegre, Brazil

Nelson RosárioUFPR, Curitiba, Brazil

O. Nuri ÖzgirginBaşkent University Faculty of Medicine, Ankara, Turkey

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Onivaldo CervantesUNIFESP, São Paulo, Brazil

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Paulo Sérgio Lins PerazzoUNEB, Salvador, Brazil

Pedro L. CoserUFSM, Santa Maria, Brazil

Pedro Luiz Mangabeira AlbernazUNIFESP, São Paulo, Brazil

Regina Helena Garcia MartinsUNESP, Botucatu, Brazil

Richard HarveyUniversity of New South Wales, New South Wales, Australia

Robert SweetMcGill University, Montreal, Canada

Robert VincentCausse Ear Clinic, Colombiers, France

Roberto Campos MeirellesUERJ, Rio de Janeiro, Brazil

Roberto Dihl AngeliUFRGS, Porto Alegre, Brazil

Roberto Eustáquio GuimarãesUFMG, Belo Horizonte, Brazil

Roberto FilipoSapienza Università di Roma, Roma, Italy

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Salvatore ConticelloUniversità degli Studi di Torino, Turin, Italy

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International Archives of Otorhinolaryngology

Volume 19, Number 2/2015

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Editorial

Original Research

99 Research Awards 2015Geraldo Pereira Jotz

100 Comparing Voice Self-Assessment with Auditory Perceptual Analysis in Patients with Multiple SclerosisVladimir Bauer, Zorica Aleric, and Ervin Jancic

106 Teleducation about Cleft Lip and Palate: An Interdisciplinary Approach in the Promotion of HealthCamila de Castro Corrêa, Thais Freire, Júlia Speranza Zabeu, Aline Martins, Rafael Ferreira, Paulo Afonso Silveira Francisconi, Jeniffer de Cássia Rillo Dutka, and Wanderléia Quinhoeiro Blasca

112 Parotid Incidentaloma Identifi ed by Positron Emission/Computed Tomography: When to Consider Diagnoses Other than Warthin TumorCarolina Bothe, Alejandro Fernandez, Jacinto Garcia, Montserrat Lopez, Xavier León, Miquel Quer, and Joan Lop

116 Preoperative Imaging Modalities to Predict the Risk of Regional Nodal Recurrence in Well-Diff erentiated Thyroid CancersMohammed K. AlNoury, Saad M. Almuhayawi, Khalid B. Alghamdi, and Khaled I. Al-Noury

121 Foreign Bodies in the Ear, Nose and Throat: An Experience in a Tertiary Care Hospital in Central NepalRamesh Parajuli

124 Diff erential Diagnosis and Treatment of Isolated Pathologies of the Sphenoid Sinus: Retrospective Study of 46 CasesThomas Ribeiro Marcolini, Maryane Cristine Safraider, Jan Alessandro Socher, Guilherme Olinto Lucena

130 Surfactant Protein A Expression in Chronic Rhinosinusitis and Atrophic RhinitisMohammad Waheed El-Anwar, Atef A. Hamed, Abd ElRaof Said Mohamed, Ahmad Abdel-Fattah Nofal, Maha A. Mohamed, and Hesham R. Abdel-Aziz

135 Mercury Exposure in a Riverside Amazon Population, Brazil: A Study of the Ototoxicity of MethylmercuryAna Hoshino, Heloisa Pacheco-Ferreira, Seisse Gabriela G. Sanches, Renata Carvallo, Nathália Cardoso, Maurício Perez, and Volney de Magalhães Câmara

141 Lipidomic Profi ling of Mastoid Bone and Tissue from Patients with Chronic OtomastoiditisFarbod Fazlollahi, Kessiri Kongmanas, Nongnuj Tanphaichitr, Jeffrey Suh, Kym Faull, andQuinton Gopen

151 Auditory Neuropathy/Dyssynchrony: A Retrospective Analysis of 15 CasesMurat Unal, and Yusuf Vayisoglu

156 Characterization of Hearing Thresholds from 500 to 16,000 Hz in Dentists: A Comparative StudyClaudia Giglio de Oliveira Gonçalves, Luciana Santos, Diolen Lobato, Angela Ribas, Adriana Bender Moreira Lacerda, and Jair Marques

161 Auditory Brainstem Response in Term and Preterm Infants with Neonatal Complications: The Importance of the Sequential EvaluationDaniela da Silva, Priscila Lopez, and Jair Cortez Mantovani

166 Health Promotion in Obstructive Sleep Apnea SyndromeCamila de Castro Corrêa, Wanderléia Quinhoneiro Blasca, and Giédre Berretin-Felix

171 The Study of Otoacoustic Emissions and the Suppression of Otoacoustic Emissions in Subjects with Tinnitus and Normal Hearing: An Insight to Tinnitus EtiologyLucieny Serra, Gabriela Novanta, Andre Lopes Sampaio, Carlos Augusto Oliveira, Ronaldo Granjeiro, and Silvia Cristina Braga

176 Olfaction in Neurologic and Neurodegenerative Diseases: A Literature ReviewMaria Dantas Costa Lima Godoy, Richard Louis Voegels, Fábio de Rezende Pinna, Rui Imamura, and José Marcelo Farfel

180 Ortner’s Syndrome: Secondary Laryngeal Paralysis Caused by a Great Thoracic Aorta AneurysmAna Claudia Alves Zangirolami, Frederico Vieira de Oliveira, and Miguel Soares Tepedino

183 Intralabyrinthine Penetrating Ventilation Tube with Preservation of Hearing: An Unusual Clinical SituationTantely Razafimahefa Raoelina, Maya Elziere, Justin Michel, and Arnaud Devèze

187 A Rare Location of Angiofi broma in the Inferior Turbinate in Young WomanAsif Salimov, and Serdar Ozer

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International Archives of Otorhinolaryngology Volume 19, Number 2/2015

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FMiv.indd 1FMiv.indd 1 17/03/15 2:03 PM17/03/15 2:03 PM

Research Awards 2015

Geraldo Pereira Jotz1

1Department of Morphological Sciences, Universidade Federal do RioGrande do Sul, Porto Alegre, RS, Brazil

Int Arch Otorhinolaryngol 2015;19:99.

Dear Colleagues,The journal International Archives of Otorhinolaryngology

has decided to award in 2015 the three best systematic reviewand/or literature review articles published in the journal in2014, highlighting contributions that laid the foundations andconcepts of universal knowledge. The purpose of the awards isto praise and acclaim distinguished authors, while also en-couraging new talent in the field of international research inotorhinolaryngology. This complies with our objective tostimulate excellence in education and welcome technologicaldevelopments through research and knowledge.

The most outstanding articles were:

Systematic Review

1°) Bittencourt, Aline Gomes; Burke, Patrick Rademaker;Jardim, Isabela de Souza; Brito, Rubens de; Tsuji, RobinsonKoji; Fonseca, Anna Carolina de Oliveira; Bento, RicardoFerreira. Implantable and Semi-Implantable HearingAids: A Review of History, Indications, and Surgery.Int. Arch. Otorhinolaryngol. 2014;18(3):303–310.

Review Article2°) Naik, Sulabha M.; Naik, Mahendra S.; Bains, Nainjot

Kaur. Cadaveric Temporal Bone Dissection: Is It ObsoleteToday? Int. Arch. Otorhinolaryngol. 2014;18(1):63–67.

3°) Campagnolo, Andrea Maria; Priston, Jaqueline; Thoen,Rebecca Heidrich; Medeiros, Tatiana; Assunção, Aída Regina.Laryngopharyngeal Reflux: Diagnosis, Treatment, andLatest Research. Int. Arch. Otorhinolaryngol. 2014;18(2):184–191.

We hope to see you again in future publications.Best regards,

Geraldo Pereira JotzEditor-in-ChiefInternational Archives of Otorhinolaryngology

Editorial

DOI http://dx.doi.org/10.1055/s-0035-1549262.ISSN 1809-9777.

Copyright © 2015 by Thieme PublicaçõesLtda, Rio de Janeiro, Brazil

Address for correspondenceGeraldo Pereira Jotz, MD, PhD,Universidade Federal do RioGrande do Sul, Porto Alegre, RS,Brazil (e-mail: [email protected]).

THIEME

Editorial 99

Comparing Voice Self-Assessment with AuditoryPerceptual Analysis in Patients with MultipleSclerosisVladimir Bauer1 Zorica Aleric1 Ervin Jancic2

1Department of Otorhinolaryngology, General Hospital Karlovac,Karlovac, Croatia

2Department of Neurology, General Hospital Karlovac,Karlovac, Croatia

Int Arch Otorhinolaryngol 2015;19:100–105.

Address for correspondence Vladimir Bauer, MD, Department ofOtorhinolaryngology, General Hospital Karlovac, A Stampara 3,Karlovac, Karlovac 47000, Croatia (e-mail: [email protected]).

Introduction

Multiple sclerosis (MS) is a neurologic diseasewith symptomsthat affect various organ systems. Voice and speech areaffected in 44% of patients with MS.1 Speech disorders aremuch more researched than voice disorders. Voice disordersare diagnosed using objective diagnostic methods. Phonatory

instability was proved through acoustic voice analysis2 andthrough electroglottography.3 Spectral voice analysis showedthat 70% of patients had symptoms of dysphonia comparedwith 33% of patients in the control group. The changes weremore notable among men than women. The basic frequency,basic frequency deviation, and jitter level are all higher.4 Evenmore importantly, objective voice changes were described by

Keywords

► multiple sclerosis► quality of life► voice quality► voice disorders

Abstract Introduction Disordered voice quality could be a symptom of multiple sclerosis (MS).The impact of MS on voice-related quality of life is still controversial.Objectives The aim of this study was to compare the results of voice self-assessmentwith the results of expert perceptual assessment in patients with MS.Methods The research included 38 patients with relapse-remittingMS (23 women and15 men; ages 21 to 83, mean ¼ 44). All participants filled out a Voice Handicap Index(VHI), and their voice sample was analyzed by speech and language professionals usingthe Grade Roughness Breathiness Asthenia Strain scale (GRBAS).Results The patients with MS had significantly higher VHI than control groupparticipants (mean value 16.68 � 16.2 compared with 5.29 � 5.5, p ¼ 0.0001). Thestudy established a notable level of dysphonia in 55%, roughness and breathiness in 66%,asthenia in 34%, and strain in 55% of the vocal samples. A significant correlation wasestablished between VHI and GRBAS scores (r ¼ 0.3693, p ¼ 0.0225), and VHI andasthenia and strain components (r ¼ 0.4037 and 0.3775, p ¼ 0.012 and 0.0195,respectively). The female group showed positive and significant correlation betweenclaims for self-assessing one’s voice (pVHI) and overall GRBAS scores, and between pVHIand grade, roughness, asthenia, and strain components. No significant correlation wasfound for male patients (p > 0.05).Conclusion A significant number of patients with MS experienced voice problems. TheVHI is a good and effective tool to assess patient self-perception of voice quality, but itmay not reflect the severity of dysphonia as perceived by voice and speech professionals.

receivedJuly 13, 2014acceptedNovember 14, 2014published onlineDecember 30, 2014



Original ResearchTHIEME

100

Dogan et al.5 In their sample, more significant differencesbetween theMS group and a control groupwere observed forjitter, shimmer, soft phonation index, and maximal phona-tion time. Using videostroboscopy, 16 of 27 patients showedincomplete glottal closure as an objective sign of vocal corddysfunction in phonation.5 All these and other studies claimthat many patients with MS suffer from vocal changes thatcan be detected and measured through diagnosticmethods.6–8

Some research has dealt with the subjective experience ofvocal changes and the impact of these changes on the qualityof life, but the results are conflicting. Dogan et al used theVoice Handicap Index (VHI) as a method for self-assessingvocal problems,5 but the authors did not describe the resultsin detail. Instead, they only concluded that the results werenot significantly different in patients with MS comparedwiththe control group. The same conclusionwas given in the studywhere only 12% of patients with MS described the difficultiesof dysphonia.9 On the other hand, the study of patients withMSperformedbyNatour showed significant differences in theself-assessment of vocal difficulties.10 VHI was used, and asignificant difference was observed between the total scoreand all subscores that were higher for patients with MS thanthe control group. The conclusion of this study is that MS canmake communicationmore difficult and cause a high sense ofvocal handicap. In our pervious study,11 the VHI score wassignificantly higher in theMS group. According to the study,12

the level of 12 points in the VHI test should be considered asthreshold for rating the biopsychosocial impact of dysphonia.Comparedwith previouslymentioned results, 44% of patientswith MS had a total VHI score higher than 12 (38% of malepatients and 48% of female patients). It is comparable withother previously mentioned results.2,3 Our present study isbased on the discrepancies regarding prevalence and impactof the voice changes in MS. We completed and equalized ourpatient sample, and we included only patients in relapse-remitted form of MS. The aim of this study is to compare theresults of voice self-assessment with the results of expertperceptual assessment. It is our assumption that vocal diffi-culties reported by patients with MS will be confirmed andrated by a listener, a speech and vocal therapist.

Methods

This is a cross-sectional case–control study, inwhich voices ofparticipants with and without MS were evaluated by self-perception questionnaire and hearing perceptual evaluation.Patients with MS were selected randomly from a list ofpatients attending Neurology Polyclinic for their routinefollow-up visits, using the Hospital Information System data-base. The control groupwas composed of individuals withouthistory of neurologic disease and disorder of communicationrecruited from hospital staff and patients’ companions. Ex-clusion criteria were endotracheal intubation within3 months before entry, history of laryngeal malignancy, anoperation performed on the larynx and vocal chords, or anacute upper airway respiratory infection. The institutionalMedical Ethical Committee approved the research.

All participantswere administered the VHI, a standardized30-point questionnaire suggested by Jacobson et al in 1997,13

to determine the effect of vocal problems on the quality of life.This is a commonly used method of biopsychosocial self-assessment of vocal difficulties. Each answer is graded 0 to 4depending on the frequency of the difficulty: 0 ¼ no difficul-ties, 1 ¼ almost never, 2 ¼ sometimes, 3 ¼ almost always,4 ¼ always. The questionnaire is divided into three subscales;the first refers to claims for self-assessing one’s own voice(pVHI), the second refers to statements regarding the effect ofvoice in daily functions (fVHI), and the third refers to the effectof vocal difficulties on the participants’ emotional status(eVHI). We used the Croatian version of the VHI.14

Hearing perceptual voice evaluation was performed usingGRBAS scale according to the Japanese Society of Logopedicsand Phoniatrics.15 The voice of each participant reading astandard text for 2 minutes was recorded on tape. The voicesample was collected in a quiet room with the microphonepositioned 5 cm away from the lips. Voice recordings wereassessed by three experienced speech therapists in a double-blind, randomized fashion. The examiners assess the gradesof dysphonia (G), roughness (R), breathiness (B), asthenia (A),and strain (S) by listening to patient’s voice, using a scale from0 to 3 where 0 ¼ regular, 1 ¼ mildly pronounced, 2 ¼ mod-erate, 3 ¼ very pronounced.

The Expanded Disability Status Score (EDSS), a standard-ized system for assessing neurologic deficit based on thegrade of nine functional systems inwhich a score of 0 is a statewith no neurologic manifestations and 10 is death by dis-ease,16 was determined for all patients.

Statistical analysis was performed using free statisticalsoftware.20 The regularity of data distribution was testedusing the Kolmogorov-Smirnov test. The VHI scores were notnormally distributed.We used descriptive statisticalmethodsto show features of groups and subgroups—mean values andstandard deviation and median and interquartile range fordatawith irregular distribution.We used nonparametric teststo measure differences in significance between the groups—chi-square for category variables and the Mann-Whitney testfor the numerical data. The correlation between groups ofparticipants was tested by the Spearman rank test. The levelof significance was set to p < 0.05.

Results

The research included 38 patients with MS, 23 women and15 men, ages 21 to 83 (mean ¼ 44). The mean length ofdisease was 12.8 years (range 1 to 48). All participants hadrelapsing-remitting type of disease. We compared themwithparticipants from a control group (n ¼ 38, 21 women, 17men) of corresponding age andwithout signs of MS and voicedisorders. A detailed overview of participant features isshown in ►Table 1. The patients with MS had significantlyhigher VHI than control group participants (mean value16.68 � 16.2 compared with 5.29 � 5.5, p ¼ 0.0001). ThepVHI, fVHI, and eVHI values were significantly higher forpatients with MS. When observed separately, women withMS had higher VHI values than men (18.39 compared with

International Archives of Otorhinolaryngology Vol. 19 No. 2/2015

Voice Self-Assessment in Patients with Multiple Sclerosis Bauer et al. 101

13.84). If we take 12 points as a cutoff point in the VHI test,47% of all patients (46% of men and 49% of women) reportednoticeable voice difficulties. Detailed results of the pVHI aredescribed in ►Table 2.

All components of the GRBAS scale were significantlyexpressed in the patient group (►Table 3). The researchersestablished a significant level of dysphonia (G ¼ 1, 2, or 3) in55%, roughness and breathiness in 66%, asthenia in 34%, andstrain in 55% of vocal samples, and 34% of participants hadone component of the GRBAS scale graded as severe.

►Table 4 shows correlation values for VHI self-assessmentand GRBAS scale auditory perceptual assessment. A signifi-cant positive correlation was established for all participantsbetween VHI and GRBAS scores (r ¼ 0.3693, p ¼ 0.0225), andVHI and A and S components (p ¼ 0.012 and 0.0195, respec-tively). If we observe only pVHI, the part that refers to sense ofvocal difficulties, the correlations are even more significant(r ¼ 0.42 to 0.53;►Table 4). This was observed both for maleand female patients. No significant correlation was found formale patients (p > 0.05). The female group showed positiveand significant correlation between pVHI and overall GRBASscores, and between pVHI and G, R, A, and S components(►Table 4).

All our participants had relatively mild functional difficul-ties (EDSS ¼ 2.7 � 1.06). The results of self-assessment andperceptive assessment could not be related with the extent ofneurologic deficit (expressed with EDSS), nor with the dura-tion of the disease. Among all the observed parameters, asignificant correlation was observed only between EDSS andA component of the GRBAS scale (r ¼ 0.6, p ¼ 0.0025) andbetween the duration of the disease and the A component ofthe GRBAS scale (r ¼ 0.53, p ¼ 0.0007), only for femalepatients.

Discussion

Phonation is not just an expiration that produces vibrationsbut a complex physiologic process that includes a coordinatedcooperation of multiple organ systems. The nervous system isespecially involved. The laryngeal part of the phonatoryfunction can be disrupted by the damages on several levelsof the nervous system. The cranial nerve X (i.e., the vagusnerve) is responsible for direct motor innervation of theintrinsic laryngeal muscles. Its fibers stem from the bodiesof cells located in the medullary reticular formation in thenucleus ambiguus. The neuropathology that may affect the

Table 2 Results obtained using VHI in patients with multiple sclerosis and control group

Patients Control group p value

Total VHI, mean (range) 16.68 (0–85) 5.29 (0–20) 0.0001

fVHI, mean (range) 4.58 (0–25) 1.34 (0–10) 0.001

pVHI, mean (range) 9.22 (0–36) 3.55 (0–6) 0.001

eVHI, mean (range) 2.88 (0–24) 0.4 (0–2) 0.004

Total VHI male, mean (range) 13.85 (0–48) 4.61 (0–16) 0.055

Total VHI female, mean (range) 18.39 (0–85) 5.42 (0–20) 0.001

Abbreviations: eVHI, emotional subscale; fVHI, functional subscale; pVHI, physical subscale; VHI, Voice Handicap Index.

Table 1 Participant characteristics

Patients Control group

Total n 38 38

Male/female 15/23 17/21

Age (mean/range) 44/21–83 43/18–72

Duration of disease, y(mean/range)

All patients 12.7/1–48

Male 12.9/1–31

Female 12.6/2–48

EDSS (mean � SD)

All patients 2.7 � 1.06

Male 2.54 � 0.96

Female 2.78 � 1.1

Abbreviations: EDSS, Expanded Disability Status Score; SD, standard deviation.


Voice Self-Assessment in Patients with Multiple Sclerosis Bauer et al.102

voice can be located even more centrally. The vagus nervenuclei are affected by the pyramidal and extrapyramidalsystem. The pyramidal system establishes its influencethrough the corticobulbar tract and the extrapyramidal sys-tem through the basal ganglia. As a demyelinating process,MS

can cause lesions at any height of the nervous system,subcortically in the white matter; in the cores of the brain-stem; in the medulla oblongata, cerebellum, peripheralnerves; in the already mentioned vagus nerve; and in nervesthat participate in respiration.17 With such complex

Table 3 Results using GRBAS scale in patients with multiple sclerosis patients and control group

Patients (n ¼ 38) Control group (n ¼ 38)

0 1 2 3 All 0 1 2 3 All p value

G 17 2 17 2 38 28 7 2 1 38 0.0025

R 13 1 21 3 38 29 4 4 1 38 0.0008

B 13 2 23 0 38 32 1 4 1 38 0.0002

A 25 1 7 5 38 33 3 2 0 38 0.0196

S 17 2 16 3 38 32 2 3 1 38 0.0071

Abbreviations: A, asthenia; B, breathiness; G, grade; R, roughness; S, strain.

Table 4 Correlation of auditory perception with self-assessment and duration of the disease

GRBAS G R B A S

All

VHI

r 0.3693a 0.2379 0.1975 0.1058 0.4037a 0.3775a

p value 0.0225a 0.1503 0.2346 0.5272 0.012a 0.0195a

pVHI

r 0.5289a 0.3408a 0.2503 0.0601 0.4433a 0.4209a

p value 0.0006a 0.0363a 0.1295 0.72 0.0053a 0.0085a

Duration

r 0.0098 0.2265 0.1008 0.0497 0.5279a 0.1589

p value 0.9532 0.1715 0.547 0.7669 0.0007a 0.3407

Male

pVHI

r 0.1822 0.2038 0.0284 0.0462 0.3368 0.2993

p value 0.5157 0.4663 0.92 0.8701 0.2196 0.2785

Duration

r 0.0359 0.1242 0.0021 0.2713 0.4442 0.0062

p value 0.8989 0.6593 0.9939 0.3281 0.0972 0.9825

Female

pVHI

r 0.7056 0.633a 0.4597a 0.182 0.4888a 0.3994a

p value 0.0002a 0.0012a 0.0273a 0.4059 0.0179a 0.059a

Duration

r 0.0301 0.2543 0.1366 0.1118 0.5777a 0.2383

p value 0.8916 0.2416 0.5341 0.6117 0.0039a 0.2736

Abbreviations: A, asthenia; B, breathiness; G, grade; GRBAS, Grade Roughness Breathiness Asthenia Strain scale total score; pVHI, physical subscale ofVoice Handicap Index; R, roughness; S, strain; VHI, Voice Handicap Index.aMarked values correlated significantly.



possibilities of emergence of dysphonia in MS, the psycho-logical, social, and emotional importance of voice contributesto the importance and the complexity of its effect on thequality of life. Our research shows that patients with MSdisplay more intense vocal difficulties than people withoutMS. The VHI score was above 12 for 47% of the participants.

The quality of life for patients with MS is a commonresearch subject. Although objective estimates of vocalchanges are well researched and objectively confirm symp-toms of dysphonia in 30 to 70% of participants,2–8 little isknown about the effect of dysphonia on the quality of life inMS. The research done on 27 female patients with MS usedVHI and GRBAS.5 The pVHI showed no significant differencebetween the patients and the control group. The results of theGRBAS scale showed markedly higher A and S components inthe MS group (p < 0.0001 for A and p ¼ 0.04 for S). Althoughexpressed somewhatmore oftenwith patients, the G, R, and Bcomponent were not significantly different. A study of 59participants with MS found no significant differences in theoverall VHI score and subscores compared with the scores ofhealthy participants (VHI ¼ 5.9, pVHI ¼ 2.4, fVHI ¼ 2.2, andeVHI ¼ 1.4).9 Only 8.7% of participants had serious vocaldifficulties (VHI > 15). Contrary to that, the work by Natouret al analyzed VHI with 39 patients with MS, and a significantdifference between patients with MS and healthy partici-pants was noticed for both male and female participants(male VHI ¼ 27.7 compared with VHI ¼ 10.1 in the controlgroup and female VHI ¼ 19 comparedwith VHI ¼ 11.4 in thecontrol group).10 The differencewas significant for total scoreand all three subscores. These scores, along with those fromour research, can be compared according to their similaritiesand differences. All four studies used the VHI scale, and twoused the GRBAS scale. The results of self-assessed VHI differgreatly. Some of that difference can be explained by thevarious degrees of total neurologic damage. Studies describ-ing insignificant differences have participants with a relative-ly low EDSS score: 1.94 (8) and 1.22 (5), although mean EDSSscore for our studywas 2.7. A score of 1.5 to 2 is interpreted asa state without neurologic incidents, with only minimal signsof damage to the functional systems. A score of 2.5 to 3 impliesmild incidents in the two functional systems. Not only doeshigher EDSS imply more total damage, the incidents, nomatter how gentle, additionally sensitize the participants toother difficulties that the EDSS does not measure. Althoughthe duration of the disease was correlated with either VHI orGRBAS scale in our study, it should be noted that there is adifference in disease duration between our sample andpatients from the previously mentioned studies (12 yearscompared with 6 and 7 years). Linguistic and cultural differ-ences among the participants, such as being speakers ofvarious languages, could affect the perception of vocal diffi-culties and were not the subject of our research.

To make our self-assessment results more valuable, theywere complemented with the results of perceptual voiceassessment. Perceptual evaluation is fundamental in assess-ing voice quality, the relevance of defects, and the defects’impact on the subject’s ability to communicate. We tried tofind the differences in the vocal perceptions between the

participants expressed by VHI and by the listeners expressedby the GRBAS scale. Because the VHI is divided into threesubscales, we expected the biggest congruence between thephysical aspect (pVHI), which describes the perception ofvocal difficulties, and the GRBAS scale. Smaller or no congru-ence was expected for the fVHI and eVHI. We found nosignificant correlation between overall VHI or its subscalesand anyof the GRBAS scale components formale participants.We found significant positive correlation between VHI andpVHI with the GRBAS overall score and the components G, B,A, and S among female participants. We found very fewstudies that compared self-assessment with the perceptualassessment for patients with MS. The results were somewhatcomparable with an already mentioned study,5,11 in which Aand S components of the GRBAS scale were graded signifi-cantly different for female patients with MS. In this case, alower EDSS score could explain the differences between theexpression of G and B components of the GRBAS scale (1.2comparedwith 2.7).We cannot explainwhy the VHI result formale participants is not in correlation with the perceptualanalysis. The VHI is a good and effective tool to assess apatient’s self-perception of the voice quality, but it may notreflect the severity of dysphonia as perceived by voice andspeech professionals. It seems that components other thanthose directly related to voice quality may contribute toresponses on voice evaluation completed by patients. Genderis one of these. Certainly the gender of patients is a factor, butwe assumed that the gender of listeners also could be a factor.All of our three voice professionals were female, possiblymore sensitized on voices of their own gender. In any case, theresearch should be repeated on a large number of maleparticipants using several questionnaires regarding the qual-ity of voice and assessors of both genders. The same degree ofdysphonia can impact different patients in very differentways. Factors such as duration of the disorder and age andsocial setting of the patient, along with the patient’s occupa-tion status, profession, general health, or other factors, mayact to blunt or accentuate the effect of dysphonia upon aparticular patient’s voice related quality of life. However,important clinical information may be missed if only thepatient’s perspective is considered. A perceptual assessmenttool such as the GRBAS scale gives valuable information aboutthe extent of altered vocal physiology.

One parameter in significant correlation with total neuro-logic deficit and the disease length is the asthenia componentof the GRBAS scale. Respiratory function is often affected byMS. Weakness of respiratory muscles is characterized bylowered maximal expiratory pressure7,10; weakness has adirect effect on the EDSS value and worsens with time. Amaintained expiratory pressure enables adequate subglotticpressure and thus the movement of vocal cords and highquality of produced voice. Otherwise, the voice is weak andasthenic. Incomplete glottal closure, described as common inMS byDogan et al,5 aswell as posterior glottal chink, definitelycontributes to vocal asthenia. The proportion of our partic-ipants with expressed subjective vocal difficulties (VHI ¼ 47%,GRBAS ¼ 55%) is comparable to the ratio of dysphonic voicesin the studies that used objective vocal analyses (30 to 70%).3–5


Voice Self-Assessment in Patients with Multiple Sclerosis Bauer et al.104

Therefore, it is justifiable to think that a significant number ofpatients with MS have and feel vocal difficulties. It is obviousthat the effect of vocal difficulties on the quality of life isusually not strong. VHI is markedly different than that ofcontrol group participants, but in most cases it implies milddifficulties. Only three participants from our research ratedtheir VHI as moderately altered, and only one patient asseverely altered (VHI ¼ 85). Unfortunately, MS is a diseasewith deficits that strongly affect mobility, independence,bladder control, sexual function, among others. When com-paredwith the difficulties described previously, it is easy to seethat vocal difficulties would get more attention if theywere anisolated vocal disorder.

It is well known that MS can lead to hearing impairment.Moderate hearing loss occurs in 23% of patients,18 anddifficulty swallowing is seen in 43% of patients with MS.19

Vocal disorders, speech, swallowing, and hearing difficultiescan cause significant problems and disrupt the quality of life,especially when occurring simultaneously. Some of the pre-viously mentioned symptoms could be the first signs of thedisease and a reason to visit the otorhinolaryngologist.Although MS is not a primary otorhinolaryngologic disorder,it should be included in a differential diagnostic process.

A lack of objective measurements limits our study. How-ever, we focused on the patients’ experience of the quality oftheir voice compared with subjective analysis performed byvoice professionals. We had a small number of patients(n ¼ 38); we plan further research with a larger number ofpatients, more parameters, and comparisons with other non-motor dysfunctions in patients with MS.

Conclusion

In our study, almost half of patients with MS feel and describemild dysphonic difficulties that affect the quality of life. Theperception of dysphonic difficulties is individual, not always inaccordancewith thehearing perceptual vocal analysis. Positivemoderate correlation between self-assessment and perceptualvocal assessment was found for our female patients with MS.

References1 Hartelius L, Svensson P. Speech and swallowing symptoms associ-

ated with Parkinson’s disease and multiple sclerosis: a survey.Folia Phoniatr Logop 1994;46(1):9–17

2 Hartelius L, Buder EH, Strand EA. Long-term phonatory instabilityin individuals with multiple sclerosis. J Speech Lang Hear Res1997;40(5):1056–1072

3 Konstantopoulos K, Vikelis M, Seikel JA, Mitsikostas DD. Theexistence of phonatory instability in multiple sclerosis: anacoustic and electroglottographic study. Neurol Sci 2010;31(3):259–268

4 Feijó AV, Parente MA, Behlau M, Haussen S, de Veccino MC,Martignago BCDF. Acoustic analysis of voice in multiple sclerosispatients. J Voice 2004;18(3):341–347

5 Dogan M, Midi I, Yazici MA, Kocak I, Günal D, Sehitoglu MA.Objective and subjective evaluation of voice quality in multiplesclerosis. J Voice 2007;21(6):735–740

6 DuranovicM, Salihovic N, Ibrahimagic A. Characteristics of voice inindividuals with multiple sclerosis. Mater Sociomed 2011;23(1):23–27

7 Chiara T, Martin D, Sapienza C. Expiratory muscle strength train-ing: speech production outcomes in patients with multiple scle-rosis. Neurorehabil Neural Repair 2007;21(3):239–249

8 Yamout B, Fuleihan N, Hajj T, et al. Vocal symptoms and acousticchanges in relation to the expanded disability status scale, dura-tion and stage of disease in patients with multiple sclerosis. EurArch Otorhinolaryngol 2009;266(11):1759–1765

9 Hamdan AL, Farhat S, Saadeh R, El-Dahouk I, Sibai A, Yamout B.Voice-related quality of life in patients with multiple sclerosis.Autoimmune Dis 2012;2012:143813. doi: 10.1155/2012/143813

10 Natour Y, Marie B, Aljunidy L. The respiratory muscle capabilitiesof Jordanian patients with multiple sclerosis. J Voice 2012;26(6):811.e15–811.e18

11 Bauer V, Aleric Z, Jancic E, Knezevic B, Prpic D, Kacavenda A.Subjective and perceptual analysis of voice quality and relation-ship with neurological disfunction in multiple sclerosis patients.Clin Neurol Neurosurg 2013;115(Suppl 1):S17–S20

12 Niebudek-Bogusz E, Kuzańska A, Woznicka E, Sliwinska-Kowal-ska M. Assessment of the voice handicap index as a screeningtool in dysphonic patients. Folia Phoniatr Logop 2011;63(5):269–272

13 Jacobson BH, Johnson A, Grywalski C, et al. The Voice HandicapIndex (VHI): development and validation. Am J Speech Lang Pathol1996;6:66–70

14 Bonetti A, Bonetti L. Cross-cultural adaptation and validation ofthe Voice Handicap Index into Croatian. J Voice 2013;27(1):130.e7–130.e14

15 Hirano M. Clinical Examination of Voice. New York, NY: Springer;1981:81–84

16 Kurtzke JF. Rating neurologic impairment in multiple sclerosis: anexpanded disability status scale (EDSS). Neurology 1983;33(11):1444–1452

17 Aronson A, Bless D. Clinical Voice Disorders. New York, NY:Thieme; 2009:73–97

18 Peyvandi A, Naghibzadeh B, Ahmady Roozbahany N. Neuro-oto-logic manifestations of multiple sclerosis. Arch Iran Med 2010;13(3):188–192

19 Thomas FJ, Wiles CM. Dysphagia and nutritional status in multiplesclerosis. J Neurol 1999;246(8):677–682

20 Wessa P. Free Statistics Software. Office for Research Developmentand Education, version 1.1.23-r7. 2012. Available at: http://www.wessa.net/. Accessed 14.04.2012.



Teleducation about Cleft Lip and Palate: AnInterdisciplinary Approach in the Promotionof HealthCamila de Castro Corrêa1 Thais Freire1 Júlia Speranza Zabeu1 Aline Martins1 Rafael Ferreira2

Paulo Afonso Silveira Francisconi3 Jeniffer de Cássia Rillo Dutka1 Wanderléia Quinhoeiro Blasca1

1Department of Speech-Language Pathology and Audiology,Universidade de São Paulo, Bauru School of Dentistry, Bauru, SaoPaulo, Brazil

2Department of Prosthesis, Universidade de São Paulo, Bauru Schoolof Dentistry, Bauru, São Paulo, Brazil

3Department of Operative Dentistry, Universidade de São Paulo,Bauru School of Dentistry, Bauru, São Paulo, Brazil


Address for correspondence Camila Corrêa, BS, MSc, Department ofSpeech-Language Pathology and Audiology, University of São Paulo,Bauru School of Dentistry, Octávio Pinheiro Brisola Street, Bauru, SaoPaulo 17012-901, Brazil (e-mail: [email protected];[email protected]).

Introduction

In Brazil, data suggest that there are �260,000 individualswith cleft lip and palate (CLP). This is the third or fourth mostcommon birth defect in the country. People with CLP and

craniofacial deformities may have functional disorders thatimpair eating, speech, and dental-occlusal relationships.1,2

Craniofacial malformations may be associated with physicaldisfiguration and communication disorders, whichmay harmthe social lives of individuals who are afflicted by them. There

Keywords

► cleft palate► speech, language,

and hearing sciences► dentistry► health education► telehealth

Abstract Introduction The Young Doctor Project (YDP) uses Telehealth and Interactive Tele-ducation instruments to promote the integration of different areas of health and to buildknowledge. This methodology can also foster public awareness on various issues relatedto health. In this context, the objective of this study was to emphasize cleft lip and palate(CLP), which is one of the most common birth defects in Brazil.Objective The study aimed to apply a model of education regarding CLP, based on thedynamics of the YDP, and to evaluate the participants’ knowledge acquired afterparticipating in the YDP.Methods The participants were 41 students, 13 to 15 years of age and at the eight-and ninth-grade levels in a private elementary school in Bauru (Brazil). To analyze theperformance of the participants, a questionnaire was administered before and after thecompletion of the training program. The training program was structured in three stepsusing: (1) interactive teleducation classes, (2) a cybertutor, and (3) practical activities.Results There was a statistically significant difference between the pre- and post-participation questionnaire results. The improved performance of participants isevidenced by the increase in the rate of correct answers on all issues.Conclusion The YDP on CLP was applied in the school setting following the three stepsrecommended by the project, and, after the implementation of the training program,there was a significant increase in participants’ knowledge of CLP. The YDP on CLPproved an effective tool in promoting health education.

receivedAugust 2, 2014acceptedDecember 14, 2014published onlineJanuary 19, 2015




106

is a high frequency of isolation, social discrimination, andbullying in this population.3 It is the role of health profes-sionals to promote health education that improves interper-sonal relationships and facilitates social inclusion ofindividuals with impairment related to malformation intosociety.4

Health education can be understood as a social practicethat enables critical and transformative reflection and behav-ioral change as well as promotes the welfare of patients.4

Besides traditional methodologies in education, informationand communication technologies can be used to disseminatehealth knowledge, especially regarding the promotion ofpositive attitudes toward health.5 The Young Doctor Project(YDP), therefore, is an initiative that involves the use ofinteractive teleducation, integrating the areas of health andeducation, to promote classroom experiences that fosterknowledge construction and exchange among elementary,middle, and high school students.6,7 The YDP is an excellentteachingmethodology that enables health education promot-ing awareness of the role of each person as an active healthagent in the community.8 The project started in 2007 andsince then has been used across the country in differentregions and sectors, establishing a link between students ofelementary, middle, and high school and teachers.9,10

When used within the area of speech-language pathologyand audiology, the YDP has addressed topics related tohearing and vocal health using various venues to conveyinformation and exchange knowledge including activitieswithin the classroom, cybertutors, and use of software suchas “Virtual Man” to promote integration of knowledge andmotivate involvement.11,12 Other YDPs have addressed thetopic of genetic syndromes13 as well as oral health.14

The current YDP addressed the topic of CLP integrating theareas of speech-language pathology, audiology, and dentistry,aiming to improve public awareness regarding one of themost common birth defects. To accomplish the task of healthpromotion, this study had two objectives: to develop andimplement a health education strategy based on the dynamicmethodologyof the YDPwhile addressing the topic of CLP andto evaluate the performance of school-aged children regard-ing the acquisition of knowledge about CLP after attendingthe YDP.

Materials and Methods

Ethical AspectsThe study was approved by the Ethics Committees of theinstitution involved in the YDP. To conduct this project,partnerships were established between two universities,one hospital specialized in the management of CLP and aprivate elementary-middle school.

ParticipantsAll 50 middle school students from eighth- and ninth-gradelevels were invited to participate in this study. Because not allstudents had access to the Internet outside the school envi-ronment, the project was conducted at the school informaticslaboratory. The sample was limited to 50 students because

there were 25 computers with Internet access available, andthe students accessed the cybertutor in pairs at a timesuitable to all. Only students who signed the informedconsent form together with their parents were included. Toselect a site to conduct the project, the researchers identifieda school that had students with CLP attending regular classesand also identified teachers and administrative staff interest-ed in the project. The participating school fit the criteria andparticipated as the site for this project.

ProceduresAfter identifying the school and the students, the participantsunderwent the YDP’s (http://www.projetojovemdoutor.org.br/) training program, which is structured in three steps.

Step 1—Lectures about the Topic of InterestAt the first meeting, the participants attended a lecture givenby the researchers, which lasted 2 hours. The class was held atthe university responsible for this study and addressed topicsrelated to CLP: predisposition to bullying and discrimination,definition, classification, embryology, etiology, diagnostics,team treatment, family structure and support, and speech-language pathology, audiology, and dental aspects of CLPmanagement. A PowerPoint (Microsoft Corp., Redmond,Washington, United States) presentation including illustra-tions and videos was prepared and used during the first class.

Step 2—CybertutorEach participant received a login ID and password to access anonline classroom on the YDP Web site (cybertutor). Theinteractive teleducation was meant to facilitate the comple-tion by the students. Participants were able to control theirown learning, accessing the cybertutor freely and establishingtheir own study schedules, within the period assigned forreview of the material. The interactive classroom allowedcollaboration between the users through mailing lists andforums to clarify doubts about the content available oncybertutor. The cybertutor development involved a partner-ship between the two universities engaged in this study.

The topics addressed in the cybertutor regarding CLPincluded definition, embryology (how and when CLP occursduring pregnancy), etiology, diagnostics, treatment, firstcaregiver concerns, and first care for babies with CLP, alongwith presentation of psychosocial issues related to bullyingand prejudice associated with CLP. After completion of theintroductory content on CLP, more specific information aboutspeech-language pathology, audiology, and dental aspectsrelated to the management of CLP were made availablethrough the cybertutor.

Step 3—Sharing the Knowledge Gained (PracticalActivities)At step 3, the participants were quizzed regarding the topicspresented during the lecture and with the cybertutor. Stu-dents were divided into five groups to take the quiz involvinggroup interactions. The group that had the highest number ofcorrect answers was encouraged to explain those topics ofmastery to the other groups. The participants were than


Teleducation about Cleft Lip and Palate Corrêa et al. 107

acknowledged by the team of researchers who advised themduring the process along with their teachers as “youngdoctors.” They were also guided to organize social activitiesaimed at disseminating the knowledge gained during theYDP. A chart demonstrating the sequence of steps takenduring the process for the students to become young doctorsin the topic of CLP is presented in ►Fig. 1.

Assessment ToolTo analyze the learning of the participants, they were admin-istered questionnaires before and after completion of the YDP.The questionnaire for the assessment of student performanceconsisted of two essay questions (questions 1 and 13) and 11multiple-choice questions (questions 2 through 12). The ques-tionnairewas developedby the research teamand reviewedbyprofessionals from the hospital specializing in the manage-ment of CLP. The essay questions addressedgeneral knowledgeof the students about CLP (question 1: What do you knowabout the cleft lip and palate?) and also investigated thestudents’ opinion regarding differences between individualswith and without cleft (question 13: Are there differencesbetween individuals with and without cleft? If yes, describe).To determine the participants’ performance in the essayquestions, the researchers searched for three main responses.The researchers expected to read answers that CLP is present atbirth (congenital anomaly); that the cleft is an opening that canaffect the lip and/or palate, changing the face and mouth; andthat the presence of CLP does not involve cognitive problemsthat can affect intellectual abilities. If the participant includedat least two of these three responses, the reply to the essayquestions was considered correct. The multiple-choice ques-tions addressed the content included in the cybertutor, withone correct answer among five possibilities. The findings wereanalyzed statistically using the Student paired-samples t test,adopting a significance level of 5%.

Results

ParticipantsAfter inviting all 50 students from eighth- and ninth-gradelevels, 41 accepted and signed the informed consent formsalong with their parents. Nine students (18%) did not acceptthe invitation, indicating that they were not interested in theproject. The participants were 16 (39%) boys and 25 (61%)girls with ages between 13 and 15 years (mean: 13 years).

Procedures

Step 1—Lectures about the Topic of InterestBefore the initial lecture with the participants, the question-naires to evaluate the knowledge of the 41 participants onissues regarding CLP were applied for baseline data. Theparticipants attended the lecture and the interaction withthe researchers to discuss the content presented about CLP.During the lecture, before discussing the information, theopinion of the students was frequently asked and they wereinvited to reflect about the content provided. During theinteractions, the researchers identified the overall level ofknowledge of the group about CLP. It was noted that theinvolvement of the participants was enthusiastic, as theyremained attentive and asked questions during the lecture.

The interaction between researchers and students was thevenue for introduction of information about CLP. The contentaddressed during the three steps of the YDP included: definition,terminology comprising technical terms related tomanagementof CLP, possible disorders and consequences related to malfor-mations, and treatment options. The information provided inthis first meeting was essential to enable the students tounderstand the issues addressed in the step 2 with the cybertu-tor. Therefore, the material provided in the initial lecture wasdesigned to foster the learning intended at the endof the project.

Fig. 1 Organizational chart of procedures performed during the Young Doctor Project in cleft lip and palate.


Teleducation about Cleft Lip and Palate Corrêa et al.108

Step 2—CybertutorAfter the initial lecture, the students were told that they had1 month to access thematerial presentedwith the cybertutor.The majority of the students (87.80%) accessed all of thesubjects in cybertutor and 75.60% accessed the two additionalforums that were available. The forums promoted interac-tions between participants and researchers and allowed theclarification of doubts about the content in the cybertutor.

Step 3—Sharing the Knowledge Gained (PracticalActivities)The students were asked to share and expand the knowledgegained in steps 1 and 3 during interactions in a group quizactivity. During the quiz, some questions/doubts regardingthe content addressed in the initial lecture and the cybertutorwere resolved (►Fig. 2). To solve these questions, the re-searchers asked the help of the participating students toclarify the doubts of their colleagues.

Students, teachers, and researchers organized pam-phlets, plays, games, workshops, and lectures about CLPto share and display within the school environment(►Fig. 3). These social actions enabled the young doctorsto disseminate knowledge to other students in theirschools, family, and community.

Data AnalysisTo assess the knowledge retained by the participants, theywere presented with questionnaires before and after theYDP. ►Table 1 presents the overall scores for the 41 partic-ipants for the 13 questions with the corresponding amount(n) and percentage (%) of correct answers. The questionnairesbefore the YDP revealed that 9 (69%) of the 13 questions wereanswered with less than 75% accuracy by 41 students (ques-tions 1, 3, 4, 6, 7, 8, 9, 10, and 12) and that 4 of these 9questions were answered with less than 50% accuracy. Thehighest rates of correct answers were observed for thecontent addressing classification, diagnostics, and differencesbetween individuals with andwithout CLP (questions 2, 5, 11,and 13). Question 11 alreadywas answeredwith 100% correctanswers before YDP.

►Table 1 also presents the overall scores obtained on thequestionnaire after the YDP. The number of correct answersincreased for all questions except question 11. The differences

between the before and after YDP conditions revealed animprovement in correct answers that ranged between 4.8 and88.8%. Inferential statistical comparison of mean correctanswers before and after the YDP revealed a statisticallysignificant difference between the two conditions(p ¼ 0.001, t ¼ 6.687).

Discussion

Interactive teleducation involves the optimization of techno-logical and educational resources for the purpose of stimu-lating interactivity, promoting self-directed learning, andmaintaining student interest and motivation.13,14 Maintain-ing student interest during educational activities in particularmay foster learning and knowledge retention.6,8,12Apreviousstudy suggested that the development of health-educationprograms should be encouraged by educational practices,especially those that involve interactive teleducation.4

The results of the YDP about CLP corroborate the findingsof previous studies,4,6,8,12–14 revealing an increase in stu-dents’ knowledge after participating in a learning programinvolving interactive teleducation. A statistically significantdifference between the number of correct answers before andafter the YDP suggested that the interactive educationalstrategies employed in this study aided the learning process.The YDP was shown to be an effective tool to disseminateinformation and to increase participants’ knowledge. Becausethe entire school community was involved in the socialactivities at step 3, a spreading effect is also expected,resulting in promotion of health education regarding infor-mation and attitudes regarding persons with CLP.

The questions that addressed definition (question 1),embryology (question 3), etiology (question 4), and treat-ment (question 8) of CLP showed the lowest rates of correctanswers before the YDP. After the program, the correctanswers for these questions increased�50% overall. Question3 showed one of the lowest increase rates after YDP. Thesefindings can be explained due to the difficulty of the assimi-lation of content related to embryologic information at themiddle school level, because this content is generally ad-dressed with students at the high school level in Brazil.15 Theresults for question 1 are noteworthy. This question showedthe lowest rate of correct answers before the YDP (9.8%). After

Fig. 2 Practical activity: group quiz.



the program, the rate of correct answers obtained was 97.6%,with nearly 100% of the participants showing that theyachieved a better understanding about CLP.

The YDP was developed within the school environmentinvolving direct and indirect participation of the school commu-nity. Including teachers and staff in health education initiativeslike the YDP can optimize efforts to build healthy environmentsand to change attitudes of the community regarding bullyingand discrimination.16 Although the objective of this project wasto measure the knowledge gained by the participants, it is theopinion of the authors that health promotion actions like theones proposed within the YDPs can lead to cooperation andcollaboration, easing the teaching of complex topics, such as CLP,while fostering supportive environments that promote inclusionof children with differences in the mainstream school system.This is exactly thescopeof theYDP.To theparticipants, becominga young doctor allows the students to take a role in healthpromotion and advocacy once they are trained to disseminatespecific knowledge in a given community.6

In this project, the young doctors acted as multipliers ofknowledge, transmitting information to other school stu-dents, teachers, staff, relatives, and their community throughsocial action. It is not possible tomeasure the exact number ofpeoplewho received the information. Even so, this project hasthe combined health and education elements that allow fordissemination of information needed to change behavior

regarding inclusion of persons withmalformations in society.At the same time, the experience helps to promote the idea ofa productive health chain.

Previous studies also revealed that the YDP can be effectivein promoting knowledge acquisition but also suggested that itis an important tool to generate behavioral changes, particu-larly related to attitudes of the participants toward socialinclusion of individuals with differences or disabilities.6,13

Question 12 approached the aspect of inclusion in this study.Before the YDP, 39% of the participants were unfavorable toinclusion initiatives, and after the YDP, only 7.3% participantsstill disapproved inclusion of people with differences relatedto CLP. The use of technology in educational programs such asthe cybertutor, for example, supports self-directed learning,allowing the participant to build knowledge before engagingin interactions within a group. When these educationalstrategies are combinedwith activities involving interactionsthat require the practical use of the information exchanged,the potential to increase knowledge and to promote behav-ioral change can be optimized and broadened.13

The YDP on CLP, as reported in this study, showed changesin the participant’s attitudes toward inclusion. Similar healtheducation projects should be encouraged, because they canmaximize dissemination of knowledge at the same time thatthey promote engagement of participants with their ownhealth and the overall health of their communities.5,6,17

Fig. 3 Practical activity: disseminating knowledge.


Teleducation about Cleft Lip and Palate Corrêa et al.110

Conclusion

In conclusion, the YDP on CLP was developed and appliedwith a group of students at amiddle school, providing a tool todisseminate information regarding CLP. An increase in theknowledge regarding the content was observed after partici-pation in the YDP, suggesting that this educational strategymay be an effective tool to multiply knowledge and tooptimize health promotion and advocacy.

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4 Gertel MCR, Maia SM. Speech therapist and school—consider-ations about the inclusive educational system: case report. RevCEFAC 2011;13(5):954–961

5 Wen CL, Silveira PSP, Azevedo RS, Böhm GM. Internet discussionlists as an educational tool. J Telemed Telecare 2000;6(5):302–304

6 Haney M, Silvestri S, Van Dillen C, Ralls G, Cohen E, Papa L. Acomparison of tele-education versus conventional lectures inwound care knowledge and skill acquisition. J Telemed Telecare2012;18(2):79–81

7 Soirefmann M, Boza JC, Comparin C, Cestari TF, Wen CL. Cybertu-tor: a teaching tool in dermatology. An Bras Dermatol 2010;85(3):400–402

8 Silva ASC, Rizzante FAP, Picolini MM, et al. Bauru School ofDentistry Tele-Health League: an educational strategy applied toresearch, teaching and extension among applications in tele-health. J Appl Oral Sci 2011;19(6):599–603

9 Blasca WQ, Maximino LP, Galdino DG, Campos K, Picolini MM.Novas tecnologias educacionais no ensino da audiologia. RevCEFAC 2010;12(6):1017–1024

10 Macea DD, Rondon S, Chaar LJ, Wen CL. Public health education foryoung students aided by technology. J Telemed Telecare 2009;15(3):159

11 Corrêa CC, Martins A, Pardo-Fanton CS, et al. Ações de teleducaçãointerativa em saúde vocal baseadas na dinâmica do projeto jovemdoutor. Distúrb Comum 2012;24(3):359–368

12 Vieira MMRM, Berretin-Felix G, Brasolotto AG. The Virtual ManProject’s CD-ROM “Voice Assessment: Speech-Language Pathologyand Audiology & Medicine,” Vol. 1. J Appl Oral Sci 2009;17(Suppl):43–49

13 Picolini MM, Blasca WQ, Richieri-Costa A, Maximino LP. Thedevelopment of a virtual learning environment in genetic syn-dromes. Rev CEFAC 2013;15(2):382–390

14 Chao LW, Silveira PSP, Böhm GM. Telemedicine and education inBrazil. J Telemed Telecare 1999;5(2):137–138

15 Freitas LAM, Barroso HFB, Rodrigues HG, Aversi-Ferreira TA.Construction of embryonic models with recycled material fordidactic using. Biosci J 2008;24(1):91–97

16 Vieira CM, Denari FE. Informative program about mental retarda-tion and inclusion: adjustment in social attitudes of childrenwithout disabilities. ABPEE 2012;18(2):265–282

17 Spinard ACP, Blasca WQ, De-Vitto LM. Genética e Fonoaudiologia:aprendizado baseado na teleducação. Pro Fono 2008;20:42–44

Table 1 Questionnaire results before and after the YDP

Questions Topics related to CLP Correct answersbefore YDP

Correct answersafter YDP

Improvement afterYDP

% n % n % n

1 Definition 9.8 4 97.6 40 88.8 36

2 Classification 85.4 35 100 41 14.6 6

3 Embryology 22 9 26.8 11 4.8 2

4 Etiology 26.8 11 65.9 27 39 16

5 Diagnostics 78 32 97.6 40 19.5 8

6 Alterations 68.3 28 100 41 31.7 13

7 Treatment 51.2 21 73.2 30 21.9 9

8 Treatment 34.1 14 80.5 33 46.3 19

9 Disorders 73.2 30 82.9 34 9.7 4

10 Treatment 56.1 23 65.9 27 9.7 4

11 Social attitudes 100 41 100 41 0 0

12 Social attitudes 61 25 92.7 38 31.7 13

13 Cleft features 95.1 39 100 41 4.8 2

Mean 58.5 24 83.3 34.1 24.8 10.1a

Minimum 9.2 4 26 11 16.8 7

Maximum 100 41 100 41 0 0

Abbreviations: CLP, cleft lip and palate; YDP, Young Doctor Project.aThe difference between the means before and after training was statistically significant according to the paired Student t test (t ¼ 6.687, p ¼ 0.001).



Parotid Incidentaloma Identified by PositronEmission/Computed Tomography: When toConsider Diagnoses Other than Warthin TumorCarolina Bothe1 Alejandro Fernandez2 Jacinto Garcia1 Montserrat Lopez1 Xavier León1

Miquel Quer1 Joan Lop1

1Department of Otorhinolaryngology, Hospital Sant Pau,Barcelona, Spain

2Department of Nuclear Medicine, Hospital de Sant Pau,Barcelona, Barcelona, Spain


Address for correspondence Carolina Bothe, MD, Department ofOtorhinolaryngology, Hospital Sant Pau, Avda. Sant Antoni Mª Claret,167, Barcelona 08025, Spain (e-mail: [email protected]).

Introduction

It is not unusual to find unexpected hypermetabolic foci as aresult of scanning using whole-body positron emission/com-puted tomography (PET/CT). Known as incidentalomas, suchfindings are not necessarily related to the tumor or diseasebeing studied. Parotid gland incidentalomas (PGIs) are defined

as new focal intraglandular deposits of radiotracer in patientswithout prior history of parotid disease.1 These deposits aremost commonly due to benign lesions such as Warthin tumor(WT),1–5 but they can also be caused by metastatic lymphnodes or malignant tumors.6

The parotid gland is the salivary gland that most commonlydevelops tumors,most of which are benign. After pleomorphic

Keywords

► parotid gland► adenolymphoma► PET scan► cigarette smoking

Abstract Introduction Parotid gland incidentalomas (PGIs) are unexpected hypermetabolic fociin the parotid region that can be found when scanning with whole-body positronemission/computed tomography (PET/CT). These deposits are most commonly due tobenign lesions such as Warthin tumor.Objective The aim of this study was to determine the prevalence of PGIs identified inPET/CT scans and to assess the role of smoking in their etiology.Methods We retrospectively reviewed all PET/CT scans performed at our center insearch of PGIs and identified smoking status and standardized uptake value (SUVmax) ineach case. We also analyzed the database of parotidectomies performed in ourdepartment in the previous 10 years and focused on the pathologic diagnosis andthe presence or absence of smoking in each case.Results Sixteen cases of PGIs were found in 4,250 PET/CT scans, accounting for 0.4%.The average SUVmax was 6.5 (range 2.8 to 16). Cytology was performed in five patients;it was benign in four cases and inconclusive in one case. Thirteen patients had a historyof smoking. Of the parotidectomies performed in our center with a diagnosis of Warthintumor, we identified a history of smoking in 93.8% of those patients.Conclusions The prevalence of PGIs on PET/CT was similar to that reported by otherauthors. Warthin tumor is frequently diagnosed among PGIs on PET/CT, and it has astrong relationship with smoking. We suggest that a diagnosis other than Warthintumor should be considered for PGIs in nonsmokers.

receivedAugust 22, 2014acceptedNovember 14, 2014published onlineDecember 29, 2014




112

adenoma, WT is the second most prevalent parotid tumor. Itpresents as a slow-growing mass in the tail of the gland, and isusually asymptomatic. It generally occurs between the sixthand seventh decades of life, predominantly in men. It has astrong association with smoking, which is therefore consid-ered a risk factor for its development.7–10 WT can be multi-centric or bilateral, and it rarely becomes malignant.

To our knowledge, only two previous studies have re-ported the prevalence of PGI, and both were performed inAsia.2,11 Furthermore, clinical features such as the role ofsmoking have not been considered in their etiology. Thepresent study aimed to determine the prevalence of parotidincidentalomas in 18F-fluorodeoxyglucose (18F-FDG) PET/CTand evaluate the presence of smoking in this group ofpatients. We also reviewed the database of parotidectomiesperformed in our hospital to assess the relationship betweenWT and smoking.


PatientsWe conducted a retrospective review of the reports of PET/CTperformed in 4,250 patients in our hospital betweenJune 2009 and February 2013. We identified 16 cases ofparotid focal uptake in patients without known disease ofthe parotid glands. The group consisted of 10 men and6 women, 48 to 88 years of age. In most cases, PET-CT wasindicated as an extension study of diagnosed neoplasms or toconfirm locoregional relapse after treatment. In two cases, itwas indicated to rule out giant cell arteritis.

We reviewed thehistory of smoking in the 16 cases: 13 hadsmoked or were current smokers. We identified additionalstudies performed on the parotid masses, finding that only 5patients underwent cytologic analysis by fine needle aspira-tion cytology (FNAC). In 8 of the 16 cases, the lesion waslocated in the right parotid gland, in 7 cases it was located onthe left, and in 1 case it was bilateral. Two patients had morethan one deposit within the same gland.

We analyzed the database of parotidectomies performedin our department in the previous 10 years and focused on thepathologic diagnosis and the presence or absence of smokingin each case.

18F-Fluorodeoxyglucose Positron Emission/ComputedTomographyCombined 18F-FDG whole-body PET/CT (Gemini TF, Philips,Amesterdam) with 64-slice CTwas performed. Standard patientpreparation included: 6-hour fasting, hydration, and serumglucose level of less than 150 mg/dL before tracer injection.Patientswere asked to rest quietly in supine position, and vesicalevacuation was done before the acquisition of images.

A dose of 3.7 MBq/kg of 18F-FDG was intravenously in-jected. Low-dose CT and PET from the base of the skull to theproximal thighs were performed, with an additional acquisi-tion of head and neck images. The total examination timewas�20 minutes. An abnormal PET-CT finding was defined as asignificant increased uptake, higher than that of the sur-rounding normal tissue.

Results

We found PGIs in 16 patients who underwent PET/CTbetween June 2009 and February 2013, corresponding to0.4% of patients undergoing PET/CT. The incidentalomas weremore frequent inmen (62.5%, 10 cases), and the average age atdiagnosis was 68 years.

The most common indication for the test was lung cancer(7 cases, corresponding to 43.75%), followed by breast cancer(3 cases), giant cell vasculitis (1 case each of Horton diseaseand Takayasu arteritis), and 1 case each of locally advancedoropharyngeal cancer, gallbladder carcinoma with liver me-tastasis, peritoneal carcinomatosis of gynecologic origin, andretroperitoneal lymphoma. (See ►Table 1.)

Five of the 16 patients underwent FNAC of the parotidmass. Three of five masses had an inflammatory componentwithout amalignant component, onewas considered amixedtumor suggestive of pleomorphic adenoma, and the otherwas inconclusive. Of the 16 patients, only the patient with thepleomorphic adenoma underwent surgery; a superficial pa-rotidectomywas performed and thehistologywas confirmed.In the remaining 11 cases, FNAC was not performed but themass was monitored by periodic clinical follow-up. Thirteenpatients (81.25%) had a history of smoking. One of the threenonsmokers was the patient with the pleomorphic adenoma.

The average standardized uptake value of parotid tumorswas 6.51, ranging from 2.87 to 16.1. The term used to describethe lesionswas hyperintense or hypermetabolic intraglandularnodule.

A review of the database of parotidectomies performed inour department revealed that 311 patients were surgicallytreated for parotid gland tumors in the previous 10 years and80 cases (25.7%) were identified as WT. This was the secondmost frequent tumor after pleomorphic adenoma. A historyof smoking was identified in 93.8% of patients with WT(75 cases). This percentage was high compared with theprevalence of smoking in pleomorphic adenomas, whichwas 38.8%. (See ►Table 2.)

Discussion

Our findings show that the prevalence of PGIs on PET/CT inour institution is similar to that reported in the two previousstudies in Asia. When analyzing the presence of cigarettesmoking in our patients with PGIs, we found it was a commonfactor in 81.25% of the patients.

In our study, FNACwas performed in five cases. Threewerenegative for malignancy without typing the lesion. One casewas a pleomorphic adenoma and the other had a nondiag-nostic smear. The remaining patients did not undergo addi-tional testing on the PGI due to the advanced stage of theprimary tumor, the absence of symptoms, or the unlikelihoodof metastasis based on clinical and tomographic features.

In the largest series published to date, Wang et al found 58PGIs in 19,333 patients, representing a prevalence of 0.3%.11

They obtained histologic confirmation in 51 patients, 41 withbenign lesions (mainly represented by pleomorphic adeno-mas and WT) and 10 with malignant tumors. In a smaller


Parotid Incidentaloma Identified by PET/CT Bothe et al. 113

sample, Horiuchi et al detected four cases of abnormal parotidFDG uptake in 1,872 PET scans, with an estimated prevalenceof 0.2%; all four cases were diagnosed with WT.2 Other seriesof cases also found WT as the main cause of PGI.1,3

As PGIs are likely benign lesions, metastasic diseaseshould not be suspected initially without histopathologicconfirmation, even in patients with known malignancyelsewhere.1,4,5 Nevertheless, these lesions warrant furtherradiologic and histopathologic correlations. Ultrasonogra-phy and FNAC are useful tools to characterize parotidmasses, avoiding erroneous staging of primary malignan-cies. The objective of cytologic diagnosis of parotid massesis the differential diagnosis between benign and malignantlesions.12 Complex histopathology and heterogeneity ofcellular patterns of salivary gland tumors explain whytyping of primary salivary malignancies by FNAC is achallenge. The accuracy of FNAC depends on the experienceof the clinician who performs the procedure and thepathologist who evaluates cytologic material.12

We found the prevalence of smoking in parotid tumorssurgically treated in our hospital was significantly higher inpatients with WT than in patients with other tumors(p < 0.001). More than 90% of patients with WT weresmokers, and cigarette use in patients with other benignand malignant neoplasms was lower, 38 and 55.4%, respec-tively. Several other authors have studied the relationbetween WT and smoking. According to their publications,between 80 and 94% of patients with WT smoke,7–9,13 andthe risk for development ofWT correlates with the level andduration of smoking.7,9 Sadetzki et al found a remarkablyhigh odds ratio (15.3) for developing WT in the presence ofsmoking.9

Conclusion

Based on the high prevalence of WT as PGI on PET/CT and thestrong relationshipwith smoking, we suggest diagnosis otherthan Warthin tumor should be considered for PGI in

Table1 Data of 16 patients with parotid incidentalomas detected by PET/CT

Patient Age (y) Sex Smoking SUVmax Cytology (FNAC)� PET/CT indication

1 64 Male Yes 4.3 and 3 Negative for malignancy Lung cancer

2 48 Female Yes 16.1 Negative for malignancy Horton disease

3 64 Female No 5.1 Not performed Breast cancer

4 66 Female No 4.55 Not performed Breast cancer

5 71 Male Yes 4.03 Inconclusive cytology Gallbladder cancer

6 88 Female Yes 3.3 Negative for malignancy Peritoneal carcinomatosis

7 54 Male Yes 5.5 Not performed Lung cancer


9 79 Female No 5.5 Pleomorphic adenoma Breast cancer

10 81 Male Yes 4 Not performed Retroperitoneal lymphoma


12 65 Male Yes 3.5 Not performed Oropharyngeal cancer

13 85 Male Yes 13.1 and 9.9 Not performed Lung cancer

14 58 Female Yes 6.8 Not performed Takayasu arteritis



Abbreviations: FNAC, fine needle aspiration cytology; PET/CT; positron emission/computed tomography; SUVmax, standardized uptake value.

Table 2 Data of parotidectomies performed in our department in the previous 10 years

Histologic diagnosis Number of cases (%) Smoking status (%)

Pleomorphic adenoma 134 (43.1%) 52 (38.8%)

Warthin tumor 80 (25.7%) 75 (93.8%)

Malignant tumors 56 (18%) 31 (55.4%)

Other diagnosis 41 (13.2%) 17 (41.5%)

Total 311 (100%) 175 (56.3%)


Parotid Incidentaloma Identified by PET/CT Bothe et al.114

nonsmokers. Further studies with histologic diagnosis andlarger samples are warranted to confirm our hypothesis.

References1 Lee SK, Rho BH, Won KS. Parotid incidentaloma identified by com-

bined 18F-fluorodeoxyglucosewhole-body positron emission tomog-raphy and computed tomography: findings at grayscale and powerDoppler ultrasonography and ultrasound-guided fine-needle aspira-tion biopsy or core-needle biopsy. Eur Radiol 2009;19(9):2268–2274

2 Horiuchi M, Yasuda S, Shohtsu A, Ide M. Four cases of Warthin’stumor of the parotid gland detected with FDG PET. Ann Nucl Med1998;12(1):47–50

3 Klijanienko J, Petras S, De Bosschere L, Paulmier B, Le Tourneau C,Rodriguez J. False-positive FDG PET/CTuptake inWarthin tumor inhead and neck oncological patients confirmed by a fine needleaspiration. Diagn Cytopathol 2012;40(3):282–284

4 Enomoto A, Nakahara H, Uchihashi T, Tsuji H, Hamada S. Fluo-rodeoxyglucose-positive Warthin tumor in a neck node mimick-ing metastasis in primary intraosseous left posterior mandibularcancer staging with positron emission tomography/computedtomography. J Oral Maxillofac Surg 2011;69(7):2052–2054

5 Iwai T, Baba J, Shibasaki M, et al. 18F-fluorodeoxyglucose-positiveWarthin tumor in a contralateral cervical lymph node mimicking

metastasis in tongue cancer staging with PET/CT. J Craniofac Surg2012;23(5):e507–e509

6 Coronado Poggio M, Couto Caro RM, Rodado Marina S, MartínCurto LM. 18F-FDG PET/TAC Semiology. Rev Esp Med Nucl 2008;27(4):284–304, quiz 305–306

7 Peter Klussmann J, Wittekindt C, Florian Preuss S, Al Attab A,Schroeder U, Guntinas-Lichius O. High risk for bilateral Warthintumor in heavy smokers—review of 185 cases. Acta Otolaryngol2006;126(11):1213–1217

8 Pinkston JA, Cole P. Cigarette smoking and Warthin’s tumor. Am JEpidemiol 1996;144(2):183–187

9 Sadetzki S, Oberman B, Mandelzweig L, et al. Smoking and risk ofparotid gland tumors: a nationwide case-control study. Cancer2008;112(9):1974–1982

10 Reddy VM, Thangarajah T, Castellanos-Arango F, Panarese A.Conservative management of Warthin tumour. J OtolaryngolHead Neck Surg 2008;37(5):744–749

11 Wang HC, Zuo CT, Hua FC, et al. Efficacy of conventional whole-body 18 F-FDG PET/CT in the incidental findings of parotidmasses.Ann Nucl Med 2010;24(8):571–577

12 Zbären P, Schär C, Hotz MA, Loosli H. Value of fine-needle aspira-tion cytology of parotid gland masses. Laryngoscope 2001;111(11Pt 1):1989–1992

13 Chedid HM, Rapoport A, Aikawa KF, Menezes AdosS, Curioni OA.Warthin’s tumor of the parotid gland: study of 70 cases. Rev ColBras Cir 2011;38(2):90–94


Parotid Incidentaloma Identified by PET/CT Bothe et al. 115

Preoperative Imaging Modalities to Predict theRisk of Regional Nodal Recurrence in Well-Differentiated Thyroid CancersMohammed K. AlNoury1 Saad M. Almuhayawi2 Khalid B. Alghamdi2 Khaled I. Al-Noury2

1Department of Otolaryngology Head and Neck Surgery, KingAbdulaziz University, Jeddah, Saudi Arabia

2Department of Otolaryngology, King Abdulaziz University, Jeddah,Saudi Arabia


Address for correspondence Mohammed K. AlNoury, MD,Department of Otolaryngology Head and Neck Surgery, King AbdulazizUniversity, P.O. Box 35135, Jeddah–21488, Saudi Arabia(e-mail: [email protected]).

Introduction

The overall increase in the incidence of thyroid cancer overthe last 2 decades can largely be attributed to increases in theincidence of papillary thyroid cancer (PTC). In 2010, there

were an estimated 44,670 new cases of thyroid cancer and1,690 deaths in the United States.1 PTC and its follicularvariant account for 80 to 90% of all primary thyroid cancers.Furthermore, follicular thyroid cancer and Hurthle cell can-cers comprise 5 to 10%, medullary thyroid cancer comprises

Keywords

► thyroid neoplasms► recurrence► ultrasonography► computed

tomography► X-ray

Abstract Introduction Thyroid cancer incidence has increased in the previous 2 decades.Preoperative identification of lymph node metastasis is a suggested risk factorassociated with recurrence following thyroidectomy.Objectives We aimed to evaluate the accuracy of preoperative radiologic investiga-tions of nodal status in determining the postoperative risk of regional nodal recurrencein cases of well-differentiated thyroid cancer.Methods This is a case series. We retrospectively reviewed data, including preopera-tive ultrasonography and/or computed tomography results, on patients who underwenttotal thyroidectomy for thyroid cancer at our hospital between 2006 and 2012.Prognostic factors for predicting recurrence, including age, sex, tumor diameter, andnodal diameter, were evaluated.Results Total thyroidectomy was performed on 24 male and 74 female patients(median age, 43 years). The median follow-up time was 21 months. Sixty-eight patientshad papillary thyroid cancer, and 30 had follicular cancer. Nodal recurrence was evidentin 30% of patients, and 4% of patients died. Identification of lymph node involvementduring preoperative radiologic investigations was strongly prognostic for recurrence:35.3% of patients with positive preoperative ultrasonography findings and 62.5% ofthose with positive preoperative computed tomography findings had recurrence(p ¼ 0.01).Conclusions Preoperative identification of lymph node metastasis on radiologicstudies was correlated with an increased risk of regional nodal recurrence in well-differentiated thyroid cancer. Computed tomography was superior to ultrasonographyin detecting metastatic nodal involvement preoperatively and is therefore recom-mended for preoperative assessment and postoperative follow-up.





116

5%, and anaplastic cancer comprises less than 1% of all thyroidmalignancies.2,3 Differentiated thyroid cancer (DTC) can oc-cur at any age but the median age at diagnosis is 49 years, andthe prevalence inwomen is 3 times greater than that inmen.4

Cervical recurrence of PTC following thyroidectomyoccursprimarily as regional lymph node (LN) metastasis, whichoccurs in up to 20% of patients with low-risk disease (men� 40 years old; women � 50 years old) and 59% of patientswith high-risk disease (older patients).3,5–7 This type oflocoregional cervical LN recurs within the first 10 yearsfollowing an initial diagnosis in 15 to 30% of patients.6,8

Current surveillance strategies to identify locoregional ornodal recurrence primarily rely on serial serum thyroglobulinmeasurements assessed in combination with cervical ultra-sonography (US) and image-guided fine needle aspirationcytology (FNAC) of suspicious lesions.9–11 Recently, certainnumber of antibodies have been developed against antigen toimprove morphologic diagnostic performances.12

Several tumor-staging systems have been developed forDTC in an attempt to include factors with prognostic value toguide the appropriate intensity of treatment and surveillance.The most relevant factors include patient age, tumor size andextent, locoregional nodal involvement, and distant metasta-ses,4 but there are few studies discussing preoperative factorsthat could predict nodal recurrence. Factors that may de-crease recurrence rates include a more comprehensive sur-gery, better tumor definition afforded by more sensitive UStechniques, and the use of routine cervical lymphadenectomyto remove LNs that could cause recurrence.13 Innovations inserology, histopathology, immunochemistry, and diagnosisthrough radiologic investigations provide us with betterunderstanding to plan the management and follow-up ofwell-differentiated thyroid cancer.

The objective of the current study was to evaluate theaccuracy of preoperative radiologic investigations of nodalstatus in determining the postoperative risk of locoregionalnodal recurrence in patients with DTC.


Patient ScreeningFollowing the hospital ethics committee approval, the pro-spectively maintained database from our tertiary hospitalwas reviewed. Ninety-eight patients met the inclusion crite-ria, which were as follows: (1) well-differentiated thyroidcancer, (2) preoperative radiologic investigations performedin our hospital, (3) patient operative report, and (4) aminimum follow-up of 12 months. Demographic data, pre-operative and postoperative US and/or computed tomogra-phy (CT) scans, FNAC findings, and surgical pathology wereevaluated. The operative and pathology reports were re-viewed, and all patients were staged according to the currentAmerican Joint Committee on Cancer (AJCC) staging systemaccording to the documented histopathologic findings.14 Theclinical course was determined, and all patients who pre-sented with recurrent disease were identified. Follow-upevaluation included a physical examination, a neck US ex-amination, measurement of serum thyroglobulin levels, and

the selective use of neck CT scanning, total body radioactiveiodine (RAI) scanning, FNAC, and histopathology reports.Patients were considered to have cervical recurrence ifthey had any of the following: nodal disease on clinicalexamination; detection of recurrence by FNAC; or an intervalincrease in activity on serial RAI imaging that promptedtreatment with RAI,13 and positive FNAC was considered asmost definitive end point of nodal recurrence.

Diagnostic Imaging ModalitiesUS of the soft tissue of the neck was performed using a high-resolution scanner. The preoperative and postoperative sta-tus of central and lateral neck compartments was determinedaccording to the sonographic appearance of the thyroid, adescription of the lesion, and the appearance of the LNs.Sonographic features suggestive of abnormal metastatic LNsinclude loss of the fatty hilus, a rounded rather than ovalshape, hypoechogenicity, cystic change, calcifications, andperipheral vascularity. Some patients underwent contrast-enhanced CT with a multidetector scanner with a recon-structed slice thickness of 3 mm for axial and coronal images.A 90-mL dose of iodinated contrast medium was adminis-tered intravenously at a rate of 3 mL/s using an automatedinjector. A 3-mL/sflush of normal saline solutionwas injectedimmediately after administration of the contrast medium toreduce artifacts induced in the subclavian vein. The scan delaywas 40 to 60 seconds. Due to different treating surgical teamsand collecting of data retrospectively, different patients re-ceived different radiologic investigation. Therefore, we divid-ed the patients into different groups: patients who receivedUS pre- and postoperatively and patients who received CTscan pre- and postoperatively.

Operative ProceduresTotal thyroidectomy was performed in cases diagnosed withDTC by FNAC. In most cases, a prophylactic central nodedissection (CND) was performed in patients with PTC andclinical stage N0 neck. A therapeutic CND and/or lateral neckdissectionwas performed in patients with evidence of centraland/or lateral neck LN metastasis at the time of the surgery.After surgery, all patients underwent thyroid-stimulatinghormone suppression treatment with oral thyroxin for6 weeks, followed by RAI therapy to improve outcome andtumor control.

Statistical AnalysesAll statistical tests were performed using SPSS version 16(IBM). Univariate statistical analysis and cross-tabulationwere performed to determine the significance of each factorin predicting cervical recurrence in DTC. The chi-square testwas used to calculate p values, and p < 0.05 was consideredstatistically significant.

Results

Between January 2006 and December 2012, 98 patientsunderwent total thyroidectomy for a diagnosis of well-differentiated thyroid cancer. Patient demographics are


Preoperative Imaging to Predict Regional Nodal Recurrence AlNoury et al. 117

shown in ►Table 1. The median age at diagnosis was43 years. The median follow-up was 21 months from thetime of the index operation. Cervical recurrence was de-fined as metastatic involvement of any cervical LN devel-oping �6 months after surgery. Cases of cervicalrecurrences were mainly isolated LNs. Of the 98 patients,71 (72%) patients underwent preoperative US, and 35 (36%)underwent preoperative CT; 8 patients underwent bothstudies. Fifty-six patients had postoperative US, and 42 hadpostoperative CT scans.

An analysis of factors predicting local recurrence is shownin ►Table 2. Positive LN recurrence was higher in mencomparedwithwomen, although the difference did not reachstatistical significance (p ¼ 0.784). In addition, age, tumorsize, and LN size were not significant risk factors for localrecurrence (p ¼ 0.608, p ¼ 0.385, p ¼ 0.875, respectively).

Positive LN status on US was not correlated with theprediction of recurrence. In contrast, positive or negativeLN status as detected by CTwas significantly correlated withpostoperative locoregional recurrence (p ¼ 0.01; ►Table 3).The US and CT analyses demonstrated similar efficacy atdetecting thyroid nodules, but differed significantly in theirability to accurately evaluate cervical LN involvement inrecurrence, with CT outperforming US (►Table 4).

Discussion

PTC is themost common endocrine neoplasiawith a tendencyfor local and regional metastasis.15 The prognostic impact of

cervical LN involvement has been investigated in manystudies, but remains a controversial issue.16–18 Initial studiessuggested that the presence of nodal metastasis had no effectin overall or disease-free survival, but this notion has beenrecently refuted.19 Patients with PTC with cervical LN metas-tasis at initial presentation aremore susceptible to recurrencethan thosewithout LN involvement.16–18 The preoperative LNmetastasis rate in our series was 64%, which was higher thanthe published 23%.19 The recurrence rate in our series (30%)was higher than that previously reported in published studies(10 to 20%),18,20 which can mostly likely be attributed todifferences in surgical techniques and treatments betweendifferent surgeons. However, the surgical management of

Table 2 Risk factors correlated with recurrence

Risk factors No. ofpatients withrecurrence (%)

p Value

Sex

Male (n ¼ 18) 8 (44.4) 0.784

Female (n ¼ 56) 22 (39.3)

Age (y)

�60 (n ¼ 58) 22 (37.9) 0.538

>60 (n ¼ 16) 8 (50)

Tumor size (cm)

�2 (n ¼ 38) 18 (47.4) 0.385

>2 (n ¼ 36) 12 (33.3)

Lymph node size (cm)

No nodal involvement(n ¼ 26)

10 (38.5) 0.875

�2 (n ¼ 36) 14 (38.9)

>2 (n ¼ 12) 6 (50)

aThe chi-square test was used to measure p values (p < 0.05 wasconsidered significant by Fisher’s exact tests).

Table 1 Patient demographics

Characteristics Number ofpatients (%)

Sex

Male 24 (24.5)

Female 74 (75.5)

Age (y)

<60 76 (77.6)

�60 22 (22.4)

Histopathology

Papillary thyroid cancer 68 (70)

Follicular cancer 30 (30)

Tumor size (cm)

�2 45 (46)

>2 53 (54)

Lymph node size (cm)

No nodal involvement 35 (36)

�2 47(48)

>2 16 (16)

Recurrence 29 (30)

Death 4 (4)

Table 3 Correlation of the preoperative lymph node status withrecurrence risk

Preoperative radiologicinvestigation

No. of patients withrecurrence (%)

p Value

Negative LN status onUS (n ¼ 20)

6 (30) 0.778

Positive LN status onUS (n ¼ 34)

12 (35.3)

Negative LN status onCT (n ¼ 15)

1 (6.67) 0.01

Positive LN status onCT (n ¼ 14)

10 (62.5)

Abbreviations: CT, computed tomography; LN, lymph node; US,ultrasonography.aThe chi-square test was used to measure p values (p < 0.05 wasconsidered significant by Fisher’s exact tests).


Preoperative Imaging to Predict Regional Nodal Recurrence AlNoury et al.118

malignant thyroid nodule was based on FNAC and frozensection evaluation, with the FNAC being more sensitive andcost-effective.21

In this study,we evaluated age, sex, tumor size, LN size, andUS and CT findings preoperatively to determine the relation-ship of these factors with nodal recurrence. Univariate analy-sis has been used, due to small sample, although it wasinsufficient in comparison with multivariate analysis. Ageand sex are considered prognostic factors (2010 AJCC stagingsystem),17,22 but these factors were not associated withcervical recurrence in our study.

In addition, tumor size and LN size were not predictive ofcervical recurrence. Preoperative LNmetastasis was associat-ed with locoregional recurrence, as has been described forother studies.18,23 In 560 Japanese patients with thyroidcancer who underwent total thyroidectomy, those with US-detectable metastasis had a significantly worse relapse-freesurvival than those with negative US findings.24 Hay et al25

and Spires et al26 showed that LN metastasis was related to ahigher recurrence rate but did not adversely influencesurvival.

Our study and those mentioned previously would indicatethat positive preoperative US findings for LNmetastasis couldindicate a more aggressive disease course and strongly pre-dict the need for additional surgery in the future, whereasnegative preoperative US LN findings indicate a lower riskwith patients less likely to need future surgery.26 However,most importantly, we determined that preoperative detec-tion of LN metastases with CT-based detection was signifi-cantly more specific, sensitive, and accurate than US-baseddetection; all patients who had negative preoperative CT LNmetastasis findings remained recurrence-free, whereas 30%of the patients with negative US LNmetastases findings wenton to develop locoregional recurrence. This strongly suggeststhat preoperative CT detection of LN metastases is a bettermodality for predicting locoregional recurrence in patientswith DTC. On the other hand, the sensitivity and specificity ofpositive LN status on US in this study were less than those inother published studies.15,27,28

Conclusions

According to this study, patients with thyroid cancer withpositive radiologic investigations (either US or CT) for LNmetastasis have an increased risk of regional node recur-rence after total thyroidectomy. CT was superior to US atdetecting metastatic LN involvement, and, therefore, on thebasis of our data, we suggest that CT should be furtherstudied and considered as a more suitable alternative to US

for preoperative evaluation and postoperative follow-upinvestigations.

References1 Davies L, Welch HG. Increasing incidence of thyroid cancer in the

United States, 1973–2002. JAMA 2006;295(18):2164–21672 Gilliland FD, Hunt WC, Morris DM, Key CR. Prognostic factors for

thyroid carcinoma. A population-based study of 15,698 cases fromthe Surveillance, Epidemiology and End Results (SEER) program1973–1991. Cancer 1997;79(3):564–573

3 Mazzaferri EL, Kloos RT. Clinical review128: current approaches toprimary therapy for papillary and follicular thyroid cancer. J ClinEndocrinol Metab 2001;86(4):1447–1463

4 Johnson NA, LeBeau SO, Tublin ME. Imaging surveillance ofdifferentiated thyroid cancer. Radiol Clin North Am 2011;49(3):473–487, vi

5 Cady B, Rossi R. An expanded view of risk-group definition indifferentiated thyroid carcinoma. Surgery 1988;104(6):947–953

6 Hay ID, Thompson GB, Grant CS, et al. Papillary thyroid carcinomamanaged at the Mayo Clinic during six decades (1940–1999):temporal trends in initial therapy and long-term outcome in 2444consecutively treated patients. World J Surg 2002;26(8):879–885

7 Simon D, Goretzki PE, Witte J, Röher HD. Incidence of regionalrecurrence guiding radicality in differentiated thyroid carcinoma.World J Surg 1996;20(7):860–866, discussion 866

8 Mazzaferri EL, Jhiang SM. Long-term impact of initial surgical andmedical therapy on papillary and follicular thyroid cancer. Am JMed 1994;97(5):418–428.Erratum in: Am J Med 1995;98:215

9 Pacini F, Molinaro E, Castagna MG, et al. Recombinant humanthyrotropin-stimulated serum thyroglobulin combined with neckultrasonography has the highest sensitivity in monitoring differ-entiated thyroid carcinoma. J Clin Endocrinol Metab 2003;88(8):3668–3673

10 Schlumberger M, Berg G, Cohen O, et al. Follow-up of low-riskpatients with differentiated thyroid carcinoma: a European per-spective. Eur J Endocrinol 2004;150(2):105–112

11 Torlontano M, Crocetti U, Augello G, et al. Comparative evaluationof recombinant human thyrotropin-stimulated thyroglobulin lev-els, 131I whole-body scintigraphy, and neck ultrasonography inthe follow-up of patients with papillary thyroid microcarcinomawho have not undergone radioiodine therapy. J Clin EndocrinolMetab 2006;91(1):60–63

12 Saggiorato E, Aversa S, Deandreis D, et al. Galectin-3: presurgicalmarker of thyroid follicular epithelial cell-derived carcinomas.J Endocrinol Invest 2004;27(4):311–317

13 Marshall CL, Lee JE, Xing Y, et al. Routine pre-operative ultraso-nography for papillary thyroid cancer: effects on cervical recur-rence. Surgery 2009;146(6):1063–1072

14 Greene FL, Page DL, Fleming ID, et al , eds. American JointCommittee on Cancer; American Cancer Society. AJCC CancerStaging Manual. 6th ed. New York, NY: Springer-Verlag; 2002

15 González HE, Cruz F, O’Brien A, et al. Impact of preoperativeultrasonographic staging of the neck in papillary thyroid carci-noma. Arch Otolaryngol Head Neck Surg 2007;133(12):1258–1262

Table 4 A comparison of the prognostic accuracy of radiologic modalities for predicting recurrence

Modality Sensitivity (%) Specificity (%) Accuracy (%) PPV (%) NPV (%)

US 66.67 77.78 74 60 82.35

CT 80 90 86.6 80 90

Abbreviations: CT, computed tomography; NPV, negative predictive value; PPV, positive predictive value; US, ultrasonography.


Preoperative Imaging to Predict Regional Nodal Recurrence AlNoury et al. 119

16 Shaha AR. Prognostic factors in papillary thyroid carcinoma andimplications of large nodalmetastasis. Surgery 2004;135(2):237–239

17 Shaha A. Treatment of thyroid cancer based on risk groups. J SurgOncol 2006;94(8):683–691

18 Pacini F, Schlumberger M, Dralle H, Ilisea R, Smith Y, Viersinga V.[European consensus on the management of patients with differ-entiated carcinoma of the thyroid from follicular epithelium].Vestn Khir Im I I Grek 2008;167(1):52–56[Article in Russian]

19 Londero SC, Krogdahl A, Bastholt L, et al; Danish Thyroid CancerGroup. Papillary thyroid microcarcinoma in Denmark 1996–2008:a national study of epidemiology and clinical significance. Thyroid2013;23(9):1159–1164

20 Dionigi G, Dionigi R, Bartalena L, Boni L, Rovera F, Villa F. Surgery oflymph nodes in papillary thyroid cancer. Expert Rev AnticancerTher 2006;6(9):1217–1229

21 Caraci P, Aversa S, Mussa A, Pancani G, Ondolo C, Conticello S. Roleof fine-needle aspiration biopsy and frozen-section evaluation inthe surgical management of thyroid nodules. Br J Surg 2002;89(6):797–801

22 Schlumberger M, Pacini F, Wiersinga WM, et al. Follow-up andmanagement of differentiated thyroid carcinoma: a Europeanperspective in clinical practice. Eur J Endocrinol 2004;151(5):539–548

23 Wada N, Masudo K, Nakayama H, et al. Clinical outcomes in olderor younger patients with papillary thyroid carcinoma: impact oflymphadenopathy and patient age. Eur J Surg Oncol 2008;34(2):202–207

24 Ito Y, Tomoda C, Uruno T, et al. Ultrasonographically and anato-mopathologically detectable node metastases in the lateral com-partment as indicators of worse relapse-free survival in patientswith papillary thyroid carcinoma. World J Surg 2005;29(7):917–920

25 Hay ID, Grant CS, van Heerden JA, Goellner JR, Ebersold JR,Bergstralh EJ. Papillary thyroid microcarcinoma: a study of 535cases observed in a 50-year period. Surgery 1992;112(6):1139–1146, discussion 1146–1147

26 Spires JR, Robbins KT, Luna MA, Byers RM. Metastatic papillarycarcinoma of the thyroid: the significance of extranodal extension.Head Neck 1989;11(3):242–246

27 Poehls JL, Chen H, Sippel RS. Preoperative ultrasonography find-ings predict the need for repeated surgery in papillary thyroidcancer. Endocr Pract 2012;18(3):403–409

28 Roh JL, Park JY, Kim JM, Song CJ. Use of preoperativeultrasonography as guidance for neck dissection in patientswith papillary thyroid carcinoma. J Surg Oncol 2009;99(1):28–31


Preoperative Imaging to Predict Regional Nodal Recurrence AlNoury et al.120

Foreign Bodies in the Ear, Nose and Throat: AnExperience in a Tertiary Care Hospital in CentralNepalRamesh Parajuli1

1Department of ENT, Head and Neck Surgery, Chitwan MedicalCollege Teaching Hospital, Bharatpur, Chitwan, Nepal


Address for correspondence Ramesh Parajuli, Assistant Professor,Department of ENT and Head and Neck Surgery, Chitwan MedicalCollege, Chitwan, Nepal, Chitwan 00977, Nepal(e-mail: [email protected]).

Introduction

A foreign body (FB) is any object in a region it is not meant tobe,where it can causeharmby itsmere presence if immediatemedical attention is not sought.1 It can be found in the ear,nose, and throat (ENT) region. FBmay be classified as animate(living) and inanimate (nonliving). The inanimate FBs canfurther be classified as organic or inorganic and hygroscopic(hydrophilic) or nonhygroscopic (hydrophobic).2 The pres-ence of FBs in the ENT region is one of the most common

causes of otolaryngologic emergencies. FBs can be introducedspontaneously or accidently in both adults and children.Generally, FBs are more common in younger children; thismay be due to various factors such as curiosity to exploreorifices, imitation, boredom, playing, mental retardation,insanity, and attention deficit hyperactivity disorder, alongwith availability of the objects and absence of watchfulcaregivers.3 The aim of this study is to analyze FBs in termsof type, site, age, and gender distribution and method ofremoval.

Keywords

► foreign body► ear► nose► throat► endoscopy

Abstract Introduction A foreign body (FB) is an object or substance foreign to the locationwhere it is found. FBs in the ear, nose, and throat are a common problem frequentlyencountered in both children and adults.Objective To analyze FBs in terms of type, site, age, and gender distribution andmethod of removal.Methods A retrospective study was performed in a tertiary care hospital in the centralpart of Nepal. The study period was from June 2013 to May 2014. The information wasobtained from hospital record books.Results A total of 134 patients had FBs in the ear, nose, or throat; 94 weremales and 40were females. Of the 134 patients, 70 (52.23%) had FB in the ear, 28 (20.89%) in thenose, and 36 (26.86%) in the throat. The FB was animate (living) in 28 (40%) patientswith FB in the ear and 1 (3.5%) patient with FB in the nose, but the FB was inanimate(nonliving) in any patient with FB in the throat, in 42 (60%) patients with FB in the ear FB,and in 27 (96.4%) patients with FB of the nose. The FB was removed with or without localanaesthesia (LA) in 98 (73.13%) patients, and only 36 patients (26.86%) required generalanaesthesia (GA). The most common age group affected was <10 years.Conclusion FBs in the ear and nose were found more frequently in children, and thethroat was the most common site of FB in adults and elderly people. Most of the FBs canbe easily removed in emergency room or outpatient department.




THIEME

Original Research 121


A retrospective study was performed in the Department ofENT, Head and Neck Surgery in a tertiary care hospital in thecentral part of Nepal. The study population includes thenumber of patients with ENT FB lodgment who presentedin the Outpatient Department (OPD) or in the emergencyroom (ER) during the 1-year study period (June 2013 toMay 2014). The data were obtained from the hospital recordbooks. Anterior rhinoscopy and otoscopy examination wereperformed to diagnose FB of the nose and ear, respectively.Rigid or flexible nasal endoscopic examination was alsoperformed in suspected cases of FB in the nasal cavity thatwas not visualizedwith anterior rhinoscopy. Similarly, exam-ination under microscope was an additional method fordiagnosis as well as useful for removal of FB of the ear.Instruments such as Jobson Horne probe, FB hook, Tilleyforceps, and crocodile forceps were used in FB removalfrom the nose and ear. In addition to the previously men-tioned instruments, syringing and suctioning were othermethods for FB ear removal. Plain X-ray of the neck wasdone in patients with a history of FB ingestion. Flexiblenasopharyngolaryngoscopy and flexible upper gastrointesti-nal endoscopy were done in cases where the FB was notvisible in X-ray to rule out presence of an FB or to determineits site of impaction. This was followed by removal of the FBfrom the oropharynx/hypopharynx and esophagus with di-rect laryngoscopy or rigid esophagoscopy, respectively.

Results

During the study period, 134 patients visited this hospitalwithFB in the ENT area; 94 weremales and 40 were females. Of the134 patients, 70 (52.23%) had FB in the ear, 28 (20.89%) in thenose, and 36 (26.86%) in the throat. The FB was removed withor without local anaesthesia (LA) in 98 (73.13%) patients, andonly 36 patients (26.86%) required general anaesthesia (GA).

Foreign Bodies in the EarA total of 70 patients presented to the hospital with FB in theear. Of these 70 patients, 28 (40%) harbored animate (living)FBs. These were 22 cases of ticks, 4 cases of cockroach, 1 caseeach of ant and aural myiasis. The rest (42; 60%) had inani-mate (nonliving) FB in their ears. Of these 42 cases ofnonliving FB, 15 were hygroscopic FB in the form of grams,peanuts, bean seed, and rice grain; the remaining 27 caseswere of nonhygroscopic FB in the form of cotton, paper,eraser, broken matchstick/cotton bud, foam, and beads.

Of the total 70 patients, 32 (45.71%) were children <10years of age.

Of 70 cases of FB in the ear, 66 were removed in the OPDand ER with or without LA and only 4 required examinationunder microscope under GA for removal of the FB; all of themwere younger than 10 years of age.

Foreign Bodies in the NoseTwenty-eight patients had FB lodged in the nose. Twenty-seven patients (96.42%), all of whomwere children<10 years

of age, had nonliving FB and only 1 patient had living FB (i.e.,maggots), an adult patient with fungating growth due tocarcinoma of maxilla. Of 27 cases with nonliving FBs, 10patients had hygroscopic FB such as bean, peanut, corn, andgrams, and 17 patients had nonhygroscopic FB such as eraser,paper, sponge, and plastic and metallic objects.

Of the total 28 patients with an FB in the nose, 27 (96.42%)were of the age group < 10 years. Most of the FBs wereremoved in the OPD and ER with the application of topicalnasal decongestant. Only 2 patients required removal of theFB under GA.

Foreign Bodies in the ThroatA total of 36 patients presented with the complaint ofingestion of FB. The most common type of FB was meatbone/bolus in the form of chicken, mutton, or buffalo meatand the most common site of the impaction was cricophar-yngeal junction in 21 patients (58.3%). The other sites of FBimpaction were oral cavity, oropharynx, hypopharynx, andthoracic esophagus.

All the ingested FBwere inanimate, with 26 (72.22%) beingorganic and 10 (27.77%) being inorganic. Organic FBs weremeat bolus and bone (fish, chicken, mutton, and buffalomeat). The inorganic FBs included denture, coin, and plasticand metallic objects.

Age of 60 or more years was the most common groupinvolved with FB in the throat, with 10 patients presentingwith FB impaction. Among all the patients who ingested anFB, 30 patients (83.33%) required GAunder GA for FB removal.

Method of FB RemovalRemoval of FB is not always easy. It requires proper instru-ments and skill. In our study, most of the nasal and aural FBswere removed in the ERor OPDwith or without LA. Out of 134FBs in the ENT, 36 (26.47%) required GA for FB removal andrest (98; 73.13%) were removed with or without LA.

Discussion

Adults and older children usually give a history of FB lodg-ment in ENT. But younger children are brought to the clinic byanxious parents or relatives. FBs may vary widely in shape,size, and composition, and the symptoms may range fromasymptomatic to acute life threatening condition.

In our study, the most common age group affected wasage < 10 years, similar to results found in many other stud-ies.4–8 This may be due to the tendency of young children tolodge objects into the natural orifices of body, accidentally orintentionally.

The ear was the most common site for FBs in youngchildren, who not only insert objects in their ears but alsointo the ears of their siblings and friends. Common ear FBsinclude cotton wool, bean, bead, paper/plastic, eraser, insect,paddy seed, and popcorn kernel. Patients usually presentwith earache, aural fullness, or ear discharge. Occasionally itmay be asymptomatic and found incidentally during routineotoscopic examination. A high incidence of living FBs (i.e.,ticks) in our study is explained by the fact that the people in


Foreign Bodies in the Ear, Nose, and Throat Parajuli122

the villages of Chitwan go to the jungle to collect fodder andgraze cattle. Many people get ticks in the ear when they go forjungle safari, as Chitwan is a famous tourist destination in ourcountry and attracts both domestic and international tourists.Examination under a microscope helps to confirm the pres-ence of FB in the ear and aids in its removal under intravenoussedation/GA to minimize trauma to the tympanic membraneand external auditory canal. It is useful especially in childrenwho are not cooperative to allow proper otoscopic examina-tion when there is associated otitis externa. Ear syringing ledto successful removal of most of the nonhygroscopic FBs.Negative pressure suctioning can be useful especially whenthere is aural or nasal discharge along with the FB.

Our study showed that there is predominance of FBs in thenose in younger children, which was seen in many otherstudies.1,3,7 Unilateral, foul-smelling, purulent nasal dischargein children must be regarded as due to FB until provedotherwise. With growth and cognitive development, the in-troduction of FB in the nostrils diminishes significantly, whichis found only in patients with psychiatric disorders. FB in thenose or ear is usually unilateral, although it can be bilateral, asin one of our patient with FB grams in both nasal cavities andanother case of FB erasers in both the external auditory canals.Sometimes there can be multiple ear or nose FBs as well.

FB ingestion is a common problem. The most frequentlyswallowed FBs in children include coins and metallic FBs(parts of playing objects), and meat bones (chicken bone/fishbone/mutton/buffalomeat) are common in adults and elderlypatients.8,9 In our study, meat bone/bolus was the mostcommon FB found inside the throat, and the most commonsite of lodgment was the cricopharyngeal region. Such pa-tients come in clusters, especially during religious festivalslike Dashain and Tihar. Heavy consumption of alcohol andeating meat simultaneously, especially during festivals, alongwith poor mastication may be the cause for meat bone/bolusimpaction in adults. In elderly, edentulous patients, defectiveperistalsis due to age-related neuromuscular incoordinationand poor masticating habits are the predisposing factors forthe cause of impaction of meat bone/bolus in the esophagus.Moreover, in elderly people there are commonly other un-derlying pathologies that cause narrowing of the digestivetract. Coin was the most common FB in the throat in childrenin our study, which is similar to other studies7,9; this may bedue to fact that the coins are often handed toyounger childrenand they accidentally swallow because of their tendency totake things into the mouth, inadequate control of deglutition,and shouting or crying while playing or eating.

Plain X-ray of soft tissue in the neck is a cost-effectiveradiologic examination method useful in the evaluation of FBin the throat. We advise X-rays in patients with history of FBingestion. Direct laryngoscopy was occasionally useful in theevaluation and removal of FB in the oropharynx and hypo-pharynx. FBs from the digestive tract are usually removed byrigid esophagoscopy. But flexible upper gastrointestinal en-doscopy is useful especially in the case of ingestion ofradiolucent FBs. Moreover, it helps to detect the site ofimpaction especially in patients with cervical spondylosiswhere neck extension is not possible and to remove the FBs,or to push the FB into the stomach (e.g., in cases of impactedmeat bolus at the distal part of esophagus).

Conclusion

FBs in the ear and nose were found more frequently inchildren, and the throat was the most common site of FBsin adults and elderly people. Most of the nasal and aural FBscan be easily removed in the ER or OPD. Parents/caretakershould not allow children to play with coins or other smallobjects to prevent the risk of FB ingestion or insertion.

References1 Sarkar S, Roychoudhury A, Roychaudhuri BK. Foreign bodies in ENT

in a teaching hospital in Eastern India. Indian J Otolaryngol HeadNeck Surg 2010;62(2):118–120

2 Carney AS, Patel N, Clarke R. Foreign bodies in the ear and theaerodigestive tract in children. In: Scott-Brown’s Otorhinolaryn-gology, Head and Neck Surgery, 7th ed. London, UK: EdwardArnold; 2008:1184–1193

3 Shrestha I, Shrestha BL, Amatya RCM. Analysis of ear, nose andthroat foreign bodies in Dhulikhel hospital. KathmanduUnivMed J(KUMJ) 2012;10(38):4–8

4 Banerjee S. Concept of foreign body—its past and present. Indian JOtolaryngol Head Neck Surg 1999;51(1, Suppl 1):23–30

5 Das SK. Aetiological evaluation of foreign bodies in the ear andnose. J Laryngol Otol 1984;98(10):989–991

6 Higo R, Matsumoto Y, Ichimura K, Kaga K. Foreign bodies in theaerodigestive tract in pediatric patients. Auris Nasus Larynx 2003;30(4):397–401

7 Ray R, Dutta M, Mukherjee M, Gayen GC. Foreign body in ear, noseand throat: experience in a tertiary hospital. Indian J OtolaryngolHead Neck Surg 2014;66(1):13–16

8 Adhikari P, Shrestha BL, Baskota DK, Sinha BK. Accidental foreignbody ingestion: analysis of 163 cases. Int Arch Otorhinolaryngol2007;11(3):267–270

9 Pokharel R, Adhikari P, Bhusal CL, Guragain RP. Oesophagealforeign bodies in children. JNMA J Nepal Med Assoc 2008;47(172):186–188


Foreign Bodies in the Ear, Nose, and Throat Parajuli 123

Differential Diagnosis and Treatment of IsolatedPathologies of the Sphenoid Sinus:Retrospective Study of 46 CasesThomas Ribeiro Marcolini1 Maryane Cristine Safraider1 Jan Alessandro Socher2

Guilherme Olinto Lucena3

1Department of Medicine, Universidade Regional de Blumenau,Blumenau, SC, Brazil

2Department of Otorhinolaryngology, Universidade Regional deBlumenau, Blumenau, SC, Brazil

3Department of Medicine, Faculdade Evangélica do Paraná, Curitiba,PR, Brazil


Address for correspondence Thomas Ribeiro Marcolini, MD,Department of Medicine, Universidade Regional de Blumenau, RuaColombo 397 apto 05, Cornélio, Procópio PR, Brazil(e-mail: [email protected]).

Introduction

Isolated disease of the sphenoid represents 1 to 2% of allsinuses infections. The sphenoid sinus has often been ne-glected because of its isolated location and difficult access.1

Furthermore, the rarity of sphenoidal involvement can beexplained by the nonspecific symptoms, the inaccessibility tothe sinus through the otorhinolaryngological physical exam-ination, and the low number of diagnoses prior to the advent

of more sophisticated technology, such as computed tomog-raphy (CT) and magnetic resonance imaging (MRI). Theisolated involvement of the sphenoid sinus in most caseshas an inflammatory origin, and a neoplastic origin is rare.2–4

Sphenoid sinus diseases are difficult to diagnose and treat,because initial symptoms are vague and difficult to recog-nize.5 The clinical presentation involves headache (mainlyretro-orbital), whichmay be associatedwith purulent rhinor-rhea, retropharyngeal drip, nasal obstruction, abnormal

Keywords

► sphenoid sinus► paranasal sinuses► sphenoid sinusitis► cerebrospinal fluid

leak► paranasal sinus

neoplasms

Abstract Introduction Isolated disease of the sphenoid is rare and has often been overlookeddue to its remote location and difficult access.Objective A retrospective study of the main causes of isolated sphenoid sinus diseaseswith discussion of the most appropriate methods of diagnosis and treatment.Methods A total of 46 cases of isolated sphenoid disease treated between Janu-ary 2008 and December 2013 were evaluated by objective ear, nose, and throatexamination and video endoscopy, computed tomography of the paranasal sinuses,and, in some cases, magnetic resonance imaging. In each case, we decided betweendrug and/or endoscopic treatment.Results We identified 12 cases of isolated sphenoiditis (26.1%), 3 cases of fungalsphenoiditis (6.5%), 3 cases of sphenochoanal polyps (6.5%), 22 cases of mucocele(47.8%), 2 cases of cerebrospinal fluid leak (4.3%), and 1 case each of meningoence-phalocele (2.1%), inverted papilloma (2.1%), fibrous dysplasia (2.1%), and squamous cellcarcinoma (2.1%).Conclusion A prevalence of inflammatory and infectious diseases was found, andendoscopic surgery for the sphenoid sinus approach is effective in treating variousdiseases of the isolated sphenoid, whether complicated or not.

receivedOctober 6, 2014acceptedNovember 1, 2014published onlineJanuary 28, 2015




124

vision, nerve deficits, and consequential damage of otherinflammatory or neoplastic processes installed. Delayed di-agnosis and treatment can result in serious complications.This is due to the anatomical relations with the brain,meninges, optic nerve, internal carotid artery, cavernoussinus, and cranial nerve (oculomotor, trochlear, ophthalmic,and maxillary branches of the trigeminal nerve andabducens).6

Before the introduction of nasal endoscopy and betterimaging technologies such as CT and MRI, the deep positionof the sphenoid sinus prevented a proper early diagnosis ofpathologies there. Moreover, its location in the skull basedoes not allow a good view through conventional radio-graphs. Routine use of endoscopic surgery and introductionof sophisticated imaging techniques has allowed more fre-quent diagnoses of these rare pathologies.7,8 The aim of thisretrospective study was to survey the leading causes ofisolated sphenoid pathologies, with discussion of the mostappropriate methods of diagnosis and treatment.

Methods

We analyzed 46 charts and examination records from Janu-ary 2008 to December 2013 in patients with isolated sphenoidpathology. At the time of the diagnosis, there was no involve-ment of other sinuses. Only tumors involving the sphenoidsinus were included. Patients were evaluated by objective ear,nose, and throat examination and by nasal-sinus video endos-copy, 3.2-mm diameter flexible and/or rigid with 30-degreeoptics. The diagnosis was confirmed by CT of the paranasalsinuses and nasal cavities, in axial and coronal sections, withand without contrast, using a 2,500- to 3,500-rads bonewindow. In some cases, MRI was used: when there waspossible bone erosion of the sinus wall, presence of cerebro-spinalfluid leak andvisual changes. Isolated sphenoiditis caseswithout complication were treated with antibiotics (levoflox-acin 500 mg/d) for 21 days associated with corticosteroid(prednisone 40 mg daily) for 7 days. The patients with wors-ening or maintenance of symptoms during treatment orwithin 6weeks after it received endoscopic surgical treatment.Benign or malignant pathologies (tumor, dysplasia, cerebro-spinal fluid leak, mucocele, fungal sphenoiditis) were treated

surgically by endoscopic transnasal or transethmoidal sphe-noid sinusotomy. Patients were postoperatively followed for atleast 6 months for inflammatory diseases and up to 4 years forother benign and malignant pathologies.

Results

We identified 12 cases of isolated sphenoiditis (26.1%), in 8female (66.6%) and 4male subjects (33.3%) from13 to 73 yearsof age; 22 of mucocele (47.8%), in 13 women and 9 men from18 to 62 years old; 3 of fungal sphenoiditis (6.5%), in 2womenand 1 man from 33 to 68 years old; 3 of sphenochoanal polyp(6.5%), in 2 women and 1 man from 30 to 52 years old; 2 ofcerebrospinal fluid leak (4.3%), in 1 female and 1male patient,17 and 49 years old; 1 of meningoencephalocele (2.1%), in a14-year-old girl; 1 of inverted papilloma (2.1%), in a 45-year-old man; 1 of osteofibrous dysplasia (2.1%), in a 41-year-oldwoman; and 1 of squamous cell carcinoma (2.1%), in a48-year-old man.

Themost common symptomswereheadache/retro-orbitalor frontal facial pain in 33 cases (71.7%), nasal obstruction in15 patients (32.6%), cerebrospinal fluid rhinorrhea in 7 cases(15.2%), and mucopurulent rhinorrhea in 6 patients (13.0%).Also, 2 patients presented with epiphora (4.3%), 5 with fever(10.9%), 2 with epistaxis (4.3%), 1 with diplopia (2.1%);another patient’s level of consciousness was altered and herequired hospitalization (2.1%).

MRI was needed in only 6 cases (13.0%). Drug treatmentwas required in 7 cases, sphenoidal sinusotomy in 30, andtransethmoidal sphenoidal sinusectomy in 8. Only 2 casesexperienced complications, in which the patients sufferedsignificant bleeding during the transethmoidal sphenoidalsinusectomy, controlled with sphenopalatine artery ligation.

Discussion

All patients with isolated sphenoiditis reported retro-orbitaland/or frontal headache, and 7 had postnasal drip, nasalobstruction, epiphora, and/or fever. There was 1 case ofconsciousness level decrease. Video endoscopy revealed 7patients (58.3%) with signs of inflammation, characterizedby mucosal edema and mucopurulent discharge arising from

Fig. 1 Axial computed tomography shows sphenoid sinus opacification and greater wing of sphenoid compatible with isolated left sphenoiditis.


Isolated Pathologies of Sphenoid Sinus Marcolini et al. 125

the sphenoethmoidal recess. CT revealed an opacity of thesphenoid sinus without any abnormalities in other paranasalsinuses. In 2 cases, CT showed signs of osteitis (►Figs. 1 and 2),and in 1 case, a sidewall erosion progressed to an extraduralempyema (►Fig. 3). Drug therapywith antibiotics and cortico-steroids was used in 7 patients, 4 cases were treated byendoscopic transnasal sphenoid sinusotomy, and 1 case wastreated through endoscopic transethmoidal sphenoid sinus-ectomy. Patients who underwent surgery were evaluatedwithvideo endoscopyweekly, at 1month postoperatively, and thenmonthly thereafter. CT of the paranasal sinuses performed 90and 180 days after surgery confirmed the absence of sphenoi-ditis evidence.

In the cases of sphenoid mucocele, the most importantsymptom was retro-orbital pain. A video endoscopy showedmucosal edema at the sphenoethmoidal recess in only 9

patients, and CT revealed a homogeneous sphenoidal masswithout adjacent bone erosion (►Fig. 4). All patients under-went endoscopic transnasal sphenoid sinusectomy decom-pression. There were no signs of recurrence at a minimum of1-year postoperative follow-up.

Of the 3 patients with fungal sinusitis, 2 presented retro-orbital pain and unilateral purulent rhinorrhea, and the othercomplained only about headache. The endoscopic examina-tion showed edema and purulent discharge at the sphenoeth-moidal recess, with postnasal drip. CT in all cases showedheterogeneous images occupying the sphenoid sinus withsigns of calcification and osteitis (►Figs. 5 and 6). Transnasalendoscopic sphenoid sinusotomy treatment was used. Thehistologic results confirmed Aspergillus presence in all cases.There were no signs of recurrence in all cases with a mini-mum of 1-year postoperative follow-up.

Fig. 2 Axial computed tomography demonstrates homogeneoussphenoid sinus opacification with direct osteitis signs.

Fig. 3 Preoperative coronal computed tomography shows left sinusopacification evolving to extradural empyema.

Fig. 4 Preoperative axial computed tomography shows homogeneousopacification on the right sphenoid sinus with no signs of adjacentbone erosion.

Fig. 5 Preoperative coronal computed tomography shows hetero-geneity occupying the right sphenoid sinus with signs of calcification.


Isolated Pathologies of Sphenoid Sinus Marcolini et al.126

The 3 patientswith sphenochoanal polyp presented symp-toms of unilateral nasal obstruction, and 2 suffered fromfrontal or retro-orbital headache. Video endoscopy was diag-nostic in all 3 cases, revealing polypoid tumor in sphenoeth-moidal recess. CT showed an expansive homogeneous imagewith evidently precise limits in the nasal cavity, with noevidence of bone erosion (►Fig. 6). Patients who underwentendoscopic transethmoidal sphenoidal sinusectomy showedno signs of recurrence for at least 1 year postoperatively.

Of the 2 patientswith cerebrospinal fluid leak, 1 reported aprevious history of hospitalization because of meningitis.Both patients reported unilateral rhinorrhea and deniedprevious nasal or sinus surgery and/or cranioencephalictrauma. In all patients, the presence of cerebrospinal fluid

leak was confirmed by studies of β2-transferrin. Endoscopicexams did not show significant changes, and CT demonstrat-ed complete or partial dehiscence with opacification of thesphenoidal sinus wall. MRI showed hyperintense signal on T2compatible to liquid occupying the sphenoid sinus. All pa-tients underwent endoscopic transnasal sphenoidal sinus-otomy to reverse the cerebrospinalfluid leakwith free graft ofinferior concha mucoperiosteum fixed with fibrin glue. Therewere no signs of recurrencewithin postoperative follow-up of3 years and 2 months.

The main symptoms were nasal in the 14-year-old girlwith meningoencephalocele, with obstruction and abundantunilateral rhinorrhea. Video endoscopy revealed the presenceof cerebrospinal fluid rhinorrhea from the left sphenoeth-moidal recess after the Valsalva maneuver. CT showed adehiscence of the bone wall with partial opacification ofthe left sphenoid sinus and meningoencephalic herniation(►Fig. 7). The patient underwent endoscopic transethmoidalsphenoid sinusectomy for correcting the mass protrusion,and the leak was closed with free graft of inferior conchamucoperiosteum fixed with fibrin glue.

The 45-year-old man with inverted papilloma presentedwith the main complaints of frontal and retro-orbital head-ache associated with nasal obstruction and right-sided epi-staxis. A polypoid tumor in the right sphenoethmoidal recesswas identified by endoscopy, and CT revealed a heteroge-neous opacification of the sphenoid sinus with contrastuptake, signs of remodeling process of the sphenoid sinuswalls, and bone erosion, with no compromise of adjacentstructures (►Fig. 8). The patient underwent a bilateral resec-tion of the tumor by endoscopic transethmoidal sphenoidsinusectomy, and the histologic studies confirmed the diag-nosis of inverted papilloma. No recurrence was noted atpostoperative follow-up of 2 years and 6 months.

The main complaint of the patient with fibrous dysplasiawas an intense frontal and retro-orbital left-sided headache.

Fig. 7 Preoperative coronal computed tomography demonstratespartial sphenoid sinus opacification and right sphenoid mass occu-pying precise limits compatible with right sphenochoanal nasal cavitypolyp.

Fig. 8 Preoperative coronal computed tomography section withdetails showing the sphenoid’s lateral wall opacification and dehis-cence with partial opacification of the lateral sphenoid’s recess.

Fig. 6 Preoperative coronal computed tomography shows hetero-geneity with calcification signs occupying the right sphenoid sinuswith osteitis signs.



The endoscopic examination revealed no significant changes,and CT showed areas of thickening and sclerosis occupyingthe left sphenoid sinus (►Fig. 9). The patient was treated byendoscopic transethmoidal sphenoid sinusectomy andshowed improvements of the disease symptoms during thepostoperative follow-up of 2 years.

In the case of squamous cell carcinoma, the symptomsincluded retro-orbital headache, nasal obstruction, left-sideddiplopia, and eventual epistaxis. The endoscopic examinationrevealed sphenoethmoidal recess edema, and CT showed anexpansive process including erosion of the bone wall of theleft sphenoid sinus (►Fig. 10). MRI identified a mass withgadolinium contrast enhancement and confirmed optic nervecompression without infiltration of other structures(►Fig. 11). The patient underwent bilateral transethmoidalsphenoid sinusectomy to resect the tumor with a safe margin

and decompression of the optic nerve. Radical resection wasnot done to preserve the other adjacent structures. Thepatient was referred for further treatment with radiotherapywith a dose of 50 Gy. No recurrence was noted during thepostoperative follow-up of 4 years and 2 months (►Fig. 12).

Conclusion

A substantial prevalence of inflammatory and infectioussphenoid sinus diseases was found, and treatment by endo-scopic approach was effective in isolated sphenoid patholo-gies, whether complicated or not.

Although video endoscopy does not show great sensitivityand specificity for the diagnosis of isolated sphenoid

Fig. 9 Preoperative coronal section computed tomographyhighlighting the right sphenoid heterogeneous opacification withsigns of remodeling in the wall of the sphenoid sinus and bone erosion.

Fig. 10 Preoperative axial computed tomography demonstratingareas of thickening and sclerosis in the left sphenoid sinus.

Fig. 11 Preoperative axial computed tomography demonstrates aheterogeneous mass occupying the sphenoid sinuses, erosion of thesinus bony wall, and compression of the left optic nerve.

Fig. 12 Postoperative sagittal magnetic resonance image shows thetumor resection.


Isolated Pathologies of Sphenoid Sinus Marcolini et al.128

pathologies, CT scans and MRI are useful for diagnosis andtreatment planning.

References1 Grillone GA, Kasznica P. Isolated sphenoid sinus disease. Otolar-

yngol Clin North Am 2004;37(2):435–4512 Lew D, Southwick FS, Montgomery WW, Weber AL, Baker AS.

Sphenoid sinusitis. A review of 30 cases. N Engl J Med 1983;309(19):1149–1154

3 Hnatuk LAP, Macdonald RE, Papsin BC. Isolated sphenoid sinusitis:the Toronto Hospital for Sick Children experience and review ofthe literature. J Otolaryngol 1994;23(1):36–41

4 Metson R, Gliklich RE. Endoscopic treatment of sphenoid sinusitis.Otolaryngol Head Neck Surg 1996;114(6):736–744

5 Martin TJ, Smith TL, Smith MM, Loehrl TA. Evaluation and surgicalmanagement of isolated sphenoid sinus disease. Arch OtolaryngolHead Neck Surg 2002;128(12):1413–1419

6 Mra Z, Roach JC, Brook AL. Infectious and neoplastic diseases ofthe sphenoid sinus—a report of 10 cases. Rhinology 2002;40(1):34–40

7 Castelnuovo P, Pagella F, Semino L, De Bernardi F, Delù G. Endo-scopic treatment of the isolated sphenoid sinus lesions. Eur ArchOtorhinolaryngol 2005;262(2):142–147

8 Socher JA, Cassano M, Filheiro CA, Cassano P, Felippu A. Diagnosisand treatment of isolated sphenoid sinus disease: a review of 109cases. Acta Otolaryngol 2008;128(9):1004–1010



Surfactant Protein A Expression in ChronicRhinosinusitis and Atrophic RhinitisMohammad Waheed El-Anwar1 Atef A. Hamed1 Abd ElRaof Said Mohamed1

Ahmad Abdel-Fattah Nofal1 Maha A. Mohamed2 Hesham R. Abdel-Aziz3

1Department of Otorhinolaryngology, Head and Neck Surgery,Zagazig University, Zagazig, Egypt

2Department of Clinical Pathology, Zagazig University, Zagazig, Egypt3Department of Pathology, Zagazig University, Zagazig, Egypt


Address for correspondence Mohammad El-Anwar, MD, Departmentof Otorhinolaryngology Head and Neck Surgery, Zagazig University,Zagazig 0020552309843, Egypt (e-mail: [email protected]).

Introduction

Pulmonary surfactant is secreted as lamellar bodies from typeII pneumocytes to reduce surface tension at the air–liquidinterface. Over 90% of pulmonary surfactant is lipid, whereasthe remainder is composed of plasma proteins and thesurfactant proteins (SPs) A, B, C, and D. SPs are hydrophilicproteins actively involved in the innate immune response toinhaled pathogens. SPs are part of the collectin family ofproteins secreted at several epithelial surfaces in the body.1

SP-A, hydrophilic sialoglycoprotein, exhibits antimicro-bial properties and interacts with a variety of respiratory

tract pathogens that have related molecular structures,called pathogen-associated molecular patterns (PAMP).PAMPs differ little from one pathogen to another but arenot found in the host.2 SP-A bind PAMPs located on micro-bial membranes via their calcium-dependent carbohy-drate-binding domains. Independent of direct binding topathogens, SP-A also interacts directly with dendritic cells,modulates subsequent T-cell responses, optimizes leuko-cyte function and chemotaxis, modifies subsequent cyto-kine/chemokine profiles, and activates complement. Thus,these proteins are crucial in the initial interaction, recog-nition, processing, and subsequent adaptive immune

Keywords

► sinusitis► rhinitis► nose

Abstract Introduction Surfactant protein A (SP-A) exhibits antimicrobial properties and inter-acts with a variety of respiratory tract pathogens.Objective The objective of this study was to detect the presence of SP-A and measureits alterations in chronic rhinosinusitis (CRS) and primary atrophic rhinitis (PAR) versushealthy controls.Methods Inferior turbinate and sinus mucosal biopsies were taken from 30 patientswith CRS, 30 patients with PAR, and 20 healthy controls. Immunohistochemical stainingfor SP-A and polymerase chain reaction (PCR) amplification of SP-Amessenger RNAwereperformed on nasal tissue samples.Results Immunostaining localized SP-A to the mucosa and submucosal glands in CRSspecimens but failed to localize it in PAR specimens. Quantitative PCR showed a high,statistically significant increase in the SP-A levels of patients with CRS when comparedwith controls (p < 0.0001) and also demonstrated a significant reduction of SP-A inpatients with PAR compared with controls (p < 0.005).Conclusion SP-A is significantly increased in CRS and decreased significantly in PARand appears to be expressed by respiratory epithelial cells and submucosal glandularelements of the sinonasal mucosa. The potential therapeutic applications of surfactantin the enhancement of mucociliary clearance need to be studied.

receivedNovember 23, 2014acceptedJanuary 10, 2015published onlineFebruary 20, 2015




130

response for a wide variety of inhaled pathogens andallergens.3 Human SP-A is encoded by two genes, SP-A1and SP-A2.2

Bronchoalveolar lavage specimens from patients withchronic bronchial asthma or pneumocystis pneumonia havedemonstrated increased SP-A.3 SP-A expression has also beenassessed in the sinonasalmucosa. In a semiquantitative study,Dutton et al found elevated SP-A in themucosa of rabbitswithintercurrent sinusitis or antibiotic-treated sinusitis comparedwith pathogen-free animals.4

Diseases of the sinonasal system, specifically chronic rhi-nosinusitis (CRS), are often associatedwith pulmonary systemdiseases. Asthma sensitive to the ingestion of aspirin may beassociated with nasal polyps. Allergic fungal rhinosinusitis inthe upper airway could be associated with allergic broncho-pulmonary aspergillosis.5 Patients with cystic fibrosis invari-ably develop CRS in addition to their debilitating pulmonarydisease. These are attributed to similar mechanisms of inspis-sated mucus, impaired mucociliary clearance, and persistentbacterial infections and inflammation. Moreover, the lungsand the paranasal sinuses share contact with inhaled patho-gens and include many of the same epithelial properties.6

SPs appear to have an important function in immunologicand rheological process of the nasal mucosa and support thetherapeutic use of liposomal nasal sprays.7

The aim of this study was to investigate whether SP-A ispresent in sinonasal mucosa and compare expression indistinct types of chronic rhinitis [CRS and primary atrophicrhinitis (PAR)] versus healthy control mucosa.


This study was performed from February 2011 to Septem-ber 2014. Informed written consent was obtained fromincluded subjects, and the Institutional Review Board ap-proved the study. The subjects were divided into threegroups; the first group included 30 patients with persistentbacterial CRS despite appropriate medical treatment andscheduled for endoscopic sinus surgery. Diagnosis of CRSdepended on Meltzer et al criteria of symptoms that persistmore than 12 weeks. Sinus mucosa was biopsied during thecourse of endoscopic sinus surgery.8

The second group included 30 patients with PAR randomlyselected from the outpatient clinic. Diagnosis of atrophicrhinitis followed Ly et al diagnostic criteria for atrophicrhinitis with two or more clinical features such as recurrentepistaxis, episodic anosmia, physician-documented nasalpurulence, or nasal crusting for �6 months with atrophiedinferior turbinatewith no detected cause.9 Cases known to besecondary atrophic rhinitis and patients with history of nasaltrauma, sinonasal surgery, granulomatous disease, or radia-tion to the sinonasal area were excluded. Nasal mucosalbiopsies were taken from inferior turbinate under localanesthesia.

The third group included 20 normal controls; nasal muco-sal biopsies were obtained from these subjects undergoingsurgery for other reasons other than CRS such as rhinoplasty(10 cases from normal inferior turbinate mucosa) and zygo-

maticomaxillary fractures (10 cases from normal maxillarysinus mucosa).

Immunohistochemical staining for SP-A and polymerasechain reaction (PCR) amplification of SP-A messenger RNAwere performed on resected nasal tissues.

Immunohistochemical AnalysisBiopsy specimens were washed three times with iced phos-phate-buffered saline for 5 minutes to remove any exogenousSP-A that may have migrated from the lower airways. Afterthawing to �20°C and overnight decalcification, followed byfixation in 4%paraformaldehyde, 4-μm-thick sectionswere cutfrom paraffin blocks containing representative tissue samples.Paraffin sections were dewaxed in xylene, rehydrated througha graded alcohol series, placed in 10 mmol/L of citrate buffer,and submitted to heat retrieval using a vapor lock for 40minutes. After heating, the slides were allowed to cool toroom temperature and briefly washed with Tris (hydroxy-methyl) amino methane-buffered saline. Endogenous peroxi-dase activity was blocked with 3% hydrogen peroxide inmethanol for 5 minutes. Normal serum (Novo stain SuperABC kit, Novocastra, Newcastle upon Tyne, England) was usedfor 30 minutes to block nonspecific immunostaining. Immu-nohistochemical staining was performed using a standardavidin-biotin peroxidase system (Novo stain Super ABC kit).The monoclonal antibody SP-A (isotype; immunoglobulin [Ig]G2a purchased from Abcam) were used in concentration of 1/200. Two slides for SP-A antibodies and isotypic control (mousemonoclonal IgG2a as negative control) were incubated over-night at room temperature. Following washing in phosphate-buffered saline, biotinylated universal secondary antibody(Novo stain Super ABC kit) was applied for 30 minutes. Thesections then were incubated with the avidin-biotin complexreagent (Novo stain Super ABC kit) for 30 minutes and devel-oped with 3, 3-diaminobenzidine tetrahydrochloride in phos-phate-buffered saline (pH 7.5) containing 0.036% hydrogenperoxide for 5 minutes. Light Mayer hematoxylin was appliedas a counterstain. The slides thenwere dehydrated in a series ofethanol and mounted with permount. Cytoplasmic or mem-branous staining was considered as positive. Two negativecontrol sections were used in each case; one was incubatedwith the secondary antibody only, the other with the primaryantibody only. Sections of human lung stained with anti SP-Aantibodies were used for positive control. Cells were consid-ered positive for anti SP-A antibodies when at least 10% of cellsshow diffuse cytoplasmic or membranous staining.

Quantitative Reverse-Transcriptase PolymeraseChain ReactionTotal RNApurificationwasperformedonall specimens (using anAurum Total RNA Mini Prep-Kit; Bio-Rad, Hercules, California,United States), including a DNase 1 digest, according to themanufacturer’s instructions. Reverse transcriptase PCR wasperformed on a portion of the purified RNA for each specimento create complementary DNA (cDNA) libraries using the iScriptcDNA Synthesis Kit (Bio-Rad) under the following conditions in aconventional thermocycler: 5minutes at 25°C, 30minutes at 42°C, and 5 minutes at 85°C.


SP-A in Chronic Rhinosinusitis and Atrophic Rhinitis El-Anwar et al. 131

Quantitative Polymerase Chain ReactionIntron-exon spanning SP-A primers were used in the quanti-tative PCR amplification reactions with the following design:5′-AAGCCACACTCCACGACTTTAGA-3′ and 5′-GCCCATTGCTG-GAGAAGACCT- 3′ (Integrated DNA Technology Inc., Coralville,Iowa, United States). Commercially available _-actin primersfor the amplification of the housekeeping gene were used:5-TCATGAAGTGTGACGTTGACATCCGT-3 and 5′-CTTAGAAG-CATTTGCGGTGCACGATG-3 (Promega, Madison, Wisconsin,United States). In a quantitative PCR detection system (MyiQ Thermal Cycler; Bio-Rad), 96-well reaction plates were usedto perform real-time, quantitative PCR with the sample wellscontaining the following: 25 μL 2_ Supermix with SYBR green (afluorescent double-strandedDNA-binding reportermolecule), 1μL eachof SP-Aprimer (100ng/μL) or _-actinprimers (100pmol/μL) for controlwells, a 1-μL sample complementary DNA, and 22μL of nuclease-free water. After an initial activation step at 95°Cfor 3minutes, amplificationwasperformedacross 45 cycleswiththe following parameters: denaturation for 30 seconds at 95°C,annealing for 20 seconds at 58°C, and elongation for 30 secondsat 72°C. Digital capture of SYBR green fluorescence was per-formed during the annealing phase for threshold cycle assess-ment. The threshold cycle valueswere automatically stored in anoutput file, and these were analyzed to determine relativestarting messenger RNA (mRNA) concentrations according tothe 2-__threshold cycle method16 using a proprietary macro(Bio-Rad)with Excel 2003XT (Microsoft Corp., Redmond,Wash-ington, United States). A melt-curve analysis confirmed speci-men uniformity and estimated product size in each sample wellin the 96-well plate. The following protocol was programmedinto the My iQ Thermal Cycler (Bio-Rad) to produce the meltcurves for each samplewell after amplification using SYBR greenas the reporter: 1 minute at 95°C, 1 minute at 55°C, and10 seconds at 55°C for 80 cycles. Finally, to verify PCR productsequence, random samples were prepared and sequenced bythe Vanderbilt University (Nashville, Tennessee, United States)DNA sequencing core laboratory. Data were compared with thehuman genome via BLAST downloadable software (http://www.ncbi.nlm.nih.gov/genome/seq/BlastGen/BlastGen.cgi?taxid=9606).

Statistical AnalysisStatistical analyses were performed using SPSS 14.0 statisticalsoftware for Windows (SPSS Inc., Chicago, Illinois, UnitedStates). The significance level was set at p < 0.05. Quantitative

data (age and quantitative PCR) in patient and control groupswere registered and compared using t test, and sex differencesbetween the three groups were analyzed by chi-square test.

Results

Eighty subjects were included in current study: 30 patientswith CRS, 30 with PAR, and 20 controls. The mean age( � standard deviation) was 37 ( � 14.2) years. The threegroups were matched for sex (p ¼ 0.84, chi-square ¼ 0.341)and sex as nonsignificant differences in age between CRS andcontrols (p ¼ 0.64, t ¼ 0.47), and nonsignificant differencesbetween PAR and controls (p ¼ 0.97, t ¼ 0.035) werereported (►Table 1). There was strong positive staining inthe nasal mucosa and submucosa in CRS, as shown in►Fig. 1.This indicates that SP-A is secreted by nasal epithelium andglandular epithelium. Faint staining for SP-A was detected inbiopsies of control specimens as shown in ►Fig. 2, andstaining for SP-A was not detected in biopsies of patientswith atrophic rhinitis (►Fig. 3). As a negative staining control,sections were incubated with nonimmune rabbit seruminstead of SP-A antibody, resulting in loss of staining.

Quantitative PCR (reported as ΔCT � standard deviation)showed highly significantly increased levels of SP-A(4.69 � 2.7) mRNA (t ¼ 6.6417, p < 0.0001) from patientswith CRSwhen comparedwith controls (10.4 � 3.15). AlthoughSP-A mRNA was decreased in patients with PAR (15.6 � 6.25)when compared with controls (10.4 � 3.15), this decrease wasstatistically significant (t ¼ 3.8778, p ¼ 0.0003).

Discussion

SP-A plays an integral part in the innate defense system. SP-Aexpression and function are altered in a variety of inflamma-tory and infectious diseases. However, the presence of SP-A inthe human paranasal sinus mucosa is not well known.7

There are few previous studies investigating surfactantand SP in the upper airway.10 The presence of lamellar bodiesand SP-A, -B, and -D have been described in the porcineeustachian tube, indicating that a surfactant-like system ispresent outside the pulmonary system.11 Woodworth et aldemonstrated the presence of lamellar bodies, SP, and mRNAin both diseased and normal human sinus tissue and reportedthat SP-A and SP-D immunolocalized to the sinonasal epithe-lium and submucosal gland secretory ducts.12 The discovery

Table 1 Age and sex of the patients (CRS, PAR) and control (Healthy) groups

Subjects CRS PAR Controls P value

n % n % n %

Sex

Male 18 60 16 53.3 128

6040

0.84aNSFemale 12 40 14 46.7

Age 29.5 � 10.1 28.3 � 9.7 29.6 � 9.6 NSb

Abbreviations: CRS, chronic rhinosinusitis; PAR, primary atrophic rhinitis; NS, not significant.aChi-square test.bDifferences between CRS and healthy (p ¼ 0.64, t ¼ 0.47) and differences between PAR and healthy (p ¼ 0.97, t ¼ 0.035).


SP-A in Chronic Rhinosinusitis and Atrophic Rhinitis El-Anwar et al.132

of surfactant production and secretion in sinonasal mucosaindicates that initial contact and interaction between patho-gens and SPs occurs after inhalation and deposition into themucus of the upper respiratory tract.

This study characterizes the differential expression of SP-Ain the nasal mucosal samples of patients with CRS usingimmunohistochemical staining and quantitative PCR. Byimmunohistochemistry, we detected strong SP-A staining inthe epithelial cells and lumen of the submucosal glands in theCRS patients. Also SP-A mRNA increased significantly in CRSpatients and decreased significantly in patients with PAR.

Strong SP-A staining in the epithelial cells and lumen of thesubmucosal glands in the CRS could be explained on the basisof the different histologic findings present in normal mucosaand CRS mucosa, because SP production mostly occurs insubmucosal glandular pathways and these patients exhibitglandular hyperplasia. This hyperplasia is more pronouncedin patients with CRS. This may have contributed more mRNAtranscripts due to a relative abundance of these cell types. Inagreement, Woodworth et al reported a significant elevationof SP-A and -D in patients with CRS associated with cysticfibrosis.6 In the sinonasal tissue of rabbits with acute bacterial

Fig. 1 Photomicrograph of chronic rhinosinusitis showing strong cytoplasmic immunostaining of surfactant protein-A in both nasal mucosa (A)and submucosal glands (B). Hematoxylin counterstain � 400.

Fig. 2 (A) Photomicrograph showing faint immunostaining of surfactant protein A (SP-A) in human nasal mucosa and submucosa of the controlgroup; hematoxylin counterstain � 400. (B) Photomicrograph of atrophic rhinitis showing negative immunostaining for surfactant protein-A(SP-A) in human nasal mucosa and submucosa; hematoxylin.

Fig. 3 Photomicrograph showing negative control with negative immunostaining for surfactant protein-A (SP-A) in nasal mucosa (A) andsubmucosa (B). Hematoxylin counterstain � 400.


SP-A in Chronic Rhinosinusitis and Atrophic Rhinitis El-Anwar et al. 133

sinusitis (inflammation but not allergy), SP-A was moreprevalent than in pathogen-free animals.3

The SP-A protein levels were significantly reduced com-pared with controls in PAR patients. A possible explanation isthat absence of significant SP-A gene up-regulation anddepletion of SP-A protein reserves in the presence of atrophicchanges of the submucosal glands and mucosa. This is inagreement with the results obtained by Sayed et al, who haddemonstrated a significant decrease in phospholipid andsurfactant concentrations in the cases with PAR comparedwith normal cases.13

Several theories tried to explain the etiology of PAR.Infection, one of the theories, can affect the surfactant systemleading to surfactant deficiency through several mechanismsincluding inflammatory cytokines produced in response tosepsis inhibiting the synthesis of surfactant.14 Activatedneutrophils are capable of cleaving SP-A and impairingsurfactant function.15 Sayed et al reported that abnormalsurfactant function in the larger airwaysmight be expected todecrease the efficiency of mucociliary clearance and alsoadversely lead to stasis of mucus.13

In addition, surfactant deficiency will impair opsonizationand phagocytosis of bacteria by macrophages, thus favoringbacterial multiplication and leukocyte infiltration. Due to alack of the opsonizing effect (due to decreased concentrationof secretory IgA and surfactant deficiency), the leukocytescannot phagocytose and destroy the bacteriawith continuousproduction of proteolytic enzymes in the secretion, which areprobably harmful to the mucous membrane, as it is knownthat inhalation of proteolytic enzymes produces emphysemain animals. Our results support the previously reportedconcept that SP has an important function in immunologicprocess of the nasal mucosa.7

The therapeutic potential of correcting abnormalities inSPs has already been demonstrated in the lower airways. Thecorrection of SP-D deficiencies with topical SP-D reversesemphysema, pulmonary lipidosis, and macrophage infiltra-tions in diseased mice.15 Gesche et al found that human (rh)KGF, betamethasone, or their combination treatment in-creased secreted surfactant phosphatidylcholines in neonatalrats with lung injuries.16 Furthermore, topical SP-A and SP-Ddecreased IgE and eosinophilia in mouse models of allergicbronchopulmonary aspergillosis.17 Because allergic broncho-pulmonary aspergillosis is very similar to allergic fungalrhinosinusitis and can be considered its counterpart in thelower airway, this could lead to future therapeutic options fordifficult to treat sinus disease, as Tan et al found that thesurfactant does not appear to elicit cellular toxicity using an invitro explant model.18

Conclusion

This study characterized the detection and expression of SP-Ain human sinus mucosa. SP-A is significantly increased in CRSand decreased in PAR and appears to be expressed byrespiratory epithelial cells and submucosal glandular ele-ments of the sinonasal mucosa. Further characterization ofthis specialized protein is warranted considering the poten-

tial therapeutic applications of surfactant in the enhance-ment of mucociliary clearance.

References1 Woodworth BA, Smythe N, Spicer SS, Schulte BA, Schlosser RJ.

Presence of surfactant lamellar bodies in normal and diseased sinusmucosa. ORL J Otorhinolaryngol Relat Spec 2005;67(4):199–202

2 Khubchandani KR, Snyder JM. Surfactant protein A (SP-A): thealveolus and beyond. FASEB J 2001;15(1):59–69

3 Wootten CT, Labadie RF, Chen A, Lane KF. Differential expression ofsurfactant protein A in the nasal mucosa of patients with allergysymptoms. Arch Otolaryngol Head Neck Surg 2006;132(9):1001–1007

4 Dutton JM, Goss K, Khubchandani KR, Shah CD, Smith RJH, SnyderJM. Surfactant protein A in rabbit sinus and middle ear mucosa.Ann Otol Rhinol Laryngol 1999;108(10):915–924

5 Shah A, Panjabi C. Allergic aspergillosis of the respiratory tract. EurRespir Rev 2014;23(131):8–29

6 Woodworth BA, Wood R, Bhargave G, Cohen NA, Baatz JE,Schlosser RJ. Surfactant protein B detection and gene expressionin chronic rhinosinusitis. Laryngoscope 2007;117(7):1296–1301

7 Schicht M, Knipping S, Hirt R, Beileke S, Sel S, Paulsen F, Bräuer L.Detection of surfactant proteins A, B, C, and D in human nasalmucosa and their regulation in chronic rhinosinusitis with polyps.Am J Rhinol Allergy 2013;27(1):24–29

8 Meltzer EO, Hamilos DL, Hadley JA, et al; American Academy ofAllergy, Asthma and Immunology (AAAAI); American Academy ofOtolaryngic Allergy (AAOA); American Academy of Otolaryngolo-gy–Head and Neck Surgery (AAO-HNS); American College ofAllergy, Asthma and Immunology (ACAAI); American RhinologicSociety (ARS). Rhinosinusitis: establishing definitions for clinicalresearch and patient care. J Allergy Clin Immunol 2004;114(6,Suppl):155–212

9 Ly TH, deShazo RD, Olivier J, Stringer SP, Daley W, Stodard CM.Diagnostic criteria for atrophic rhinosinusitis. Am J Med 2009;122(8):747–753

10 Ooi EH, Wormald PJ, Carney AS, James CL, Tan LW. Surfactantprotein D expression in chronic rhinosinusitis patients and im-mune responses in vitro to Aspergillus and alternaria in a nasalexplant model. Laryngoscope 2007;117(1):51–57

11 Paananen R, Glumoff V, Sormunen R, Voorhout W, Hallman M.Expression and localization of lung surfactant protein B in eusta-chian tube epithelium. Am J Physiol Lung Cell Mol Physiol 2001;280(2):L214–L220

12 Woodworth BA, Neal JG, Newton D, et al. Surfactant protein A andD in human sinus mucosa: a preliminary report. ORL J Otorhino-laryngol Relat Spec 2007;69(1):57–60

13 Sayed RH, Abou-Elhamd KE, Abdel-Kader M, Saleem TH. Study ofsurfactant level in cases of primary atrophic rhinitis. J LaryngolOtol 2000;114(4):254–259

14 Arias-Díaz J, Vara E, García C, Gómez M, Balibrea JL. Tumournecrosis factor-alpha inhibits synthesis of surfactant by isolatedhuman type II pneumocytes. Eur J Surg 1993;159(10):541–549

15 Zhang L, Ikegami M, Dey CR, Korfhagen TR, Whitsett JA. Reversibilityof pulmonary abnormalities by conditional replacement of surfactantprotein D (SP-D) in vivo. J Biol Chem 2002;277(41):38709–38713

16 Gesche J, Fehrenbach H, Koslowski R, et al. rhKGF stimulates lungsurfactant production in neonatal rats in vivo. Pediatr Pulmonol2011;46(9):882–895

17 KishorU,MadanT, SarmaPU, SinghM,Urban BC, ReidKB. Protectiveroles of pulmonary surfactant proteins, SP-A and SP-D, against lungallergy and infection caused by Aspergillus fumigatus. Immunobi-ology 2002;205(4–5):610–618

18 Tan NC, Cooksley CM, Paramasivan S, Vreugde S, Wormald PJ.Safety evaluation of a sinus surfactant in an explant-based cyto-toxicity assay. Laryngoscope 2014;124(2):369–372


SP-A in Chronic Rhinosinusitis and Atrophic Rhinitis El-Anwar et al.134

Mercury Exposure in a Riverside AmazonPopulation, Brazil: A Study of the Ototoxicity ofMethylmercuryAna Hoshino1,2 Heloisa Pacheco-Ferreira3 Seisse Gabriela G. Sanches4 Renata Carvallo5

Nathália Cardoso1 Maurício Perez1 Volney de Magalhães Câmara1

1Universidade Federal do Rio de Janeiro, Instituto de Estudos emSaúde Coletiva, Rio de Janeiro, Brazil

2Department of Otorhinolaryngology, Faculdade de Medicina daUniversidade de São Paulo (FMUSP), São Paulo, Brazil

3Universidade Federal do Rio de Janeiro, Faculdade de Medicina, Rio deJaneiro, Brazil

4Universidade de São Paulo, FOFITO, São Paulo, Brazil5 FMUSP, FOFITO, São Paulo, São Paulo, Brazil


Address for correspondence Ana Hoshino, PhD, Department ofOtorhinolaryngology, Hospital das Clínicas, R. Vigário Albernaz, 843/74, São Paulo 04134-021, Brazil (e-mail: [email protected]).

Introduction

The health impacts due to acute and/or chronic exposure arealready well defined for most chemicals. Organizations such

as the US Environmental Protection Agency andWorld HealthOrganization established exposure limits, but many peoplelive above this limit. Environmental contamination withpersistent pollutants showing toxic and cumulative effects

Keywords

► mercury► methylmercury

compounds► auditory pathways► medial olivocochlear

system► otoacoustic emissions► ototoxicity

Abstract Introduction Mercury poisoning causes hearing loss in humans and animals. Acuteand long-term exposures produce irreversible peripheral and central auditory systemdamage, andmercury in its various forms of presentation in the environment is ototoxic.Objective We investigated the otoacoustic emissions responses in a riverside popula-tion exposed to environmental mercury by analyzing the inhibitory effect of the medialolivocochlear system (MOCS) on transient otoacoustic emissions (TEOAE).Methods The purpose of the research was to evaluate the entire communityindependently of variables of sex and age. All of the participants were born and livedin a riverside community. After otolaryngologic evaluation, participants were receivedtympanometry, evaluation of contralateral acoustic reflexes, pure tone audiometry, andrecording of TEOAEs with nonlinear click stimulation. Hair samples were collect tomeasure mercury levels.Results There was no significant correlation between the inhibitory effect of theMOCS, age, and the level of mercury in the hair.Conclusions The pathophysiological effects of chronic exposure may be subtle andnonspecific and can have a long period of latency; therefore, it will be important tomonitor the effects of mercury exposure in the central auditory system of the Amazonpopulation over time. Longitudinal studies should be performed to determine whetherthe inhibitory effect of theMOCS on otoacoustic emissions can be an evaluationmethodand diagnostic tool in populations exposed to mercury.

receivedNovember 6, 2014acceptedDecember 11, 2014published onlineFebruary 19, 2015



THIEME


on organisms, as in the case of mercury, has reached globalproportions due to the persistence and mobility of thesesubstances.1 Because it is very toxic and has the tendency tobioaccumulate and because of biomagnification along thefood chain, mercury is recognized as a potentially dangerouspollutant in the marine environment.

Historically, two great exposures to methylmercury haveoccurred. In Minamata (Japan), women showed hair mercuryconcentrations between 10 and 100 µg/g. The other greatexposure occurred in Iraq, where women with elevated hairmercury levels (�10 µg/g) had children in whom neurologicalterations were observed after birth.2 Passos and Mergler,studying mercury exposure in Amazon communities,reported that even different bioindicators have revealedmean hair mercury levels higher than 15 µg/g, which arethe highest reported mercury levels in the world.3

The Amazon region occupies a prominent place in thisscenario,4 and the riverside populations of that region sufferthe health impacts associated with environmental contami-nation. Fish, the main source of dietary protein for riversidepopulations, is consumed at an average of 406 g/d and iscontaminated with mercury.5 The consumption of mercury-contaminated fish gives rise to a series of neurotoxic effectsthat are observed in pregnant women, fetuses, and newborns,particularly when they are exposed to mercury during braindevelopment. The prenatal exposure may result in smallalterations in development that appear over time.6

Mercury poisoning causes hearing loss in humans andanimals. In a systematic review, the authors conclude thatacute and long-term exposures produce irreversible periph-eral and central auditory system damage and that mercury inits various forms of presentation in the environment isototoxic.7 The consequences of hearing loss can be irrevers-ible, with clinical alterations that often go undetected byneurologic examination and biological markers, thus jeopar-dizing the quality of life of these individuals. Follow-up ofototoxicity is important for detecting hearing changes, there-by suggesting new treatment strategies for the patient andintervention when a disabling hearing impairment occurs.8

Mercury is a liquid that expands or shrinks very accuratelyaccording to the temperature. These particular properties areextremely useful for various applications such as the produc-tion of electronic measuring equipment, fluorescent lamps,batteries, dental amalgam, and cosmetics (in small amounts).The mercury used can be released to the environment atdifferent stages of the production, such as industrial gas,industrial waste, and contaminated waste. Thus, it candestroy the soil, air, and water, affecting environmental andhuman health directly.4

Once mercury enters the aquatic ecosystems, the micro-organisms transformed into a highly toxic form of mercurycalled methylmercury, which is accumulated and biomagni-fies in fish and shellfish and those who consume them. Thelevels of methylmercury in some fish species can reachmillions of times above the levels present in water from itssurroundings.

Even at low doses of exposure, mercury, especially meth-ylmercury, can penetrate the placenta and appear in breast

milk. This can disturb development of brain functions and cancreate deficits in language skills, memory, attention, andmotor and visual skills. When exposure to mercury combineswith malnutrition, the risk grows considerably.

Mercury exposure can be monitored through hair, blood,breastmilk, urine, and nails. The level ofmercury in blood andurine shows a recent exposure. Levels in hair reflect long-term exposure.9 The Agency for Toxic Substances and DiseaseRegistry estimated that the minimum risk level for methyl-mercury intake would be 0.3 mg/kg per day.10 This value isbased on estimated level of adverse effects (NOAEL) of 1.3mg/kg per day calculated for methylmercury hair levels. Theestimated level of adverse effects was obtained from a studyof the Seychelles population that observed the neurodevel-opment of children.9

Our objective was to analyze the inhibitory effect of themedial olivocochlear system (MOCS) with transient otoa-coustic emissions (TEOAE) responses in a riverside populationexposed to environmental mercury. To date,this survey isunprecedented in a population exposed to mercury.

Methods

BackgroundThe INPTAm (Instituto Nacional de Pesquisa Translacional daAmazônia) is formed by a group of researchers interested inthe health of the environment of the Amazon and its impli-cations for the health of the local population, conductingresearch in several areas and expecting aggregate knowledgeof suitable proposals for improving the quality of life ofriverside populations of the Amazon. The INPTAm also worksto transfer the knowledge to local professionals, recovery, andenvironmental protection.

The area selected for the proposed research was thecommunity of Lago do Puruzinho, located in Amazonas state,Brazil. This population was selected for the study because thepeople are subsistence farmers who are exposed to environ-mental methylmercury from fish, the primary source ofanimal protein in their diet.

This study was conducted according to the ethical aspectsrecommended by Resolution 196/96 of the National HealthCouncil on studies involving humans, and informed consentwas obtained from the research subjects. The research wasapproved by the Ethical Committee (Institutional ReviewBoard/EC) from the Instituto de Estudos em Saúde Coletiva(IESC), under number 79/2011.

All of theparticipantswerebornand lived inPuruzinho.Noneof the participants had a history of any type of auditory problemor otologic surgery. During the study period, the total populationof Puruzinho was 102 people. Of these, 83 agreed to participatein the study. Among those who agreed to participate, 25 did notmeet the inclusion criteria. The inclusion criteriawere as follows:absence of otologic alteration, confirmed by otolaryngologicevaluation and hearing thresholds � 20 dB hearing level deter-mined by pure tone audiometry (0.5 to 8 kHz in octave inter-vals). Children who did not undergo audiometry were includedin the study because they had TEOAEs, type A tympanograms(peak pressure between þ20 and �40 daPa; admittance


Mercury Exposure in a Riverside Amazon Population Hoshino et al.136

between 0.3 and 1.2 mL), and presence of contralateral acousticreflexes. Individuals who showed an absence of TEOAEs or ahearing loss (at any level, of any type, and affecting either ear)were excluded from the study.

The studywas conducted among 58 individuals (30 femalesand 28 males) with a mean age of 17.3 years (range 1 to 47).The purpose of the research was to evaluate the entirecommunity independently of variables of sex and age. Afterotolaryngologic evaluation, participants had tympanometryand investigation of contralateral acoustic reflexes (AT-235Interacoustics, Denmark, Middelfart), pure tone audiometry(AC-33 Interacoustics), and recording of TEOAEs (ILO v6, dualchannel, Otodynamics, London, United Kingdom) with nonlin-ear click stimulation (260 sweeps) presented at 80 � 2 dBpeak sound pressure level (SPL). We considered that otoacous-tic emissions were present when a signal-to-noise ratio above3 dB occurred in at least three frequency bands. The tests,including tonal audiometry, were performed in a very quiet,althoughnot acoustically treated, room. The earphones used inpure tone audiometry were AUDIOCUP (Amplivox Ltd, Oxford-shire, UK) (noise-reducing headset enclosures).

To evaluate the inhibitory effect of the MOCS, a test oflinear click-evoked otoacoustic emissions at 60 � 2-dB peakSPL was performed, with and without the contralateralpresentation of white noise at 60 dB SPL (contralateralacoustic stimulation, CAS). White noise was generated bythe equipment and delivered in the contralateral ear by asecond channel. The presentation mode was alternate, with10 seconds of TEOAE recording collected without CAS beingfollowed by 10 seconds collected with CAS. The alternatingpattern was continued until 260 sweeps in each conditionwere completed. The final result was the sum of the alternateresponses obtained in each condition. The inhibitory effectwas calculated from the response values by subtracting thevalue obtained in the condition with contralateral noise fromthe value in the condition without contralateral noise (inhib-itory effect ¼ TEOAE without CAS � TEOAE with CAS). Aninhibitory effect was considered to be present when theresult showed a positive value.

The collection of mercury from hair samples was con-ducted by professionals from the Universidade Federal deRondônia. Hair samples underwent digestion and chemicaloxidation with nitric acid (HNO3) and potassium perman-ganate (KMnO4). A total mercury assay was conducted atthe Universidade Federal de Rondônia, utilizing an atomicabsorption spectrophotometry technique through genera-tion of cold vapor (Flow Injection Mercury System, FIMS-400 by Perkin Elmer, Waltham, Massachusetts). Themercury assay was performed on 38 individuals who con-sented to participate in the study. The mean mercury levelin the hair samples was 11.26 µg/g (range 2.93 to 23.45).Participants were classified into three groups according todetected levels of mercury: group 1, up to 6 µg/g; group 2,from 6.1 to 14.9 µg/g; and group 3, �15 µg/g.

For the statistical analysis, the measures of central ten-dency and dispersion were calculated for all the numericalvariables, and the measures of frequency and percentagefrequency were calculated for all the categorical and ordinal Table

1Measuresof

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denc

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ission

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theleft

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,Brazil,

2012

Nonlin

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Linea

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twithoutnoise,

RE

Linea

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sien

twithnoise,

RE

Difference

linea

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dwithout

noise,

RE

Linea

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sien

twithoutnoise,

LELinea

rtran

sien

twithnoise,

LEDifference

linea

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sien

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dwithout

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LE

N58

5647

4747

4949

49

Mea

n14

.20

13.00

9.34

8.13

1.20

7.14

5.88

1.26

Med

ian

13.80

12.70

9.20

8.10

1.20

5.90

4.40

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Stan

dard

deviation

4.87

4.55

5.57

5.74

1.03

4.71

4.77

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0.40

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22.80

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5.30

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Mercury Exposure in a Riverside Amazon Population Hoshino et al. 137

variables. The comparison between the discrete variablesnoted in tables was calculated by the chi-square test withYates’ correction. In the correlation analysis, the Pearson testwas used to measure the linear association between twovariables. The comparison of means was performed by un-paired Student t test. The data were analyzed using theStatistical Package for the Social Sciences program (SPSSversion 14.0, IBM, NY, USA). The level of significance wasset at p < 0.05.

Results

The presence of nonlinear TEOAEs was observed in all re-search participants for the right ear and in 56 participants forthe left ear (►Table 1). It is important to emphasize that, inthe absence of response, the affected side was excluded forevaluation of TEOAEswith linear click stimuli. Among 58 rightears, 47 showed linear TEOAEs for stimulations of 60 dB SPL(sound pressure level) with and without contralateral noise,and thesewere considered for analysis. Among 56 left ears, 49

showed linear TEOAEs and were considered for analysis. Themean linear TEOAEs were 9.34 dB for the right ear and7.14 dB for the left ear. The mean value of the inhibitoryeffect of the MOCS was 1.20 dB for the right ear and 1.26 dBfor the left ear.

There was no significant correlation between the values ofthe inhibitory effect for the right and left ears and age (►Fig. 1).Furthermore, there was no significant correlation betweenvalues of the inhibitoryeffect andhairmercury levels (►Fig. 2).

Among the 58 participants, 7 (12%) showed an absence ofinhibitory effect of the MOCS. The mean level of mercury inthe hair of these individuals was 12.78 µg/g. Five participantshad hair mercury levels above the acceptable limit accordingto theWorld Health Organization (6.0 µg/g). Two participantswere not tested (►Table 2).

►Table 3 shows participants classified into three groupsaccording to their hair mercury levels. Individuals whoshowed altered results of the inhibitory effect of the MOCSin both right and left ears were excluded from each group(groups 1, 2, and 3). The mean amplitude of the inhibitory

Fig. 1 Inhibition of medial olivocochlear system values of the right ear and left ear compared with ages of the population from PuruzinhoCommunity, Amazon, Brazil, 2012.

Fig. 2 Inhibition of medial olivocochlear system values of the right ear and left ear compared with hair mercury levels in the population ofPuruzinho Community, Amazon, Brazil, 2012.



effect of the MOCS was lower for participants with higherconcentrations of mercury in their hair.

Discussion

The same team that analyzed the mercury levels for thisproject also analyzed hair mercury levels of the Puruzinhopopulation in 2005 to 2006.5 Themedian values in the femaleand male populations were 12.93 µg/g (range 2.48 to 57.04)and 18.41 µg/g (range 2.28 to 70.56), respectively. In thepresent study, themedian valuewas 10.91 µg/g (range 2.93 to23.45). The population remains exposed to high levels ofmethylmercury that are comparable to the historical greatexposures of populations in other parts of the world.

Several studies concerning the inhibitoryeffect of the efferentpathway demonstrated an asymmetry between the right andleft ears, suggesting an advantage of effectiveness of the right earand dominance of the left hemisphere.11 The inhibitory effect ofthe MOCS and the measure of the TEOAE amplitude in childrenand adults are valid indicators of peripheral auditory lateraliza-tion; however, they are independent.12 Such asymmetrymay berelated to the effectiveness of the efferent system in processingacoustic signals and, consequently, to its role in auditory perfor-mance with background noise. The Puruzinho populationshowed higher amplitude of responses evaluated by the TEOAEsfor the right ear; however, the inhibitory effect of the efferentpathway was equivalent for both sides, and an advantage of theright ear was not seen, contrary to other studies.

We observed that the amplitude of responses of the rightand left ears showed a tendency to decrease as the mercury

levels increased significantly. A previous study reported thathigher biological marker values are associatedwith increasedpresence of signals and symptoms of exposure.13

The inhibitory effect values of the MOCS in the Puruzinhopopulation did not present a significant correlation with age.This may be because the Puruzinho population is a youngpopulation, with a mean age of 17.3 years and a maximumage of 47 years.

It was not possible to correlate mercury levels with theinhibition of otoacoustic emissions; similarly, it wasnot possible to conclude that the 12% (7 participants) of thepopulationwho showed absence of the inhibitory effect of theMOCS had alterations arising from exposure to methylmer-cury. Population studies concerning the inhibition of otoa-coustic emissions cannot be found in the literature.

The population presented high levels of long-term expo-sure to methylmercury. The lack of a significant relationshipbetween the results of the auditory evaluation and hairmercury levels at the time the hair samples were collectedshould not be interpreted as implying that mercury does notalter the efferent auditory system. For this analysis, a longi-tudinal monitoring of the population and a higher number ofindividuals participating in the research would be necessary.

The pathophysiological effects of chronic exposure may besubtle and nonspecific and have a long period of latency14;therefore, it will be important to monitor the effects ofmercury exposure in the central auditory system of thePuruzinho population over time.

Limitations can be pointed out for this study: the numberof residents that agreed to participate in the study; the young

Table 2 Amplitude values of linear TEOAE, inhibitory effect, and mercury levels for participants with altered examinations from thepopulation of Puruzinho Community, Amazon, Brazil, 2012

Patient no. Linear TEOAEwithout noise(dB), RE

Linear TEOAEwith noise(dB), RE

Difference(dB)

Linear TEOAEwithout noise(dB), LE

Linear TEOAEwith noise(dB), LE

Difference(dB)

Mercurylevels (µg/g)

1 Excluded Excluded Excluded 3.80 3.80 0.0 ND

2 3.80 3.80 0.0 0.90 0.00 0.9 ND

3 8.20 8.10 0.1 3.10 3.40 �0.3 6.51

4 5.20 5.20 0.0 2.60 2.20 0.4 10.89

5 Excluded Excluded Excluded 3.30 3.90 �0.6 13.46

6 5.30 7.30 �2.00 2.10 1.50 0.6 15.53

7 11.40 9.80 1.60 7.40 7.40 0.0 17.53

Abbreviations: LE, left ear; ND, not determined; RE, right ear; TEOAE, transient otoacoustic emissions.

Table 3 Suppression mean distribution in individuals participating in the hair mercury level research across three groups

Average G1 0–6 µg/g Hg (n ¼ 10) G2 6.1–14.9 µg/g Hg (n ¼ 13) G3 � 15 µg/g Hg (n ¼ 10)

Hg 4.39 9.97 19.03

Suppression of RE 1.17 1.22 0.84

Suppression of LE 1.05 1.28 0.95

Abbreviations: LE, left ear; RE, right ear.Note: G1 had exposure within allowable limit according to the World Health Organization, G2 had exposure between 6.10 to 14.90 µg/g, and G3 hadexposure � 15 µg/g.


Mercury Exposure in a Riverside Amazon Population Hoshino et al. 139

population that prevented comparisons between age groups;and the difficulty in finding a control group that was notexposed to mercury in a riverside population.

Conclusions

The Amazon population, and in particular the riversidepopulation in which fish is a main source of dietary protein,should be clinically monitored. Some studies, taking intoaccount utilization of different methodologies, point towardthe possibility that the Amazon region is seeing the signifi-cant silent action of methylmercury in exposed riversidepopulations,14 with the presumable appearance of the clini-cal expression of mercury poisoning in the Amazon.

Although the literature refers to mercury as ototoxic,damaging the afferent auditory system, this study cannotclaim that methylmercury alters the function of the MOCS.Existing studies indicate that the inhibitory effect of theMOCS is a promising tool for evaluation of the efferentauditory pathways, and its specificity is very important fordiagnosing central auditory disorders.

The present study proposes that longitudinal monitoringstudies of the riverside population should be performed to learnwhether analysis of the inhibitory effect of the MOCS bymeasuring otoacoustic emissions may be used as an evaluationmethod and diagnostic tool in populations exposed to mercury.

It is important to emphasize that this study is an initial andexploratory study; therefore, the results may be used as areference for new studies of other populations exposed tomethylmercury and may help to open new frontiers formultidisciplinary research.

Declaration of InterestThe authors report no conflicts of interest.

FundingThis work was supported by the INCT for TranslationalResearch in Health and Environment in the AmazonRegion (Public notice MCT/CNPq no 015/08–NationalInstitutes of Science and Technology).

AcknowledgmentsI would like to thank Dra. Denise Carvalho (INPETAM) andDra. Jaqueline Quintanilha Brucha (UFRJ) for the scientificcontributions.

References1 Dias ACL, Guimarães JR, Malm O, Costa PA. Mercúrio total em

músculo de cação Prionace glauca (Linnaeus, 1758) e de espadarteXiphias gladius Linnaeus, 1758, na costa sul-sudeste do Brasil esuas implicações para a saúde pública. Cad Saude Publica 2008;24(9):2063–2070

2 Castoldi AF, Johansson C, Onishchenko N, et al. Human develop-mental neurotoxicity of methylmercury: impact of variablesand risk modifiers. Regul Toxicol Pharmacol 2008;51(2):201–214

3 Passos CJS, Mergler D. Human mercury exposure and adversehealth effects in the Amazon: a review. Cad Saude Publica 2008;24(4, Suppl 4):s503–s520

4 OPAS/OMS. Cooperación técnica entre Brasil, Bolivia e Colômbia:teoría y prática para el fortalecimiento de la vigilancia de saludde poblaciones expuestas a mercúrio. 2011. Available at: http://www.paho.org/bra/index.php?option¼com_docman&task¼doc_download&gid¼1428&Itemid¼423. Accessed October 21, 2013

5 Oliveira RC, Dórea JG, Bernardi JVE, Bastos WR, Almeida R,Manzatto AG. Fish consumption by traditional subsistence villag-ers of the Rio Madeira (Amazon): impact on hair mercury. AnnHum Biol 2010;37(5):629–642

6 Giordano G, Costa LG. Developmental neurotoxicity: some old andnew issues. ISRN Toxicol 2012:2012:814795

7 Hoshino ACH, Ferreira HP, Malm O, Carvallo RM, Câmara VM. Asystematic reviewofmercury ototoxicity. Cad Saude Publica 2012;28(7):1239–1248

8 American Academy of Audiology (AAA)—Position Statement andClinical Practice Guidelines. Ototoxicity monitoring. Octo-ber 2009. Available at: http://www.audiology.org/publications-resources/document-library/ototoxicity-monitoring Acessed inJanuary 2014

9 ATSDR—Agency for Toxic Substances and Disease Registry. Toxico-logical profile for mercury. Atlanta, GA: U.S. Department of Healthand Human Services, Public Health Service; 1999

10 ATSDR—Agency for Toxic Substances and Disease Registry. Tech-nical support document for a methylmercury reference dose as abasis for fish consumption screening values (FCSVs). [cited 2013Jan 31]. Available at: http://www.atsdr.cdc.gov/hac/pha/Methyl-mercuryReference/TechnicalSupportDocumentforAMethylcer-curyReferenceDoseasaBasisforFCSVs9-10-2009.pdf. Accessed inJanuary 17, 2012

11 Khalfa S, Collet L. Functional asymmetry of medial olivocochlearsystem in humans. Towards a peripheral auditory lateralization.Neuroreport 1996;7(5):993–996

12 Durante AS, Carvallo RMM, da Costa FS, Soares JC. [Characteristicsof transient evoked otoacoustic emissions in newborn hearingscreening program]. Pro Fono 2005;17(2):133–140

13 Choi AL, Weihe P, Budtz-Jorgensen E, et al. Methylmercury expo-sure and adverse cardiovascular effects in Faroese whaling menenvironmental. Environmental Health Perspectives 2009;117(3):367–372

14 Pacheco-Ferreira H. [Estudo dos efeitos a saúde em populaçõesribeirinhas expostas ao mercúrio na Amazônia] [PhD Thesis].Belém, Brazil: Universidade Federal do Pará; 2001



Lipidomic Profiling of Mastoid Bone and Tissuefrom Patients with Chronic OtomastoiditisFarbod Fazlollahi1 Kessiri Kongmanas2 Nongnuj Tanphaichitr2 Jeffrey Suh3 Kym Faull1

Quinton Gopen3

1Department of Psychiatry and Biobehavioral Sciences, UCLA, LosAngeles, United States

2Department of Biochemistry/Microbiology/Immunology, Universityof Ottawa, Ottawa, Ontario, Canada

3Department of Otorhinolaryngology–Head and Neck Surgery, UCLAHealth System, Los Angeles, California, United States


Address for correspondence Farbod Fazlollahi, BS, Department ofPsychiatry and Biobehavioral Sciences, UCLA, 760 Westwood Plaza,CHS 68-146, Los Angeles, CA 90095, United States(e-mail: [email protected]).

Introduction

Chronic otomastoiditis refers to an inflammatory conditioninvolving the middle ear and mastoid compartments. In the

United States alone, chronic otomastoiditis is estimated toaccount for over 30 million physician visits each year with anannual cost of roughly 3 to 5 billion dollars, and it remains acommon indication for antibiotic therapy and surgery.1

Keywords

► mastoiditis► lipid metabolism► mass spectrometry► flow injection analysis

Abstract Introduction Chronic otomastoiditis causes pain, otorrhea, and hearing loss resultingfrom the growth of tissue within the normally hollow mastoid cavity.Objectives In this report, we used a lipidomics approach to profile major mastoid boneand tissue lipids from patients with and without otomastoiditis.Methods The bone dust created during mastoidectomy, as well as the mastoid tissue,was analyzed from seven patients. Bone dust was also collected and analyzed in anadditional four otologic cases (parotidectomy requiring mastoidectomy). Samples weresubjected to a modified Bligh/Dyer lipid extraction, then high-performance thin-layerchromatography (HPTLC), combined gas chromatography/electron impact-mass spec-trometry (GC/EI-MS), and flow-injection/electrospray ionization-tandem mass spec-trometry (FI/ESI-MSMS). Data were analyzed for identification and profiling of majorlipid components.Results HPTLC revealed the presence of various lipid classes, including phosphatidyl-cholines, cholesterol, and triacylglycerols. GC/EI-MS analysis revealed the presence ofcholesterol and several fatty acids. FI/ESI-MSMS analysis revealed a host of phospha-tidylcholines, phosphatidylethanolamines, and cholesteryl esters.Conclusion We used a lipidomics approach to develop an efficient (both in time andtissue amount) methodology for analysis of these tissues, identify the most abundantand common lipid species, and create a base of knowledge from which more focusedendeavors in biomarker discovery can emerge. In an effort toward improved patientcategorization and individualized intervention, the ultimate goal of this work is tocorrelate these lipid molecules to disease state and progression. This is the first reportedstudy of its kind on these tissues.

receivedSeptember 25, 2014acceptedNovember 6, 2014published onlineDecember 8, 2014



THIEME


Interestingly, this pathology is also found in a significantproportion of indigenous children from the United States,Canada, Northern Europe, Australia, and New Zealand, theimplication of which is obscure.2 Chronic otomastoiditis canresult in hearing loss, vertigo, facial paralysis, sigmoid sinusthrombosis,meningitis, increased intracranial pressure, brainabscess, and in rare cases can even lead to death. In thiscondition, the normally air-filled middle ear and mastoidcompartments become infiltrated and opacified by soft tis-sue–likematerial, which frequently requires surgical removalbecause resolution of symptoms is not achieved by antibiotic,anti-inflammatory, or other treatments.

When the surgically removed mastoid tissue is evaluatedby histologicmethods, chronic inflammation is almost alwaysseen as demonstrated by an infiltration of polymorphonucle-ar cells and lymphocytes as well as a deposition of densefibrotic tissue. Interestingly, standard swab and plate culturetechniques rarely identify any associated pathogens. Severalpossibilities have been proposed to account for the produc-tion of this material, including infectious, autoimmune, neo-plastic, and allergic processes. Despite the negative resultswith swab and plate techniques, an infectious etiology shouldnot be excluded as there are plausible explanations as to whypathogens have evaded detection. These include the presenceof biofilms, polysaccharide matrices created by bacteria thatprotect or shield them from the host response. Once envel-oped within the biofilm, bacteria are difficult to detect bystandard swab and plate techniques. The true relationshipbetween mastoiditis and this material is still unknown. Thiscontinues to be a significant barrier toward clinical progressfor treatment.

Lipids play crucial roles in cell, tissue, and organ physiolo-gy; not only do they provide an energy reservoir and serve asstructural components of membranes, but they also act assignaling molecules and hormone precursors. Consequently,their steady-state concentrations are under tight homeostaticcontrol,3,4 perturbation of which leads to disease and injury.Lipidomics aims to profile lipids such that the data sets can beused to compare control and disease states. Relating thelipidome to the proteome and the genome is an emergingfield that provides an experimental platform with whichcomplex biochemical systems can be related to genetic,developmental, and environmental influences; superim-posed over these factors, the effect of disease states can bestudied. With the ultimate goal of identifying lipid biomark-ers of otomastoiditis, the first step is to identify the majorlipid components of the involved tissues. In this report, weused high-performance thin-layer chromatography (HPTLC),combined gas chromatography (GC)/electron impact (EI)-mass spectrometry (MS), and flow-injection (FI)/electrosprayionization (ESI)-tandem mass spectrometry (MSMS) to qual-itatively identify the major lipid classes in the mastoid tissueand mastoid compartment bone dust (BD). A necessary firststep toward elucidating the composition and mechanismsbehind its production, this information is fundamental tofurther identification of clinical biomarkers for otomastoidi-tis, and subsequently efficient patient diagnosis andtreatment.

Methods

SolventsChloroform, methanol, and formic acid (FA) were purchasedfrom Fisher Scientific (San Jose, California, United States).Triethylamine (TEA), butylated hydroxytoluene (BHT), andammonium acetate (AA) were purchased from Sigma Aldrich(St. Louis, Missouri, United States).

Patients Recruitment and Biological Sample CollectionWith approval from and strict adherence to the MedicalInstitutional Review Board guidelines and regulations, sam-pleswere collected after written consent was obtained beforesurgery from patients with and without chronicotomastoiditis.

Recruited patients included those who showed clinicalsymptoms consistent with the diagnosis such as otorrhea,hearing loss, and otalgia. Furthermore, these patients showedthe presence of pathologic mastoid tissue, as revealed by acomputed tomography scan. Four additional cases of patientsundergoing mastoidectomy during exposure of the facialnerve for parotid tumors were recruited to participate inthis study and provide samples for the BD group.

Sample CollectionIn the operating room, mastoid tissue was excised into sterileplastic specimen cups, and BD was collected after drillingthrough the mastoid bone. Specimens were stored on ice fortransportation to the laboratory, where they were thentransferred to 1.5-mL microcentrifuge tubes, to which 500µL of ice-cold deionized water was added. Tubes were mixedbriefly to wash and remove excess blood, then centrifuged atlow speed (2,000g). The supernatant was discarded and theprocess was repeated twice. Samples were then stored at�80°C.

Sample ExtractionTo minimize hydrolysis or degradation, the workupwas doneon ice as swiftly as possible. Samples were thawed at roomtemperature and then placed on ice. A small piece of chronicotomastoiditis tissue (COT) from each patient was placed onan ethanol-cleaned watch glass, also on ice, and diced(roughly 500-µm2 pieces) with a no. 10 scalpel (sterilizedfor 30 minutes in ethanol) to aid in homogenization. The BDwas already amenable to homogenization and extraction, sothis consideration was unnecessary. To prevent oxidation,BHT (final concentration 300 µM) was added to all solutionsand solvents used throughout the lipid extraction procedures.Roughly 1 mg of tissue from each patient’s COT was pooledinto a glass test tube and roughly 20 mg of BD from eachpatient were pooled into another tube, and 1mL of deionizedwater containing BHT (dH2O þ BHT) was added to both. Thepooled samples were then homogenized while on ice, eitherwith an ultra-sonic cell disrupter (Branson, three 15-secondbursts with a 5-minute interval in between) or a BrinkmanPolytron Homogenizer (30 seconds).

After settling, 10 µL aliquots of the supernatants werediluted 1:100 in dH2O þ BHT and the optical density


Lipidomic Profiling of Mastoid Tissue Fazlollahi et al.142

determined at 417 nm against a water blank (Beckman DU640B spectrophotomer).5 A 60-µL aliquot of freshly drawnhuman blood was diluted in 1 mL dH2O þ BHT, sonicated asdescribed above, and the OD417nm determined. From acomparison of the absorption intensities, control humanblood samples were then prepared at an appropriate dilutionin dH2O þ BHT to match the hemoglobin, and theoretically,the blood content of each tissue sample.

The remainder of each homogenized suspension and thematched human blood samples (990 µL each) were thensubjected to a modified Bligh-Dyer lipid extraction.6,7 In anaqueous suspension, 1,237.5 µL of chloroform containing 300µM BHT (CHCl3 þ BHT) and 2,475 µL methanol containing300 µM BHT (MeOH þ BHT) were added to each sample toachieve a chloroform:methanol:water ratio of 1:2:0.8, v/v/v.Samples were mixed thoroughly, and after 30 minutes at 27°C, additional CHCl3 þ BHT (1,237.5 μL) and dH2O þ BHT(1,237.5 µL) were added to give the final chloroform:metha-nol:water ratio of 1:1:0.9, v/v/v. The samples were mixedthoroughly again, centrifuged (2,000g for 3 minutes), and thechloroform (bottom) phase was transferred with a glasssyringe to 15-mL glass capped test tubes. The extractionwas repeated once using a volume of CHCl3 þ BHT identicalto that removed in the previous step. The pooled chloroformphases were dried under a gentle stream of dry nitrogen gas,capped, and stored at �20°C.

High-Performance Thin-Layer ChromatographyHPTLC was performed to elucidate the complexity of the tissuesamples and broadly identify major lipid classes.7 Each samplewas redissolved in 100 µL of chloroform, from which a 5-µLaliquot was loaded as a 0.8-cm band onto a prewashed andpreactivated HPTLC plate (HPK silica gel 60 Å, 200-µm thick-ness, 10 � 10 cm, Whatman, Kent, United Kingdom), pre-washed in chloroform-methanol 1:1 (v/v), dried, andimmediately prior to use heat-activated at 100°C for 1 hour,7

alongwith lipid standards as a separate band (lactosylceramide[CerLac, 2 µg; Sigma Aldrich], triacylglycerol [TAG, 5 µg; SigmaAldrich], phosphatidylcholine [PtdCho; 5 µg; Avanti PolarLipids, Alabaster, AL], cholesterol [C, 5 µg; Avanti Polar Lipids],and sphingomyelin [CerPCho, 5 µg; Sigma Aldrich]). Afterdeveloping in chloroform/methanol/water (65:25:4, v/v/v),the platewas sprayedwith orcinol (0.02% in 50% H2SO4; SigmaAldrich) and heated (120°C, 2 to 3 minutes). An image of thestained platewas recorded digitally. The plate was then furtherstained with Coomassie Brilliant Blue G-250 (0.03% in 30%methanol/100 mM NaCl; Biorad, Hercules, CA) and destainedwith methanol (30% in 100 mMNaCl).8,9 Another image of theplate was recorded digitally.

The remaining 90 µL of each samplewas loaded as an 8-cmband onto a longitudinally prescored HPTLC plate and devel-oped as described above. A narrow strip of this plate (5%) wasdetached (along the score) and stained with orcinol asdescribed above. Based on the orcinol staining pattern theunstained part was divided into four zones (zone 1, relative tofront (Rf) 0.0 to 0.24; zone 2 Rf 0.24 to 0.47; zone 3 Rf 0.47 to0.71; zone 4 Rf 0.71 to 1.0), and the silica from each wasscraped into a clean glass tube (one zone/tube). The scrapings

were treated with chloroform/methanol (1 mL, 1:1, v/v), andafter centrifugation (2,000g, 5 minutes) the supernatantswere collected into a new glass tube and the silica pelletwas re-extracted once. The pooled supernatants weresubjected to a Bligh-Dyer extraction by the addition of water(0.9 mL), the phases separated by centrifugation (2,000g,5 minutes) and the chloroform phases were removed to cleanglass vials. The Bligh-Dyer extraction was repeated once andthe pooled chloroform phases were dried under a stream ofnitrogen gas, resulting in four HPTLC fractions per sample.The dried samples were stored at �20°C.

Gas Chromatography/Mass Spectrometry of the High-Performance Thin-Layer Chromatography ZonesEach lipid fraction extracted from the HPTLC plate wasredissolved in chloroform:methanol (500 μL,1:1). Fiftymicro-liters of each sample were transferred to glass conical GCinjector vials and dried under a nitrogen stream. Each samplewas then treated with 50 µL N,O-bis(trimethylsilyl)trifluor-oacetamide containing 10% (v/v) trimethylchlorosilane andincubated (60°C, 60 minute) to convert all carboxyl, amino,and hydroxyl functional groups to their corresponding tri-methylsilyl derivatives.10,11 The derivatized samples werethen placed in an autosampler (Agilent 7683 series, FosterCity, California, United States) from which an aliquot of each(1 μL, equivalent of 0.2% of the contents of the HPTLC fraction)was injected onto a bonded-phase nonpolar fused silicacapillary column (Phenomenex ZB-5, phenyl/dimethylpoly-siloxane 5/95, 60 m � 0.25 mm, 0.10-μm film thickness,Phenomenex, Torrance, California, United States; injectorport 250°C) and eluted (constant flow, 1 mL/min) withultra-high-purity helium (Agilent 6890A GC system) over a63-minute temperature ramp (min/°C: 0/50, 3/50, 53/300, 63/300). The end of the column (GC/MS transfer line at 250°C)was directly inserted into the EI source (180°C, 70 eV) of atime-of-flight mass spectrometer (Waters GCT, calibratedwith perfluorotributylamine immediately prior to the analy-sis of each batch of samples) scanning from m/z 40 to 650(0.9 s/scan at a resolution [full width at half maximum] of7,000, lock-mass from column bleed at 207.0329C5H15Si3O3) with a 15-minute solvent delay. Data werecollected with instrument manufacturer-supplied software(MassLynx version 4.1, Waters Corporation, Milford, MA).Each sample was individually prepared and run in duplicate.

Identifications were based on comparison of spectra aver-aged over the width of the GC peaks within the total ionchromatogram, with background subtraction, to the NationalInstitute of Standards and Technology (NIST) 2008 MassSpectral Library (version 2.0f). These were based on NISTmatch factors of at least 750, indicating strong concordancebetween the unknowns and the library spectra, and acceptablevisual concordance between the unknown and library spectra.

Electrospray Ionization/Tandem Mass SpectrometryTo screen for the major lipid classes on the basis of tandemmass spectra, identical lipid extracts were prepared as de-scribed above and dissolved in chloroform:methanol (500 μL,1:1). To provide the appropriate milieu for effective


Lipidomic Profiling of Mastoid Tissue Fazlollahi et al. 143

ionization, a 50-μL aliquot of each stock solution was diluted1:5 (v/v) in chloroform:methanol:formic acid (200 μL,50:50:0.1, v/v/v). Another 50 μL of each stock was dilutedsimilarly in chloroform:methanol:triethylamine (200 μL,50:50:0.1, v/v/v), and a third 50-μL aliquot was dilutedsimilarly in chloroform:methanol:ammonium acetate (200μL, 50:50:10 mM, v/v/c). Ten microliters of each of thesedilutions, from each sample, were analyzed on an Agilent6460 triple quadrupole mass spectrometer by flow injectioninto an Agilent Jet Stream ion source (gas temperature 300°C,gas flow 6 L/min, nebulizer pressure 45 psi, sheath gastemperature 50°C, sheath gas flow rate 10 L/min, capillaryvoltage 4,500 V, nozzle voltage 2,000 V) with chloroform:methanol (50:50) as the running solvent (100%, 0.050 mL/min). Data were collected with instrument manufacturer-supplied software (Agilent Mass Hunter version B.05.00).

Using published methods as a guide,12–21 optimal con-ditions for precursor, product, and neutral loss ion scans forthe PtdCho/CerPCho, cholesteryl esters (CE), and sulfatedlipids (S) were developed in-house using authentic standards(►Table 1). Optimization for each compound class was per-formed with solutions prepared in chloroform:methanol(50:50, v/v), 1 pmol/μL using 10-μL injections, containingeither 10 mM AA (CE), 0.1% TEA (phosphatidyl serine[PtdSer], phosphatidyl inositol [PtdIns], and S), or 0.1%FA (PtdCho/CerPCho and phosphatidyl ethanolamine[PtdEtn]).12 The standards utilized for this purpose were:PtdCho, diheptanoyl phosphatidylcholine and 1-heptadeca-noyl-1-lyso-phosphatidylcholine (Sigma Aldrich); PtdIns,L-α-phosphatidylinositol sodium salt from Glycine max (soy-bean; Sigma Aldrich); PtdSer, 3-sn-phosphatidyl-L-serinesodium salt from bovine brain (Sigma Aldrich); Cer, ceramidefrom bovine spinal cord (Sigma Aldrich); PtdEtn, 3-sn-phosphatidylethanolamine from bovine brain (SigmaAldrich). Instrument parameters (collision energy, fragmen-tor, and cell accelerator voltage) were manually tuned withrepeat injections until maximal signal intensity for precur-sors ions were obtained in each scan modality. Limits ofdetection for these compounds were then determined by

preparing serial dilutions of the standard solutions andrunning them at the aforementioned optimized conditionsuntil the dilution at which a signal 3 times as intense asbackground was determined. Optimized instrument param-eters and limits of detection for CE and S were previouslydetermined on this instrument via similar means.22–24

Results

High-Performance Thin-Layer ChromatographyProfilesHPTLC was used to estimate the magnitude of blood contam-ination in the extracts. The OD417nm provided a convenientmethod for measuring hemoglobin, and therefore blood thatinevitably comprises a portion of each sample. The contribu-tion of blood to the lipid profiles of the tissue samples wasinsignificant (►Fig. 1). Co-spotted matched blood controlsamples for each tissue sample revealed relatively minorbands with the exception of C and possibly TAG, althoughthe tissue samples were vastly stronger for these bands aswell.

HPTLC was also used to gauge the complexity of the lipidprofile in the samples and to screen for possible majordifferences between samples. Visual inspection of the platesrevealed multiple bands in the region of PtdCho, CerPCho,bands in the region of CerLac, relativelyminor bands betweenCerLac and C, and strong bands at the Rf of C and TAG(►Fig. 1). The results suggested that the samples were notsufficiently complex to warrant prefractionation ahead oftandem MS.

Gas Chromatography/Electron Impact-MassSpectrometry Profiles of High-Performance Thin-LayerChromatography ZonesThe total ion chromatograms revealed a peak for C in zone 4extracts from each of the sample groups (►Fig. 2), which wasoverloaded in the COTsample, and on-scale for the BD sample.In addition, less intense peaks were identified in both zone 4extracts for palmitic, oleic, and stearic acids. Glycerol and

Table 1 Scan modalities for tandem mass spectrometric screens of major lipid classes

Lipid class Dilutionused

Ions orneutral mass

Polarity Type ofscan

Fragmentor Collisionenergy

LOD(injected in 10 μL)

PtdCho/CerPCho FA m/z 184 Positive Precursor 100 20 450 fmol

PtdSer TEA 87 Negative Neutral loss 45 21 40 pmol

PtdEtn FA 141 Positive Neutral loss 141 10 5 pmol

PtdIns TEA m/z 241 Negative Precursor 200 50 50 pmol

Cer FA m/z 264.1and 265.3

Positive Precursor 150 30 100 pmol

CE AA m/z 369.4 Positive Precursor 80 7 –

S TEA m/z 97 Negative Precursor 190 85 200 fmol

Abbreviations: AA, ammonium acetate; CE, cholesteryl esters; Cer, ceramide; CerPCho, sphingomyelin; FA, formic acid; LOD, limit of detection;PtdCho, phosphatidyl choline; PtdSer, phosphatidyl serine; PtdEtn, phosphatidyl ethanolamine; PtdIns, phosphatidyl inositol; S, sulfated lipids; TEA,triethyl ammonia.Notes: LOD for CE not found due to high background at low concentrations. The LOD was determined from serial dilutions of standards prepared in-house, and the assigned value was the amount injected that gave a signal �3 times more intense than background.



phosphatewere also identified; however, their presence in allfour zones of both COT and BD relegated them to the status ofcontaminants. Based on an estimated limit of detection of1 ng injected, the inferred content of any free fatty acids in thesamples must be greater than�5.5 ng per mg of COT and 260pg per mg of BD.

Flow-Injection/Electrospray Ionization-Tandem MassSpectrometry ProfilesThe MSMS approach to identify the components of the majorlipid classes using flow injection with appropriately selectedscan modalities (►Table 1) proved to be a fast and efficient

screen. Using the redissolved Bligh/Dyer extracts, there wasno evidence of ion suppression as learned from repeatinjections of 10-fold concentrated samples from which therewas the expected increase in signal intensity. The lack of ionsuppression observed during flow injection validated thedecision to forgo chromatography. Consequently, the analy-ses were both faster and shorter, adding to the efficiency ofthis screening method. The criteria for selection of significantions from these spectra were that the same signal must bepresent in repeat injections from agiven sample, and the peakintensity must be at least fivefold larger than the localbaseline. Such stringent criteria were used to restrict thisreport to the most abundant species and keep the number ofassigned peaks to a manageable level. The estimated limits ofdetection (LOD) for the different lipid classes varied from 200and 450 fmol, to 5, 40, 50, and 100 pmol injected from S,PC/SM, PE, PS, PI, and C, respectively (►Table 1). Theseinjected LODswere determined by injection of serial dilutionsof the authentic standards as discussed above, and equate totissue LODs between 125 fmol/mg (S) and 62.5 pmol/mg(Cer). Ion assignments in the collected spectra were made asprotonated (MHþ) and ammoniated (M þ NH4þ) moleculeswhen FA and AA solvents were used, respectively.

The complexity of the spectra varied for the differentclasses, and representative MSMS spectra for the majorlypresent lipid classes (►Fig. 3) show the most abundantspecies in each. ►Tables 2, 3, and 4 show precursor ions forfour major lipid classes identified (PtdCho, PtdEtn, Cer, andCE, respectively) and the identity of themost intense ion fromeach sample. Signals for PtdIns, PtdSer, and S were notdetected in any sample.

The parents of m/z 184 positive ion scan modality, used toscreen for choline-containing lipids, relies on the facile pro-duction of the C5H15PO4Nþ fragment ion produced duringcollisionally activated dissociation (CAD) from all PtdChos bycleavage of the moiety attached to the sn3 position of theglycerol backbone.25 Even with stringent selection criteria, inFA, this scan modality revealed a complex array of signals inthe COT sample with more than 20 species present(►Fig. 3, ►Table 2, even-numbered m/z values). However,the same scan and selection criteria in FA solvent revealedonly two signals in the BD sample. The pair of ions at m/z 758and 760, presumably differing by a single double bond,dominated both profiles and are assigned as 34a:2 and34a:1 diacyl phosphatidylcholine, respectively. When identi-cal samples were diluted in AA solvent, the profile from COTwas even more complex, and astonishingly the profile fromBD was equally complex with 24 and 20 assigned signals(even-numbered m/z values), respectively. Sprinkled amongthe even m/z signals are a small number of odd massassignments.

The neutral loss of 141 Da positive ion scanmodewas usedto profile PtdEtns.21 This was based on the facile loss of theC2H8O4NP molecule from the protonated PtdEtn molecularions during CAD. COTrevealed up to 8 and 14 different speciesin FA- and AA-containing solvent, respectively(►Fig. 3, ►Table 3). BD revealed two and nine differentspecies in the same solvents, respectively. Themost abundant

Fig. 1 Orcinol-stained (top panel) and Coomassie blue-stained (bot-tom panel) high-performance thin-layer chromatography (HPTLC)displays of (left to right): 1, standards (5 μg/compound except CerLac 2μg); 2, mastoid tissue; 4, bone dust; 6, blank. Matched blood controlsfor each sample are in the adjoining lanes (for example, for lane 2 thematched blood control is in lane 3). The plates were run and firststained with orcinol and then Coomassie as described in the Methods.The HPTLC-purified samples that were separately prepared for massspectrometric analysis were: zone 1, Rf 0.0 to 0.24; zone 2 Rf 0.24 to0.47; zone 3 Rf 0.47 to 0.71; zone 4 Rf 0.71 to 1.0. Abbreviations: C,cholesterol; CerLac, lactosylceramide; CerPCho, sphingomyelin; TAG,triacylglycerol; PtdCho, phosphatidylcholine.



species in all samples were the pair of ions at m/z 768 and766, corresponding to 38a:4 and 38a:5 diacyl phosphatidyl-ethanolamine, respectively. The profiles also revealed a fewodd mass assignments.

The parents of m/z 369 positive ion scan modality in AA-containing solvent was used to profile CEs.24 This scanmodality was based on the facile fragmentation of the allCE (M þ NH4)þ parents yielding an intense ion for thecholesterol-containing fragment (C27H49) during CAD. COTand BD revealed 22 and 2 different molecular species, respec-tively, of which the signal at m/z 666 corresponding to thelinoleate ester was most intense in both (►Fig. 3, ►Table 4).

The scan modality for PtdSer, the neutral loss of 87 Da inthe negative ionmode inTEA solvent, was based on the loss ofthe serine functionality (C3H5NO2) from the (M-H)� parention common to these compounds.21 The scan modality forPtdIns was based on the parents of the common fragment ionat m/z 241 (C6H10O8P) in the negative ion mode in TEA

solvent formed in abundance under CAD conditions withthese compounds.26 The scanmodality for Cer, parents of m/z264, was based on the formation of a common positiverearrangement CAD fragment ion C18H34N displayed bythese compounds in FA solvent.27 The scan modality for Swas based on extensive in-house experience with theselipids28,29 in which the common negative fragment ion atm/z 97 (HSO4�) forms the basis of the scan in the negative ionmode in TEA solvent. Despite careful optimization of scanparameters for each of these modalities, using authenticstandards, signals for PtdIns, PtdSer, Cer, and S from COTand BD were below their respective limits of detection(►Table 1).

Discussion

There is no one technique capable of conveniently displayingall lipids in a sample. As such, we have used a combination of

Fig. 2 Combined gas chromatography/electron impact-mass spectrometry total ion current chromatograms from high-performance thin-layerchromatography (HPTLC) zone 4 extracts. Total ion current chromatograms of HPTLC zone 4 trimethylsilyl (TMS)-derivatized eluates from bonedust (top panel) collected during mastoidectomy and tissue excised from the mastoid cavity (bottom panel) in patients with chronicotomastoiditis. The dominant peak in each chromatogram (retention time 54.06 minutes) was identified as cholesterol. Other tentativeidentifications include: glycerol (20.54 minutes), palmitic acid (37.02 minutes), oleic acid (40.59 minutes), and stearic acid (41.04 minutes). Theregion from 15.00 to 20.00 minutes is populated by byproducts of the derivatization reaction and excess derivatizing reagents, and the large peakat 26.25 minutes in the bottom panel was identified at butylated hydroxytoluene. To aid in visualizing smaller peaks, the ordinate of eachchromatogram was regionally expanded as follows: top panel: 15.03 to 17.71 (�10), 17.79 to 19.89 (�24), 19.99 to 52.00 (�186), 54.50 to 63.00(�54); bottom panel: 15.03 to 17.51 (�16), 17.69 to 25.99 (�36), 27.49 to 47.50 (�54), 47.70 to 53.50 (�36), 54.50 to 63.00 (�24). The fattyacids observed in these chromatograms were present in abundance below the limit of confident identification, thus the spectra were not of highquality.



Fig. 3 Representative tandem mass spectrometry spectra of the majorly present lipid classes in mastoid tissue. (a) Precursor ion scan of m/z369.4 shows cholesteryl esters and similar compounds. (b) Precursor ion scan of m/z 184 of samples shows phosphatidylcholines, sphingomyelins,and similar compounds. (c) Neutral loss scan of m/z 141 of samples shows phosphatidylethanolamines and similar compounds. (d) Precursor ionscan of m/z 264.1 shows ceramides and similar compounds. Compounds not shown here were present below the limit of detection.



HPTLC, GC/EI-MS, and FI/ESI-MSMS to probe the lipidome ofthese pathologic tissues. The different techniques providecomplementary and partially overlapping profiles. To ourknowledge, this is the first study of its kind to evaluate thelipid characteristics ofmastoid bone and tissue, and it is a vitalfirst step toward elucidation of putative biomarkers forotomastoiditis. The HPTLC profiles of Bligh-Dyer extractedsamples provide convenient visualization of the major lipidcomponents in one display,with the singlemajor limitation ofan LOD of �0.5 µg/compound (1 nmol with a molecularweight of 500). HPTLC is therefore a convenient way to gaugethe complexity of any sample, and is particularly useful in thiscase where there is no precedence for analysis of middle earbone and tissues. In contrast, MS has unrivalled sensitivity

and specificity with the single major limitation being the lackof a scan modality that can be applied to all compounds.Because this work was not sample limited, we have used acombination of HPTLC and MS to provide a global qualitativeanalysis of the lipid components of bone and tissue from thetwo patient classes.

Inclusion of a blood control for each tissue type on theHPTLC chromatogram showed a relatively sparse bandingpattern, indicating an insignificant contribution of blood tothe tissue extracts. A variable blood contamination is un-avoidable when working with any tissue and bone samplesexcised from living patients.

The HPTLC profiles revealed a well-defined lipidome withsignificant quantities in the C- and choline-containing

Table 2 Tabulation of the major ions evident in the precursor of m/z 184 scan modality, characteristic of phosphatidylcholine-containing species

Tissue type Ion-pairreagent

Ionsa Identification of mostintense species

COT FA 496.4, 544.4, 703.5,b 720.3, 732.3, 734.6, 735.7,b

746.7, 758.5, 760.4, 768.4, 770.7, 772.2, 782.5,784.6, 786.6, 794.4, 796.4, 808.6, 810.5, 834.4, 836.8

34a:1 diacyl phosphatidylcholine

AA 520.4, 522.4, 544.1, 675.1,b 703.6,b 717.8,b 720.5,732.7, 734.5, 744.8, 746.6, 748.5, 758.6, 760.6, 768.4,782.6, 784.6, 786.6, 794.6, 796.3, 806.4, 808.5, 810.6,812.7, 832.6, 834.4, 836.5

34e:2 alkyl ether phosphatidylcholine

BD FA 703.6,b 758.4, 760.6 34a:2 diacyl phosphatidylcholine

AA 496.6, 520.4, 544, 675.1,b 676.3, 689.6,b 703.5,b 732.4,734.3, 744.1, 746.6, 748.3, 758.5, 760.5, 768.3, 782.5,784.5, 786.3, 794.2, 799.7,b 802.5, 806.4, 808,810.4, 813.8b

Unknown

Abbreviations: AA, ammonium acetate; BD, bone dust; COT, chronic otomastoiditis tissue; FA, formic acid.Notes: This listing includes only the signals that were more intense than fivefold above baseline. Close inspection of the spectra reveals many lessintense signals. The LOD was determined from serial dilutions of standards prepared in-house, and the assigned value was the amount injected thatgave a signal �3 times more intense than background.aMost intense shown in bold.bOdd-numbered m/z values.

Table 3 Tabulation of the major ions evident in the neutral loss of 141-Da scan modality, characteristic ofphosphatidylethanolamine-containing species

Tissuetype

Ion-pairreagent

Ions (most intense in bold font) Identification of most intense species

COT FA 740.5, 742.5, 744.5, 746.5, 747.6,b 764.5, 766.2, 768.5,769.5,b 770.6

38a:4 diacyl phosphatidylethanolamine

AA 716.4, 718.5, 740.5, 742.4, 744.5, 746.5, 754.5, 764.5,766.4, 768.4, 769.5,b 770.5, 792.5, 793.5,b 794.5, 796.6


BD FA 766.5, 768.2 38a:5 diacyl phosphatidylethanolamine

AA 716.5, 718.5, 740.5, 742.6, 744.5, 764.5, 766.5, 768.4,770.6


Abbreviations: AA, ammonium acetate; BD, bone dust; COT, chronic otomastoiditis tissue; FA, formic acid.Notes: This listing includes only the signals that were more intense than fivefold above baseline. Close inspection of the spectra reveals many lessintense signals. The LOD was determined from serial dilutions of standards prepared in-house, and the assigned value was the amount injected thatgave a signal �3 times more intense than background.aMost intense shown in bold.bOdd-numbered m/z values.



regions. By visual inspection, and considering that theamount of BD used in these analyses was 20 times by wetweight that of the tissue, BD had a lower lipid concentrationacross the chromatogram as compared with COT; however,there was no gross difference between the chromatograms,other than the difference in opacity attributable to therelatively higher richness of lipid content in tissue. Thismeans themajor lipid components of these tissues are similarin type, but not in amount. It is unlikely that a smallsubpopulation of each sample pool could dominate thechromatogram because each sample in each pool is presentin roughly equal quantities.

The GC/EI-MS lipidomic profiling of the trimethylsilyl-oximederivatives of Bligh-Dyer extracts of COT and BD samplesrevealed a similar lipid complexity; however, BD was compara-tively deficient in C. This phenomenon can likely be attributed tothe relatively higher concentration of C inherent to tissue ascompared with bone. The low abundance of free fatty acids is ofnote. These compounds are important components of interme-diary metabolism. Their low concentrations could be indicativeof a possible acellular nature of the samples.

The MSMS work provided a more sensitive and directedprofiling of the lipid content of these samples. The mostabundant lipids detectedweremembers of the PtdEtn, PtdCho,and CE classes. PtdChos are the principal phospholipids inhuman cell membranes and are a pulmonary surfactant.30

They are typically found on the exoplasmic side of the cellmembrane and play a role in membrane-mediated signaling.PtdChos are also mainly structural components of cellularmembranes, but also serve roles in intracellular signaling,differentiation, proliferation, and programmed cell death. Asexpected, a varietyof PtdChosweredetected inCOTwith eitherFA or AA solvents (►Table 2). The unexpected result was theemergence of signals from BD in AA solvent that were notobserved in FA solvent. This observation is of methodologicalsignificance relevant to future lipidomic screening experi-ments. There is considerable concordance between the signalsobserved for COT (either solvent) and BD (AA solvent). Appli-cation of the nitrogen rule to this data setmeans all quaternarynitrogen-containing species would be recorded at an even-numbered m/z. The relatively few odd-valued signals areunlikely to be choline-containing species, and unraveling theirstructures is beyond the scope of this work.

PtdEtns are found in all living cells and account for nearly aquarter of all phospholipids.31–33 In human physiology, they

are typically found in neural tissue, serving roles in mem-brane fusion and cytokinesis, lipoprotein release in the liver,and, pathologically, prion propagation without the involve-ment of other proteins or nucleic acids. In bacteria, however,PtdEtns are the principal phospholipids; therefore, theirrelative abundance (►Table 3) may be indicative of a bacterialpresence despite the failure of the swab and plate cultureexperiments previously referred to. Application of the nitro-gen rule to putative PtdEtns would mean the assigned parentions would be even when protonated (in FA solvent) and oddwhen ammoniated (only in AA solvent). The presence of thesame ion in both FA and AA solvents implies that at least someof the ions detected in AA solvent are protonated. The fewodd-value assignments are not likely to be PtdEtns.

Interestingly, CEs are among the major contributors tolocal lipidome in mastoid tissue and bone and are present inmuch greater abundance and variety in mastoid tissue. Asshown in ►Table 4, both cholesteryl arachidonate and cho-lesteryl linoleate are present in mastoid tissue, but onlycholesteryl linoleate is present both tissue types studiedhere. These findings could be potentially significant. Accord-ing to Do et al, cholesteryl arachidonate and cholesteryllinoleate show microbicidal effects in vitro, including againstPseudomonas aeruginosa,24 a common human pathogenwhose infection causes generalized inflammation and poten-tially sepsis. The differential presence of these antimicrobiallipids across the different tissue types analyzed and thesignificance of the variety of these lipids in the idiopathicmastoid tissue will be a topic of future study.

PtdSer, PtdIns, Cer, and S were present below the limit ofdetection in these samples (40 pmol, 50 pmol, 100 pmol, and200 fmol, respectively, injected; ►Table 1). This is an unex-pected result as PtdIns and PtdSer are relatively commoncellular membrane structural phospholipids and sulfatedlipids play roles in cell-surface signaling.34,35 The significanceof thisfinding is obscure at themoment. This is not to say thatthese lipid classes do not exist in mastoid tissue and/ormastoid bone, but rather their respective concentrations inthese tissues is low, and a more sensitive targeted assay forthem is required. The latter will be explored in future work.

Conclusions

These results represent a first step to characterize the lipidprofile of patients with otomastoiditis and to identify

Table 4 Tabulation of the major ions evident in the precursor of m/z 369 scan modality, characteristic of cholesteryl esters

Tissuetype

Ion-pairreagent

Ions (most intense in bold font) Name of mostintense species

COT AA 404.3, 614.6, 640.1, 642.6, 664.6, 666.6, 668.6, 670.6,690.5, 692.5, 694.7, 696.5, 714.8, 716.5, 718.5, 720.4,722.7, 744.6, 746.5, 748.8, 750.6, 778.5

Cholesteryl linoleate

BD AA 404.2, 666.6 Cholesteryl linoleate

Abbreviations: AA, ammonium acetate; BD, bone dust; COT, Chronic otomastoiditis tissue.Notes: This listing includes only the signals that weremore intense than 5-fold above baseline. Close inspection of the spectra reveals many less intensesignals. The limit of detection for this scan modality is listed in ►Table 1.



potential biomarkers of the disease. As there is a dearth ofliterature on lipidomic profiling in bone, this report sought todevelop an efficient and catholic analytical method for thesetissues. Given the large and diverse role of lipids in tissuestructure and the regulation of local homeostasis, the lip-idome of the mastoid compartment is a logical place to beginlooking for candidate biomarkers. The techniques developedin this studywill be exploited in further analyses of individualpatient samples in the search for relative abundance of thesemajor species as well as a broader global lipidomecomparison.

References1 Rovers MM, Schilder AG, Zielhuis GA, Rosenfeld RM. Otitis media.

Lancet 2004;363(9407):465–4732 Morris PS, Richmond P, LehmannD, LeachAJ, GunasekeraH, Coates

HL. New horizons: otitis media research in Australia. Med J Aust2009;191(9, Suppl):S73–S77

3 Ameer F, Scandiuzzi L, Hasnain S, Kalbacher H, Zaidi N. De novolipogenesis in health and disease. Metabolism 2014;63(7):895–902

4 de Kroon AI, Rijken PJ, De Smet CH. Checks and balances inmembrane phospholipid class and acyl chain homeostasis, theyeast perspective. Prog Lipid Res 2013;52(4):374–394

5 Luthra A, Denisov IG, Sligar SG. Spectroscopic features of cyto-chrome P450 reaction intermediates. Arch BiochemBiophys 2011;507(1):26–35

6 Bligh EG, Dyer WJ. A rapid method of total lipid extraction andpurification. Can J Biochem Physiol 1959;37(8):911–917

7 KatesM. Techniqueof lipidology: isolation, analysis and identificationof lipids. In: Burdon RH, ed. Laboratory Techniques in Biochemistryand Molecular Biology. New York, NY: Elsevier; 1986:100–278

8 Furimsky A, Vuong N, Xu H, et al. Percoll gradient-centrifugedcapacitated mouse sperm have increased fertilizing ability andhigher contents of sulfogalactosylglycerolipid and docosahexae-noic acid-containing phosphatidylcholine compared to washedcapacitated mouse sperm. Biol Reprod 2005;72(3):574–583

9 Iida N, Toida T, Kushi Y, et al. A sulfated glucosylceramide from ratkidney. J Biol Chem 1989;264(10):5974–5980

10 Fiehn O, Wohlgemuth G, Scholz M, et al. Quality control for plantmetabolomics: reporting MSI-compliant studies. Plant J 2008;53(4):691–704

11 Faull KF, Anderson PJ, Barchas JD, Berger PA. Selected ion monitor-ing assay for biogenic aminemetabolites and probenecid in humanlumbar cerebrospinal fluid. J Chromatogr A 1979;163(4):337–349

12 Pulfer M, Murphy RC. Electrospray mass spectrometry of phos-pholipids. Mass Spectrom Rev 2003;22(5):332–364

13 Hutchins PM, Moore EE, Murphy RC. Electrospray MS/MS revealsextensive and nonspecific oxidation of cholesterol esters in humanperipheral vascular lesions. J Lipid Res 2011;52(11):2070–2083

14 Leiker TJ, Barkley RM, Murphy RC. Analysis of diacylglycerolmolecular species in cellular lipid extracts by normal-phase LC-electrospraymass spectrometry. Int JMass Spectrom 2011;305(2–3):103–109

15 Murphy RC, Leiker TJ, Barkley RM. Glycerolipid and cholesterolester analyses in biological samples by mass spectrometry. Bio-chim Biophys Acta 2011;1811(11):776–783

16 Hsu FF, Turk J. Characterization of phosphatidylinositol, phospha-tidylinositol-4-phosphate, and phosphatidylinositol-4,5-bisphos-phate by electrospray ionization tandem mass spectrometry: amechanistic study. J Am SocMass Spectrom 2000;11(11):986–999

17 Gu M, Kerwin JL, Watts JD, Aebersold R. Ceramide profiling ofcomplex lipidmixtures by electrospray ionizationmass spectrom-etry. Anal Biochem 1997;244(2):347–356

18 Hsu FF, Turk J. Studies on phosphatidylserine by tandem quadru-pole and multiple stage quadrupole ion-trap mass spectrometrywith electrospray ionization: structural characterization and thefragmentation processes. J Am Soc Mass Spectrom 2005;16(9):1510–1522

19 Murphy RC, Axelsen PH.Mass spectrometric analysis of long-chainlipids. Mass Spectrom Rev 2011;30(4):579–599

20 Hsu FF, Turk J. Electrospray ionization/tandem quadrupole massspectrometric studies on phosphatidylcholines: the fragmenta-tion processes. J Am Soc Mass Spectrom 2003;14(4):352–363

21 Brügger B, Erben G, Sandhoff R, Wieland FT, Lehmann WD.Quantitative analysis of biological membrane lipids at the lowpicomole level by nano-electrospray ionization tandem massspectrometry. Proc Natl Acad Sci U S A 1997;94(6):2339–2344

22 Hsu FF, Bohrer A, Turk J. Electrospray ionization tandem massspectrometric analysis of sulfatide. Determination of fragmenta-tion patterns and characterization of molecular species expressedin brain and in pancreatic islets. Biochim Biophys Acta 1998;1392(2–3):202–216

23 Duffin K, Obukowicz M, Raz A, Shieh JJ. Electrospray/tandemmassspectrometry for quantitative analysis of lipid remodeling inessential fatty acid deficient mice. Anal Biochem 2000;279(2):179–188

24 Do TQ, Moshkani S, Castillo P, et al. Lipids including cholesteryllinoleate and cholesteryl arachidonate contribute to the inherentantibacterial activity of human nasal fluid. J Immunol 2008;181(6):4177–4187

25 Hunt AN, Postle AD. Mass spectrometry determination of endo-nuclear phospholipid composition and dynamics. Methods 2006;39(2):104–111

26 Sherman WR, Ackermann KE, Bateman RH, Green BN, Lewis I.Mass-analysed ion kinetic energy spectra and B1E-B2 triple sectormass spectrometric analysis of phosphoinositides by fast atombombardment. Biomed Mass Spectrom 1985;12(8):409–413

27 Liebisch G, Drobnik W, Reil M, et al. Quantitative measurement ofdifferent ceramide species from crude cellular extracts by electro-spray ionization tandem mass spectrometry (ESI-MS/MS). J LipidRes 1999;40(8):1539–1546

28 Marbois BN, Faull KF, Fluharty AL, Raval-Fernandes S, Rome LH.Analysis of sulfatide from rat cerebellum and multiple sclerosiswhite matter by negative ion electrospray mass spectrometry.Biochim Biophys Acta 2000;1484(1):59–70

29 Kongmanas K, Xu H, Yaghoubian A, et al. Seminolipid quantifica-tion by colorimetric assay and ESI-LC-MS/MS-multiple reactionmonitoring: compensation of seminolipid expression in Cgtþ/�mice. J Lipid Res 2010;51(12):3548–3558

30 Bernhard W, Hoffmann S, Dombrowsky H, et al. Phosphatidylcho-line molecular species in lung surfactant: composition in relationto respiratory rate and lung development. Am J Respir Cell Mol Biol2001;25(6):725–731

31 Vance JE, Tasseva G. Formation and function of phosphatidylserineand phosphatidylethanolamine in mammalian cells. Biochim Bio-phys Acta 2013;1831(3):543–554

32 Zemski Berry KA, Turner WW, VanNieuwenhze MS, Murphy RC.Characterization of oxidized phosphatidylethanolamine derivedfrom RAW 264.7 cells using 4-(dimethylamino)benzoic acid de-rivatives. Eur J Mass Spectrom (Chichester, Eng) 2010;16(3):463–470

33 Zemski Berry KA, Murphy RC. Electrospray ionization tandemmass spectrometry of glycerophosphoethanolamine plasmalogenphospholipids. J Am Soc Mass Spectrom 2004;15(10):1499–1508

34 Gardocki ME, Jani N, Lopes JM. Phosphatidylinositol biosynthesis:biochemistry and regulation. Biochim Biophys Acta 2005;1735(2):89–100

35 Takahashi T, Suzuki T. Role of sulfatide in normal and pathologicalcells and tissues. J Lipid Res 2012;53(8):1437–1450



Auditory Neuropathy/Dyssynchrony:A Retrospective Analysis of 15 CasesMurat Unal1 Yusuf Vayisoglu1

1Department of Otorhinolaryngology, Mersin University,Mersin, Turkey


Address for correspondence Yusuf Vayisoglu, MD, Department ofOtorhinolaryngology, Mersin University, Mersin 33079, Turkey(e-mail: [email protected]).

Introduction

Auditory neuropathy/dyssynchrony (AN/AD) is a hearingdisorder characterized by an absent or atypical auditorybrainstem response (ABR), with preservation of the co-chlear microphonics (CM) and/or otoacoustic emissions(OAEs).1,2 In 1996, Starr et al first described this rare entity,drawing on their observations in 10 patients.3 The authorssuggested that these patients were probably similar tothose previously reported cases with a paradoxicalabsence of ABRs and only a slight impairment of puretone thresholds but in whom CMs or OAEs had not beenrecorded.3–5 Also In 1998, Doyle et al reported eightpatients with normal transient evoked OAEs (TEOAEs)and distortion product OAEs (DPOAEs) combined withthe absence or marked abnormalities of ABRs.6 Starr et al

suggested that this type of hearing impairment is due to adisorder that impairs auditory nerve function and mayhave as one of its causes a neuropathy of the auditorynerve, occurring either in isolation or as part of a systemicneuropathic process.3 Clinically, the diagnostic criteria ofAN/AD is defined as (1) sensorineural hearing loss, usuallybilateral, of any degree; (2) normal outer hair cell functionas evidenced by the presence of OAEs and/or CM; (3) absentor atypical ABR; (4) understanding of speech worse thanwould be predicted from the behavioral or pure toneaudiometry; (5) absent acoustic reflexes to the ipsilateraland contralateral tones a 110-dB hearing level.1,7–9

In this retrospective study, we investigated the audiologi-cal findings, history, and clinical manifestations of patientsdiagnosed with AN/AD at our clinic.

Keywords

► auditory neuropathy/dyssynchrony

► otoacoustic emission► auditory brainstem

response

Abstract Introduction Auditory neuropathy/dyssynchrony (AN/AD) comprises a spectrum ofpathology affecting the auditory pathways anywhere from the inner hair cells to thebrainstem. It is characterized by an absent or atypical auditory brainstem response(ABR) with preservation of the cochlear microphonics and/or otoacoustic emissions(OAEs).Objective Retrospective analysis of patients with AN/AD.Methods Fifteen patients with AN/AD were included in this study and their recordswere retrospectively investigated.Results Possible etiology of AN/AD was neonatal hyperbilirubinemia in three patients,family history of hearing loss in three patients, consanguineous marriage in twopatients, head trauma in two patients, mental motor retardation in one patient,cerebrovascular disease in one patient, and there was no apparent cause in threepatients.Conclusion Otolaryngologists should keep in mind the diagnosis of AN/AD especiallyin patients complaining of difficulty in hearing and speech and audiological evidence ofdisassociation between pure tone and speech audiometry. ABR and OAE testing isrecommended in these patients for AN/AD diagnosis.

receivedJune 4, 2014accepted after revisionAugust 25, 2014published onlineNovember 28, 2014



THIEME


Patients and Methods

Fifteen patients with AN/AD were included in this study, andtheir records were retrospectively investigated. All of thepatients’ medical and otologic histories were recorded. Acomplete ear, nose, and throat examination was performed.Otoscopic examinations were done by an otolaryngologistbefore testing to rule out any external or middle ear patholo-gy that could affect audiometric measurements. Then puretone audiometry (250 to 8,000 Hz), tympanometry, andacoustic reflex measurement (500 to 4,000 Hz) were donein a standard fashion (Interacoustic AC 40 and AZ 26,Denmark, Assens). DPOAEs were measured at click levels of65 (L1) and 55 dB (L2) peak sound pressure for the F1 and F2components (Homoth Medizinelektronik GmbH&Co, KG,Germany, Hamburg). DPOAE-grams were recorded in one-quarter-octave steps over a frequency range of F2 from 0.5 to6 kHz. DPOAE values were plotted on a DPOAEgram, whichshows the emission level as a function of the F2 frequency.ABRs were recorded using Homoth ABR equipment.Electrodes were placed on the forehead as a ground electrodeand on both mastoids as active electrodes. The responseswere filtered with a bandpass of 100 to 3,000 Hz. Alternatepolarity clicks of 100-millisecond duration were presentedmonaurally with a repetition rate of 16.4/s. Each ear wastested separately, and all responses were replicated twice.Patients underwent assessment in a state of natural sleep. Inthis retrospective study, we did not include genetic research;we investigated only according to the history of patients.

Results

Eleven male and four female patients diagnosed with AN/AD(age range: 2 to 52 years and median age: 19.3 years) were

included in this study. Possible etiology of AN/ADwas neonatalhyperbilirubinemia in three patients. These patients had ahistory of exchange transfusion because of bilirubin levels over20 mg/dL. Three patients had family history of hearing loss,two patients had consanguineous marriage, two patients hadhead trauma, one patient had mental motor retardation(psychomotor retardation), one patient had cerebrovasculardisease, and there was no apparent causes in three patients.

Thirteen patients had bilateral and two patients hadunilateral AN/AD.

None of the patients with AN/AD had middle or inner earanomalies on computed tomography or magnetic resonanceimaging. ►Table 1 summarizes the patients’ demographic,clinical, and audiological features (see also ►Fig. 1 for a leftand right ear ABR and DPOAE recording of a case of AN/AD).

Eight of 15 patients’ pure tone audiometric results showedprofound hearing losses; however, seven of them had mild tomoderate hearing loss. Acoustic reflex tests were also absentin all of the patients.

Discussion

AN/AD is characterized by a unique pattern of hearing lossand distorted ABR with preservation of outer hair cell func-tion.10,11 AN/AD comprises a spectrum of pathology affectingthe auditory pathways anywhere from the inner hair cells tothe brainstem. Thus it is difficult to define the disorder ascochlear or retrocochlear. Increased clinical suspicion sup-ported by appropriate diagnostic tests is needed to establishan accurate diagnosis.12,13

The clinical findings for auditory neuropathy are associat-ed with several diagnoses including hyperbilirubinemia,neurodegenerative diseases, Charcot-Marie-Tooth syndrome,and other sensorimotor neuropathologies, mitochondrial

Table 1 Clinical and audiological features of the patients with AN/AD

Patient No. Age (y) Sex ABR DPOAE History

1 2 M Bilateral absent Bilateral present Consanguineous marriage

2 3y M Bilateral absent Bilateral present Mental motor retardation

3 52 F Bilateral absent Right ear (þ) Trauma

4 13 F Bilateral absent Bilateral present Blood exchange due to hyperbilirubinemia

5 2 M Bilateral absent Right ear (þ) No apparent cause

6 2 F Bilateral absent Bilateral present Blood exchange due to hyperbilirubinemia

7 7 F Bilateral absent Bilateral present Family history of hearing loss

8 4 M Bilateral absent Bilateral present No apparent cause

9 30 M Bilateral absent Bilateral present Trauma

10 5 M Bilateral absent Bilateral present Consanguineous marriage

11 2 M Bilateral absent Bilateral present Blood exchange due to hyperbilirubinemia

12 52 M Bilateral absent Bilateral present Family history of hearing loss

13 29 M Bilateral absent Bilateral present No apparent cause

14 6 M Bilateral absent Bilateral present Family history of hearing loss

15 50 M Bilateral absent Bilateral present Cerebrovascular disease

Abbreviations: ABR, auditory brain stem response; AN/AD, auditory neuropathy/dyssynchrony; DPOAE, distortion product otoacoustic emission.


Auditory Neuropathy/Dyssynchrony Unal, Vayisoglu152

disorders, and ischemic-hypoxic neuropathy resultingfrom asphyxia.14 Also, experimental animal models forauditory neuropathy have been proposed using the carbo-platin ototoxicity and ischemic-hypoxic neuropathy method-ologies.12,15,16 In our series, three patients with AN/ADhad neonatal hyperbilirubinemia, three patients had familyhistory of hearing loss, two patients had consanguineousmarriage, two patients had head trauma, one patient had

mental motor retardation, and one patient had cerebrovas-cular disease. We did not find any specific etiology in threepatients.

Recently twomechanisms havebeen proposed for explain-ing the abnormalities of auditory function: (1) impairedsynchrony among nerve fibers and/or (2) reduced neuralinput.17–19 It is generally thought that absent or severelydistorted ABR is highly likely to be related to impairment of

1 2 3 4 5 6 7 8 9 (ms) 1 2 3 4 5 6 7 8 9 (ms)RIGHT (lmp = 2485 Ohm) LEFT (lmp = 3115 Ohm)

30

80

70

60

50

F1 = 500Hz / 69.00 dBSPL F1 = 500Hz / 66.70 dBSPL

F1 = 625Hz / 60.90 dBSPLF1 = 625Hz / 60.80 dBSPL

F2/F1 = 1.25 F2/F1 = 1.25 M. duration = 1 min 21 sec 340 ms

DP1 = 1.21 dBSPL S/N = 8.54 dB (86%) 6.02 dB (97%) DP1 = -0.19 dBSPL S/N = 2.96 dB (86%) 0.42 dB (97%)

DP2 = -3.14 dBSPL S/N = 1.83 dB (86%) 0.35 dB (97%)DP2 = 15.50 dBSPL S/N = -7.09 dB (86%) -9.05 dB (97%)

M. duration = 1 min 21 sec 480 ms

DP

1 [u

nrea

dabl

e]/d

BS

PL

DP

1 [u

nrea

dabl

e]/d

BS

PL

35

30

25

20

15

10

5

0

-5

35

30

25

20

15

10

5

0

-5

1.000

2.000

3.000

4.000

5.000

6.0007.0008.0009.00010.000

1.000

2.000

3.000

4.000

5.000

6.000

7.0008.0009.00010.000

40

Fig. 1 Left and right ear auditory brain stem response and distortion product otoacoustic emission recording of a case of auditory neuropathy/dyssynchrony. Abbreviations: dBSPL, decibels sound pressure level; DP, distortion product; F1 and F2 used as formulation markers.


Auditory Neuropathy/Dyssynchrony Unal, Vayisoglu 153

neural synchrony in the auditory pathways. Similar to theprevious reports, we could not obtain any response to theclick stimulus.

The prevalence of AN/AD in patients with hearing lossranged from 0.5 to 15% according to studies in literature.10,11

Because there are so many possible causes for AN/AD, it isdifficult to estimate its exact prevalence.14 Davis and Hirshsuggested that 1 in every 200 hearing-impaired children hadABR findings inconsistent with pure tone findings.20 Perma-nent bilateral hearing loss is seen in 1.4 per 1,000 live births.Therefore the incidence of AN/AD is likely to be 1.4 per 10,000live births. According to Rance et al, AN/AD would be presentin 2.3 per 1,000 infants with risk factors for hearing loss. Thus,if OAE tests alone are used for hearing screening of infantswith risk factors, 11% of infants with permanent hearing losswill be missed.14,21 These data are important for designingthe newborn hearing screening protocols especially indeveloping countries.

Patients with AN/AD complain of hearing disability, espe-cially in the presence of noise, and tend to have word-recognition scores that are disproportionately poorer thanwould be expected by audiometric thresholds. In our series,all of the patients were admitted to our department with thecomplaint of hearing and speech disability. Only one of themhad understandable speech. Pure tone audiometric thresh-olds are variable in patients with AN/AD, and the degree ofhearing loss can range from mild to profound sensorineuraltype. Eight of our patients' pure tone audiometric resultsshowed profound hearing losses, and seven had mild tomoderate hearing loss. Acoustic reflex tests were also absentin all of the patients. Because OAEs and CM are dependent onthe integrity of cochlear outer hair cells and are “preneural”events, they may be present and normal in AN/AD; however,absent or grossly abnormal ABRs are seen.14,22 Similar find-ings were observed in our cases.

The hallmark of AN/AD is an abnormal ABR readingtogether with a normal OAE reading. However, there is alack of actual diagnostic procedures for AN/AD. Other testsmay also be used as part of a more comprehensive evaluationof an individual’s hearing and speech-perception abilities.Electrocochleography (ECoG), which objectively assesses co-chlear potentials, is the indicated clinical procedure to ana-lyze CMs. Although transtympanic ECoG yields recordingswith higher amplitudes and lower test–retest variability, ithas the disadvantage of being an invasive procedure. Extra-tympanic ECoG, therefore, is clinically more useful in thiscontext, supporting an audiological diagnosis and increasingknowledge about cochlear function in AN/AD. ECoG is themost appropriate procedure for assessing cochlear functionand helping identify CMs. CMs play an important role in thediagnosis of AN/AD. It is necessary and helpful to diagnose thesites of lesion in patients with AN/AD by analyzing thepatterns of CM amplitudes. Therefore, recently ECoG hasbeen used for diagnosing AN/AD.8,23

Some patients with AN/AD lost their OAE over the periodof time but there was no associated change in pure tonethresholds. It has also been reported that some patients withAN/AD do not have OAEs but rather evidence of hair cell

function was evident from CM. Therefore these authorssuggested that presence of CM with absent ABR seems tobe reliable criteria for diagnosing AN/AD.24,25

There has been controversy regarding whether to providehearing devices (hearing aids, personal radiofrequency[frequency modulation] systems, or cochlear implants) tochildren with AN/AD and whether to offer aural-oral orvisual-manual modes of habilitation.14,26 Berlin et al sug-gested that conventional amplification has little beneficialeffect on AN/AD patients.27 On the contrary, Cone-Wessonet al concluded that nearly 50% of children demonstrate somebenefit from the use of conventional hearing aids and that atrial of amplification is warranted.14,21 Cochlear implantationmay also be an option for hearing rehabilitation. Although theoutcome of cochlear implantation in children with AN/ADmight vary, it is favorable in most cases. Cochlear implanta-tion seems a justified hearing rehabilitation option forchildren with AN/AD and limited benefits from conventionalhearing aids.28–30 In our series, we obtained some develop-ment in hearing and speech abilities using a conventionalhearing aid in one case.

The current position statement of The Joint Committeeon Infant Hearing (2007) calls for (1) physiological hearingscreening of all infants before they are 1 month old, (2) con-firmation of the hearing loss before 3 months of age, and(3) commencement of an interdisciplinary interventionprogram before the infant is 6 months old. Moreover, thescope of disorders targeted for identification has beenexpanded to include neural hearing loss (especially AN/AD)in addition to sensorineural and permanent conductivehearing loss.31

Conclusion

Although AN/AD affects only a small portion of all personswith hearing loss, the infant, child, or adult with AN/AD isoften most disabled by the hearing disorder because of thelack of knowledge about its cause and, more importantly, itstreatment. Continued research regarding the causes andpathologies underlying this disorder is needed.10 However,it is also necessary to develop methods to reduce false-negative screening results and to provide accurate diagnosisfor the disorder.2 A combined OAE and ABR screening proce-dure may be considered to overcome the limitations of OAE-only procedures especially in high-risk infants. Also, it shouldbe kept in mind that adult AN/D cases may be related withsystemic neurological diseases.

References1 Madden C, Rutter M, Hilbert L, Greinwald JH Jr, Choo DI. Clinical

and audiological features in auditory neuropathy. Arch Otolar-yngol Head Neck Surg 2002;128(9):1026–1030

2 Lee JSM,McPherson B, Yuen KCP,Wong LLN. Screening for auditoryneuropathy in a school for hearing impaired children. Int J PediatrOtorhinolaryngol 2001;61(1):39–46

3 Starr A, Picton TW, Sininger Y, Hood LJ, Berlin CI. Auditoryneuropathy. Brain 1996;119(Pt 3):741–753


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4 Worthington DW, Peters JF. Quantifiable hearing and no ABR:paradox or error? Ear Hear 1980;1(5):281–285

5 Kraus N, Ozdamar O, Stein L, Reed N. Absent auditory brain stemresponse: peripheral hearing loss or brain stem dysfunction?Laryngoscope 1984;94(3):400–406

6 Doyle KJ, Sininger Y, Starr A. Auditory neuropathy in childhood.Laryngoscope 1998;108(9):1374–1377

7 Rapin I, Gravel J. “Auditory neuropathy”: physiologic and patho-logic evidence calls for more diagnostic specificity. Int J PediatrOtorhinolaryngol 2003;67(7):707–728

8 Shi W, Ji F, Lan L, et al. Characteristics of cochlear microphonics ininfants and young children with auditory neuropathy. Acta Oto-laryngol 2012;132(2):188–196

9 Roush P, Frymark T, Venediktov R,Wang B. Audiologicmanagementof auditory neuropathy spectrum disorder in children: a systematicreview of the literature. Am J Audiol 2011;20(2):159–170

10 Nikolopoulos TP. Auditory dyssynchrony or auditory neuropathy:understanding the pathophysiology and exploring methods oftreatment. Int J Pediatr Otorhinolaryngol 2014;78(2):171–173

11 Sinha SK, Barman A, Singh NK, Rajeshwari G, Sharanya R. Involve-ment of peripheral vestibular nerve in individuals with auditoryneuropathy. Eur Arch Otorhinolaryngol 2013;270(8):2207–2214

12 Sawada S,Mori N,Mount RJ, Harrison RV. Differential vulnerabilityof inner and outer hair cell systems to chronic mild hypoxia andglutamate ototoxicity: insights into the cause of auditory neurop-athy. J Otolaryngol 2001;30(2):106–114

13 Marco J, Morant A, Orts M, Pitarch MI, Garcia J. Auditory neuropa-thy in children. Acta Otolaryngol 2000;120(2):201–204

14 Cone-Wesson B, Rance G. Auditory neuropathy: a brief review.Curr Opin Otolaryngol Head Neck Surg 2000;8(5):421–425

15 Harrison RV. An animal model of auditory neuropathy. Ear Hear1998;19(5):355–361

16 Salvi RJ, Wang J, Ding D, Stecker N, Arnold S. Auditory deprivationof the central auditory system resulting from selective inner haircell loss: animalmodel of auditory neuropathy. ScandAudiol Suppl1999;51(Suppl 51):1–12

17 Kraus N, Bradlow AR, CheathamMA, et al. Consequences of neuralasynchrony: a case of auditory neuropathy. J Assoc ResOtolaryngol2000;1(1):33–45

18 Starr A, McPherson D, Patterson J, et al. Absence of both auditoryevoked potentials and auditory percepts dependent on timingcues. Brain 1991;114(Pt 3):1157–1180

19 Kırkım G, Serbetçioğlu MB, Ceryan K. Auditory neuropathy inchildren: diagnostic criteria and audiological test results. KulakBurun Bogaz Ihtis Derg 2005;15(1–2):1–8

20 Davis H, Hirsh SK. The audiometric utility of brain stem responsesto low-frequency sounds. Audiology 1976;15(3):181–195

21 RanceG, Beer DE, Cone-Wesson B, et al. Clinicalfindings for a groupof infants and young children with auditory neuropathy. Ear Hear1999;20(3):238–252

22 Maris M, Venstermans C, Boudewyns AN. Auditory neuropathy/dyssynchrony as a cause of failed neonatal hearing screening. Int JPediatr Otorhinolaryngol 2011;75(7):973–975

23 Anastasio ART, Alvarenga KdeF, Costa Filho OA. Extratympanicelectrocochleography in the diagnosis of auditory neuropathy/auditory dyssynchrony. Braz J Otorhinolaryngol 2008;74(1):132–136

24 Sanyelbhaa Talaat H, Khalil LH, Khafagy AH, Alkandari MM, ZeinAM. Persistence of otoacoustic emissions in childrenwith auditoryneuropathy spectrum disorders. Int J Pediatr Otorhinolaryngol2013;77(5):703–706

25 Mittal R, Ramesh AV, Panwar SS, Nilkanthan A, Nair S, Mehra PR.Auditory neuropathy spectrum disorder: its prevalence and audi-ological characteristics in an Indian tertiary care hospital. Int JPediatr Otorhinolaryngol 2012;76(9):1351–1354

26 De Leenheer EM, Dhooge IJ, Veuillet E, Lina-Granade G, Truy E.Cochlear implantation in 3 adults with auditory neuropathy/auditory dys-synchrony. B-ENT 2008;4(3):183–191

27 Berlin CI,Morlet T, Hood LJ. Auditory neuropathy/dyssynchrony itsdiagnosis and management. Pediatr Clin N Am 2003;50:331–340

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Auditory Neuropathy/Dyssynchrony Unal, Vayisoglu 155

Characterization of Hearing Thresholds from 500to 16,000 Hz in Dentists: A Comparative StudyClaudia Giglio de Oliveira Gonçalves1 Luciana Santos1 Diolen Lobato1 Angela Ribas1

Adriana Bender Moreira Lacerda1 Jair Marques1,2

1Post-Graduation Program in Communication Disorders,Universidade Tuiuti do Paraná, Curitiba, Paraná, Brazil

2Department of Audiology, Universidade Tuiuti do Paraná,Curitiba, Brazil


Address for correspondence Claudia Giglio de Oliveira Gonçalves,MSc, PhD, Universidade Tuiuti do Paraná, Rua Sidenei Antonio RangelSantos, 238 Curitiba, Paraná 82010330, Brazil(e-mail: [email protected]).

Introduction

Noise is present in different environments of human society,including the workplace, and may cause irreversible damage tothe human body.1 Loud noise is present in dentists’workplaces,whether from the clinical equipment used, such as dental drills,suction tubes, amalgamators, air compressors (when located inthe room), suction pumps, autoclaves, and air conditioners, or

from external sources, such as ambient noise including trafficfrom nearby vehicles and other urban noise.2–4

High-level noise exposure may damage the auditory sys-tem in dentists. Since the 1950s, some studies have shownhigh sound pressure levels in high-speed equipment indentists’ workplaces.5 In 1959, the American Dental Associa-tion recommended periodic audiological evaluations for den-tists due to noise exposure.6

Keywords

► noise induced► dentist► hearing loss

Abstract Introduction High-level noise exposure in dentists’ workplaces may cause damages tothe auditory systems. High-frequency audiometry is an important tool in the investiga-tion in the early diagnosis of hearing loss.Objectives To analyze the auditory thresholds at frequencies from 500 to 16,000 Hz ofdentists in the city of Curitiba.Methods This historic cohort study retrospectively tested hearing thresholds from500 to 16,000 Hz with a group of dentists from Curitiba, in the state of Paraná, Brazil.Eighty subjects participated in the study, separated into a dentist group and a controlgroup, with the same age range and gender across groups but with no history ofoccupational exposure to high levels of sound pressure in the control group. Subjectswere tested with conventional audiometry and high-frequency audiometry and an-swered a questionnaire about exposure to noise.Results Results showed that 81% of dentists did not receive any information regardingnoise at university; 6 (15%) dentists had sensorineural hearing impairment; significantdifferences were observed between the groups only at frequencies of 500 Hz and 1,000,6,000 and 8,000 Hz in the right ear. There was no significant difference between thegroups after analysis of mean hearing thresholds of high frequencies with the averagehearing thresholds in conventional frequencies; subjects who had been working asdentists for longer than 10 years had worse tonal hearing thresholds at high frequencies.Conclusions In this study, we observed that dentists are at risk for the development ofsensorineural hearing loss especially after 10 years of service.

receivedApril 1, 2014accepted after revisionJuly 23, 2014published onlineNovember 28, 2014




156

Many studies showed that dentists are at riskof developingnoise-induced hearing loss (NIHL).7–10 The damage caused byexposure affects the inner ear and causes an irreversiblehearing loss. The auditory thresholds show a classic sign ofNIHL in the audiometric notch at the frequencies in the rangebetween 3 and 6 KHz.11–13

Because early diagnosis of NIHL is prioritized, additionalaudiological tests are used to add value to basic audiologicalevaluations; among them, high-frequency audiometry (from8,000 Hz) is used.14 Studies show that high-frequency audi-ometry is an important tool in the investigation of the basalcochlear response as well as an instrument that helps in theearly diagnosis of NIHL.15–18 Research using high-frequencyaudiometry has suggested that with increasing noise expo-sure and age on dentists, there is a decrease in auditorythresholds.9,19–21

As a result of these considerations, this study aimed toanalyze the auditory thresholds at frequencies from 500 to16,000 Hz of dentists in the city of Curitiba.

Methods

This historic cohort study retrospectively tested hearingthresholds from 500 to 16,000 Hz in a group of dentistsfrom Curitiba, in the state of Paraná, Brazil. The study tookplace from August to December 2011.

The dentists’ time of professional practice was consideredin the analysis. The auditory threshold of dentists was alsocompared with a group of subjects with the same age rangeand gender, but with no history of occupational exposure tohigh levels of sound pressure (control group).

This study was approved by the Ethics Research Commit-tee, number 017/2008. The procedures were performed aftereach participant signed the informed consent form—TCLE(Resolution MS/CNS/CNEP, Number 196/96, October 10,1996). Subjects were excluded if they had compromisedmiddle ear or previous hearing damage.

To be included in the dentist group, the subject had to be anactive dentist. To be included in the control group, subjectshad to not be exposed to occupational noise and to be of thesame sex and age as one of the dentists for paired data.

The group of dentists was composed of 40 subjects,10 men and 32 women. The subjects’ ages ranged from 23 to61 years (mean 40.55 years, standard deviation 9.87 years,median 41.5 years) and time of practice was 1 to 39 years(mean 16.32 years and standard deviation 9.67 years). Sub-jects in the dentists’ groupworked 6 to 12 hours daily, but thisvariable was not considered in this study because accuratenoise exposure assessments in hours/years are difficult insuch cases. All participants were handed a questionnaireabout noise exposure.

The control group had a total of 40 subjects. This group wascomposed of students and subjects from different professionalactivities who did not present noise exposure. They wereall volunteers who were invited to participate in this study.For the analysis of thedata, each subject fromcontrolgroupwasmatched one-to-onewith someone from the group of dentists.

All 80 subjects submitted to ear canal inspection to ruleout possible obstructions that could compromise the hearingtest. All subjects were at acoustic rest for at least 14 hoursbefore audiometry.

Pure tone hearing thresholds were tested by conventionalaudiometry (frequencies from 500 to 8,000 Hz using a Mad-sen Itera II model audiometer (GN Otometrics Schaumburg,IL, USA), calibrated according to the International Organiza-tion of Standards 389/64 standards) and high-frequencyaudiometry with the same equipment but with HDA 200headphones (Sennheiser, Old Lyme, CT, USA) with a soundintensity in decibels hearing loss (dB HL), frequency 9,000 to16,000 Hz, calibrated according to the American NationalStandards Institute S3.6/69.

The hearing normality criterionwas defined for aerial tonehearing thresholds up to 25-dB HL for the frequency rate of500 to 8,000 Hz. In the case of changes in pure tone airconduction thresholds, pure tone audiometry was performedby bone conduction.22

For the analysis of high-frequency hearing thresholds (from9,000Hz), as there areno standardizednormal results,weusedthe group of subjects not exposed to occupational noise(control group) for comparison with the group of dentists.

Statistical methods that enabled the determination ofsignificant audiologic assessment between groups’ results

Table 1 Conventional hearing thresholds in the groups (n ¼ 80)

Frequency (Hz) Right ear Left ear

Dentists Controls p Dentists Controls p

Avg σ Avg σ Avg σ Avg σ

500 15.12 3.84 9.00 5.09 0.0000a 17.00 3.89 9.25 6.26 0.0000a

1,000 12.00 5.04 8.50 5.91 0.0056a 10.00 4.94 9.38 6.12 0.6164

2,000 9.25 6.36 7.38 5.43 0.1601 7.87 7.15 7.63 7.34 0.8778

3,000 7.38 7.59 6.75 6.06 0.6850 7.88 6.59 10.25 7.07 0.1241

4,000 9.62 9.63 7.75 7.07 0.3240 9.37 8.26 10.37 8.04 0.5847

6,000 18.63 12.86 12.75 9.27 0.0216a 19.25 11.30 15.38 7.02 0.0691

8,000 15.75 13.71 10.63 8.49 0.0478a 15.50 15.01 15.63 7.94 0.9630

Abbreviations: Avg, average; σ, standard deviation.aSignificant differences between the mean thresholds according to Student t test at a significance level of 0.05.


Hearing Thresholds in Dentists Gonçalves et al. 157

were used for data analysis, considering a 0.05 significancelevel (5%).

Descriptive statistics were applied. Student t test was usedto compare the thresholds of the groups and the analysis oftonal hearing thresholds as a relation of time of service inyears for dentists’ group.

Results

Forty dentists from Curitiba took the hearing tests andanswered a questionnaire about exposure to noise in theirworkplace. We observed that 81% of dentists did notreceive any information regarding noise during theiracademic training; however, 51% said they knew theeffects of noise on health. All (100%) acknowledged theexistence of noise in their workplace, and 54% believe thatnoise to be of medium intensity and 24% believe it to be ofloud intensity. Only two dentists used hearing protectiondevices.

Six (15%) dentists had sensorineural hearing impairment,only 1 (2.5%)with unilateral hearing loss (left ear) and 1 (2.5%)individual with hearing impairment among the group notexposed to occupational noise.

Auditory thresholds for the 40 dentists were analyzedand compared with the group of 40 subjects not exposedto occupational noise. Comparison of hearing thresholdsin conventional frequencies between groups is shownin ►Table 1. There were differences between the frequenciesof 500 Hz in both ears and in 1,000, 6,000, and 8,000 Hzfor the right ear; results were worse in dentists comparedto controls.

The results of pure tone hearing thresholds at higherfrequencies are shown in ►Table 2. No differences betweenthe groups for the high frequencies were observed. Tocompare conventional and high-frequency auditory thresh-olds, dentists were separated into two groups: those withover 10 years of experience and those with less than10 years, shown in ►Tables 3 and 4. Differences were notedbetween the frequencies of 2,000, 4,000, and 6,000 Hz forthe right ear, and the average conventional auditory tonethresholds were worse among those with more than10 years of experience.

There was no difference at all for frequencies in bothears, and those who had been working as dentists for longerthan 10 years had worse tonal hearing thresholds at highfrequencies.

Table 3 Conventional thresholds among dentists with time in service of up to 10 years (n ¼ 14) and over 10 years (n ¼ 26)


0–10 y Over 10 y p 0–10 y Over 10 y p


250 13.21 4.21 14.62 4.88 0.3705 18.21 5.41 15.19 5.00 0.0842

500 13.93 2.89 15.77 4.17 0.1503 17.50 5.10 16.73 3.14 0.5576

1,000 11.43 4.57 12.31 5.33 0.6051 10.36 4.14 9.81 5.38 0.7417

2,000 6.07 5.25 10.96 6.33 0.0183a 6.79 5.41 8.46 7.97 0.4868

3,000 4.29 4.32 9.04 8.49 0.0580 6.43 6.02 8.65 6.86 0.3147

4,000 4.29 3.85 12.50 10.61 0.0083a 6.43 5.35 10.96 9.17 0.0982

6,000 13.21 6.96 21.54 14.41 0.0495a 15.71 8.29 21.15 12.35 0.1487

8,000 10.00 6.79 18.85 15.51 0.0502 10.71 6.46 18.08 17.61 0.1411


Table 2 High-frequency hearing thresholds in the groups (n ¼ 80)


Dentists Controls p Dentists Controls p


9,000 20.13 16.96 17.37 16.41 0.4634 19.50 14.36 17.75 14.67 0.5913

10,000 22.00 18.36 20.50 17.39 0.7085 22.13 18.50 21.00 16.53 0.7750

11,200 25.63 18.75 22.38 20.41 0.4605 25.75 20.15 23.75 19.80 0.6555

12,500 24.25 22.49 22.88 22.73 0.7864 26.75 24.22 24.00 25.40 0.6216

14,000 26.13 21.50 30.00 23.53 0.4443 29.00 23.10 29.25 24.01 0.9623

16,000 34.63 21.73 33.13 21.41 0.7567 34.88 20.98 33.82 21.70 0.8271

Abbreviations: Avg, average; σ, standard deviation.


Hearing Thresholds in Dentists Gonçalves et al.158

Discussion

In researching information on the effects of exposure to noiseat work for dentists, it was observed thatmost had no trainingabout noise in their formal education and 50% of the subjectsknew the harmful effects of noise on health; however, onlytwo dentists claimed to use hearing protection device eventhough they recognized that there was noise at work. Suchfindings corroborate another study inwhich 48 dentists, ages22 to 55, participated, with only one reporting the use ofhearing protection.23

In another study on the perception of noise from dentists,49% of the 163 professionals surveyed felt that the noise intheir workplace was of medium intensity. Only 3% knew theeffects of noise on health and used hearing protection.10

Evenwith the presence of noise in the dentist’s workplace,a lack of information is still dominant regarding the educa-tional process for these professionals, and the lack of hearinghealth is shown in the low level of use of protection.

In the present study, we observed 6 (15%) cases of dentistswith sensorineural hearing impairment. Other studies havefound cases of hearing loss in dentists in various percentages.In a study conducted in the city of Cascavel, Paraná, with85 dentists between 25 and 60 years of age and of bothgenders, 43.5% were found to have audiograms suggestive ofNIHL.9 In another study of 198 dentists in the state of Paraná,27% had sensorineural hearing loss suggestive of NIHL.10

Another study in Recife of 50 dental professionals of bothgenders aged between 25 and 54, and with work experiencebetween 3 and 29 years, found 28 (56%) dentists had hearingloss.24 In a study in the city of João Pessoa with 48 dentists,conventional audiometrywas performed and 52.17% subjectshad bilateral hearing loss, taking into account the frequenciesof 3, 4, and 6 kHz.23

Dentists had lower mean hearing thresholds than thecontrol group, and in the right ear, this difference wassignificant at frequencies of 500, 1,000, 6,000, and 8,000 Hz(►Table 1). A study with dentists in Bauru, Sao Paulo, alsofound the right ear to haveworse hearing thresholds than theleft ear.21 Other studies identified dentists presenting worse

thresholds in their left ears.25,26 In relation to the laterality ofthe hearing thresholds, an unexpected finding was that thedentists’ group had worse hearing thresholds in the right ear,because both ears are exposed to the noise simultaneously.Future research needs to address this issue.

Even considering that hearing thresholds at high fre-quencies were higher than those at conventional frequen-cies, there was no difference between the groups. Studiesanalyzing high-frequency audiometry in dentists observeda predisposition to hearing impairment; however, in thesestudies control groups were not used as a comparison todentists.9,21

After 10 years of working as a dentist, there werereduced hearing thresholds in conventional audiometryfor the right ear at frequencies of 2,000, 4,000, and6,000 Hz, configured as an acoustic notch characteristicof hearing loss due to exposure to high sound pressurelevels. In dentists with over 10 years’ experience, all highfrequencies showed significant worsening. These findingscorroborate research that found that older workers with alonger time of service are most vulnerable to hearingimpairment.9,27 It is noteworthy that the age factor mayalso affect the results, especially in high-frequency hearingthresholds, because those with longer service as a dentistare also older.23,28

Conclusion

This study showed that dentists are at risk for the develop-ment of sensorineural hearing loss, especially after 10 years ofservice. However, there were significant differences betweenthe group of dentists and the control group (which was notexposed to occupational noise) only at frequencies of 500 Hzin both ears and 1,000, 6,000 and 8,000 Hz in the right ear.Even with the average hearing thresholds at high frequenciesbeing worse than in conventional frequencies, there were nosignificant differences between groups.

We suggest further studies to compare groups exposed toand not exposed to occupational noise to better understandthe pattern of hearing damage.

Table 4 Thresholds at high frequencies among dentists with time in service of up to 10 years (n ¼ 14) and over 10 years (n ¼ 26)


0–10 y Over 10 y p 0–10 y Over 10 y p


9,000 10.00 5.19 25.58 18.62 0.0041a 12.50 7.78 23.27 15.74 0.0216a

10,000 11.43 6.02 27.69 20.26 0.0059a 12.14 7.52 27.50 20.46 0.0103a

11,200 13.57 7.19 32.12 19.91 0.0018a 12.14 5.08 33.08 21.45 0.0010a

12,500 9.64 9.50 32.12 23.63 0.0016a 8.21 7.23 36.73 24.33 0.0001a

14,000 8.21 9.12 35.77 20.03 <0.0001a 9.29 12.22 39.62 20.49 <0.0001a

16,000 16.07 16.66 44.62 17.20 <0.0001a 17.86 16.26 44.04 17.32 <0.0001a



Hearing Thresholds in Dentists Gonçalves et al. 159

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Auditory Brainstem Response in Term andPreterm Infants with Neonatal Complications:The Importance of the Sequential EvaluationDaniela da Silva1 Priscila Lopez1 Jair Cortez Mantovani2

1Ophthalmology/ENT and Head and Neck Surgery Department,Faculdade de Medicina de Botucatu, Botucatu, São Paulo, Brazil

2 Ophthalmology/ENT and Head and Neck Surgery Department,Faculdade de Medicina de Botucatu, UNESP, Botucatu, Brazil


Address for correspondence Daniela da Silva, MSc, Departamento deOftalmologia/Otorrinolaringologia e CCP, Faculdade de Medicina deBotucatu, Distrito de Rubião Júnior s/n, Botucatu 18618970, Brazil(e-mail: [email protected]).

Introduction

Neonatal hearing screening in newborns with complicationsshould be performed using the evoked otoacoustic emissions(OAE) and auditory brainstem response (ABR) because theyare complementary tools and together provide a completeevaluation of the auditory system.1,2 The ABR test is anoninvasive neurophysiologic assessment of brainstem mat-uration in babies and could be a useful electrophysiologic testto verify neuronal myelination in preterm infants.3

Furthermore, the ABR has shown fewer false-positives andlower referral rates compared with OAE, is less sensitive to

noise and middle ear disorders even in very prematureinfants, and is an essential tool in the diagnosis of hearingloss in the pediatric population.4–7

TheABR in neonatesmayhave a highvariabilityof response.The frequentlyobservedprolonged latencies canbe considerednormal due to the characteristics of the maturation process ofthe auditory system, explained by the hypothesis that thestructures that generate the ABR core components take moretime to completely mature.8–10 Therefore, the literature re-mains unclear whether these changes in the ABR responseswould be permanent, thus indicating hearing loss, or whetherthey could be normalized with increasing age.11–14

Keywords

► neonatology► evoked potentials► auditory brainstem

Abstract Introduction Literature data are not conclusive as to the influence of neonatalcomplications in the maturational process of the auditory system observed by auditorybrainstem response (ABR) in infants at term and preterm.Objectives Check the real influence of the neonatal complications in infants by thesequential auditory evaluation.Methods Historical cohort study in a tertiary referral center. A total of 114 neonatesmet inclusion criteria: treatment at the Universal Neonatal Hearing Screening Programof the local hospital; at least one risk indicator for hearing loss; presence in bothevaluations (the first one after hospital discharge from the neonatal unit and the secondone at 6 months old); all latencies in ABR and transient otoacoustic emissions present inboth ears.Results The complications that most influenced the ABR findings were Apgar scoresless than 6 at 5 minutes, gestational age, intensive care unit stay, peri-intraventricularhemorrhage, and mechanical ventilation.Conclusion Sequential auditory evaluation is necessary in premature and term new-borns with risk indicators for hearing loss to correctly identify injuries in the auditorypathway.

receivedFebruary 25, 2014acceptedApril 26, 2014published onlineJune 13, 2014


Copyright © 2015 by Thieme PublicaçõesLTDA, Rio de Janeiro, Brazil

THIEME


According to Kohelet et al,15 the ABR response in full-termand preterm newborns of the same chronological age wassimilar, regardless of gestational age. Nevertheless, Sleifer etal observed that absolute and interpeak latencies werestatistically different between 4-month-olds and 12-month-olds.16 At 20 months, only wave I was similar. Theauthors concluded that the maturation of the auditorypathway occurs in a different way between full-term andpreterm newborns, and that gestational age must be consid-ered in the ABR analysis, mainly in infants younger than20 months. In a recent study, Roopakala et al compared theABR results in 25 preterm and 25 full-term infants andobserved a significant increased latency of ABRwaveform V in preterm infants.3

However, the ABR response in full-term and pretermnewborns may be impacted by neonatal complications thatare considered of risk for hearing loss and, consequently, maydelay the maturation process.17 Most of the studies show theinfluence of neonatal complications that are considered ofrisk for hearing loss in the ABR results. Some authors reportedthat neonates with transient low Apgar scores had a signifi-cant increase in I to V interval at very high click rates in thefirst 3 days of life.11 In another study, very low-birth-weightinfants had prolonged interpeak III to V.12 The ABR inextremely preterm infantswith bronchopulmonary dysplasiashowed a significant increase in wave V and interpeakintervals I to V and particularly III to V.13 Extremely low birthweight and mechanical ventilation may, as well, cause abnor-mal ABR.14

The question is whether these ABR findings will be recov-eredwith increasing age or neurodevelopment. The objectiveof this study was to understand the real influence of neonatalcomplications considered of risk for hearing loss, in infants,using the sequential ABR evaluation.

Methods

Study PopulationThis historical cohort study was conducted in a tertiaryreferral center from October 2008 to October 2010. Theinclusion criteria were: treatment at the Universal NeonatalHearing Screening Program of the local hospital; at least onerisk indicator for hearing loss according to the Joint Commit-tee on Infant Hearing18; presence in both evaluations (thefirst uponneonatal unit discharge, and the second at 6monthsof age); all the ABR latencies; transient otoacoustic emissionsconfirmed in both ears; consent form signed by the new-born’s parents or guardians.

For comparison, gestational age was divided into threecategories: term � 37 weeks, premature between 31 and36 weeks, and extremely premature < 31 weeks.

Audiological AssessmentFor the ABR analysis, the rarefaction click stimulus waspresented by the 3Ω insertion phone, with intensity of80-dB nHL (normal Hearing Level) and a presentation rateof 20.1 c/s (clicks/seconds) with a bandpass filter of 100 and3,000 Hz and average of 1,024 stimuli on Interacoustics EP15

Eclipse (Interacoustics A/S, Assens, Denmark). Duplicaterecordings weremade in response to each stimulus conditionto examine reproducibility. The ABR was captured throughelectrocardiogram disposable electrodes (Neuroline, AmbuA/S, Baltorpbakken, Denmark), after cleaning the skinwith electrocardiogram/electroencephalograph abrasive gel(NUPREP,Weaver and Company, Aurora, USA). The impedancelevel was kept between 1 and 3 kΩ for the electrodes; theactive electrode was positioned in Fz, the reference electrodein M1 and M2, and the ground electrode in Fpz. No sedativeswere used.

Variable and Statistical AnalysesPredictor variables included gender, gestational age, birthweight < 1,500 g, low Apgar score, infection, intensive careunit (ICU) stay, hyperbilirubinemia, peri-intraventricularhemorrhage, use of mechanical ventilation and ototoxicmedication. Outcome variables included absolute latencies(I, III, and V) and interpeak latencies (I to III, III to V, and I to V)of the ABR in both ears.

ABR change (in percentage) at 6months of age (moment 2)was calculated in relation to the ABR performed upon hospitaldischarge (moment 1) using the following expression: ABR at6 months of age � ABR postdischarge / ABR postdischarge0.100%.

The relation between risk indicator for hearing loss andvariation of ABR were analyzed using Mann-Whitney andKruskal-Wallis test, followed by Dunn for multiple compar-isons of the gestational age. Statistical analysiswas performedwith SPSS v15 and Graph Pad Prism v5 software (SPSS Inc.,Chicago, IL, USA). A p value of < 0.05 was accepted as statisti-cally significant.

EthicsThis study was approved by the Research Ethics Committee,process no. 402/08.

Results

A total of 114 newborns, 51 (45%) girls and 63 (55) boys, metthe inclusion criteria. Other sample data are shownin ►Table 1.

►Table 2 shows that Apgar score lower than 6 in the firstminute, gestational age, and ICU stay best correlated with thevariation of the absolute ABR latencies (p < 0.05).

►Table 3 shows that ICU stay, peri-intraventricular hem-orrhage, mechanical ventilation, and gestational age showedthe best relationship to the interpeak variation (p < 0.05).

Discussion

The objective of our cohort study was to verify the realinfluence of the neonatal complications considered of riskfor hearing loss, by analyzing the variation of the ABRparameters between two moments: at hospital dischargeand at 6 months of age. This second evaluation momentwas chosen based on the need of an early diagnosis of hearingloss and intervention by 6 months of age. Obviously, children


ABR in Term and Preterm Infants with Complications da Silva et al.162

with a history of hearing risk indicators are more likely tohave hearing impairment and then require periodic evalua-tion even if they had an initial normal auditoryassessment.18,19

Newborns with risk indicators for hearing loss have somecoexistent neonatal complications. However, such complica-tions are not considered by most of the studies, whichcompare only one risk indicator in healthy babies in onlyone moment or in the first days of life.

This study intended to show the real influence of riskindicators coexisting with prematurity or not and yet clarifywhether the findings in the first evaluation remain abnormalin the second one even with increasing age, considering thepresent risks. Therefore, in this study, an ABR follow-up wasperformed in twomoments, thefirst at hospital discharge andthe second at 6 months of age, to assess whether thesepossible alterations are transient or permanent.

Results show that ABR latencies and interpeaks werehigher in extremely preterm infants than in full-term new-borns, confirming other published results.3,16,17 Apparently,this difference decreases with increasing age, possibly due tothe maturational process.16 Sleifer et al reported differencesbetween these children, with very similar results at24 months of age.16 Furthermore, Turchetta et al observedan improvement over time of the estimated hearing thresh-old in ABR follow-up in preterm and full-term newborns.20

In the second assessment, when we analyzed the percent-age of changes between these two moments, extremelypremature infants showed greater reduction in the absolutelatency of wave I in both ears. Despite the statistical signifi-cance, this finding has no clinical relevance, because thevalues are similar to those in older children and healthyadults.10,17 This result shows that the peripheral transmis-sion matures faster as compared with subsequent waveformsand the same reasoning cannot be applied to reductions ofinterpeaks I to III, III to V, and I to V between the twoassessments due to improper myelination of auditory path-way and improper efficacy of higher-order neurons in infantpopulation.21

Table 1 Description of the 114 newborn according to gestationalage and risk indicators for hearing loss

Variables n (%)

Gestational age < 31 wk 29 (25%)

Gestational age 32–36 wk 49 (43%)

Gestational age �37 wk 36 (32%)

Birth weight <1,500 g: no 79 (69%)

Birth weight <1,500 g: yes 35 (31%)

Apgar < 4 (1 min): no 59 (52%)

Apgar < 4 (1 min): yes 55 (48%)

Apgar < 6 (5 min): no 94 (82%)

Apgar < 6 (5 min): yes 20 (18%)

Infection: no 71 (62%)

Infection: yes 43 (38%)

Intensive care units: no 38 (33%)

Intensive care units: yes 76 (67%)

Hyperbilirubinemia: no 109 (96%)

Hyperbilirubinemia: yes 05 (4%)

Peri-intraventricular hemorrhage: no 101 (89%)

Peri-intraventricular hemorrhage: yes 13 (11%)

Mechanical ventilation: no 53 (46%)

Mechanical ventilation: yes 61 (54%)

Ototoxic drugs: no 74 (65%)

Ototoxic drugs: yes 40 (35%)

Table 2 The p values of the relation between the variation of ABR absolute latencies and gestational age as well as risk indicators forhearing loss

Characteristics Right ear Left ear

I III V I III V

Apgar < 6 (5 min): no (n ¼ 94) � yes (n ¼ 20) 0.433 0.016 0.164 0.032 0.006 0.026

GA: <31 wk (n ¼ 29) � 32–36 wk (n ¼ 49) � � 37 wk (n ¼ 36) 0.018a 0.683 0.217 0.043 0.278 0.258

ICU: no (n ¼ 38) � yes (n ¼ 76) 0.320 0.025 0.815 0.134 0.157 0.907

BW < 1,500 g: no (n ¼ 79) � yes (n ¼ 35) 0.110 0.636 0.606 0.054 0.654 0.958


Infection: no (n ¼ 71) � yes (n ¼ 43) 0.255 0.338 0.309 0.684 0.312 0.205

Hyperbilirubinemia: no (n ¼ 109) � yes (n ¼ 5) 0.841 0.668 0.447 0.140 0.175 0.533

PIVH: no (n ¼ 101) � yes (n ¼ 13) 0.107 0.206 0.380 0.083 0.820 0.199

MV: no (n ¼ 53) � yes (n ¼ 61) 0.084 0.154 0.360 0.201 0.666 0.189

Ototoxic drugs: no (n ¼ 74) � yes (n ¼ 40) 0.764 0.960 0.204 0.667 0.838 0.268

Sex: F (n ¼ 51) � M (n ¼ 63) 0.468 0.346 0.174 0.934 0.401 0.130

Abbreviations: ABR, auditory brainstem response; BW, birth weight; GA, gestational age; ICU, intensive care unit; MV, mechanical ventilation; PIVH,peri-intraventricular hemorrhage.a < 31 6¼ � 37 (Dunn test for multiple comparisons).


ABR in Term and Preterm Infants with Complications da Silva et al. 163

The percentage of change in interpeak latencies I to III andIII to Vwas higher in preterm infants than in full-term infants,giving the impression that auditory maturation occurs morerapidly in preterm infants. Both showed change in the ABRvalues between the assessments, characterized by reducedlatencies due to the maturation process. But in the secondassessment, preterm infants had higher values, showing thatthe improvement might be related to the maturation of theauditory pathways that was not complete at birth andchanged according to gestational age.20,22

On the other hand, not only gestational agemay cause ABRalterations, but other neonatal complications may as wellcontribute to auditory maturation impairment. Despite beingfrequently studied, both aspects are not consideredconcurrently.

These studies evaluated healthy babies, without risk in-dicators for hearing loss, although it is very common thatpreterm neonates have many associated risk factors, hencethe need for a detailed analysis of the factors to whichpreterm infants may have been exposed and that may havecontributed to these remaining differences after the end ofmaturation.

In this study, newborns whose Apgar score was less than 4at 1 minute after birth and normal after 5 minutes showed nosignificant increase in the ABR latencies, but those infantswho had a low Apgar score at 5 minutes after birth hadsignificant differences in the absolute latency of wave III inboth ears.

According to these findings, the low Apgar score at 5minutes caused worse damages in the auditory pathway,despite increasing age. Jiang et al observed an increase ininterpeak I to V only within the first 3 days of life, as well asnormal results within 1 month of life for the newborns who

had a low Apgar score but had good recovery after 5minutes.14 The authors highlighted that the ABR changesobservedwithin the first 3 days of life are of little significancebecause they normalized with the development of the matu-ration process. Therefore, a low Apgar score at 1minute poseslow risk for auditory changes, and a persistently low scoremay affect the auditory pathways.

An interesting finding in this study that has not beenreported in the literature is that the infants who had a longerICU stay, without considering the primary status of suchadmission, presented clear alterations between the firstand second ABR assessments. Reductions in the absolutelatency of wave III in the right ear, interpeak I to III in bothears, and in interpeak III to V in the left ear were statisticallysignificant between the first and second assessments.

Other conditions contribute to the ABR differences, such asperi-intraventricular hemorrhage, maternal infection, cho-rioamnionitis, and neonatal infection or sepsis. These con-ditions have been associated with the development ofneonatal brain damage and adverse neurodevelopmentaloutcomes.23–26 In this study, alterations in the ABR compo-nents were observed in these infants, showing that theoccurrence of this injury deserves special attention as itsassociation with hearing loss is rarely reported in literature.

Likewise, the use of mechanical ventilation, regardless ofthe primary disease, changes the ABR recordings and is,therefore, a confounding factor along with other hearingrisk indicators.14,27 This was observed in this study whennewborns using mechanical ventilation showed reducedinterpeak I to III for the right ear and reduced interpeaklatency I to V for the left ear.

Nevertheless, other conditions frequently associatedwith hearing loss, such as low birth weight, infections,

Table 3 The p values of the relation between the variation of ABR interpeaks and gestational age as well as risk indicators for hearingloss

Characteristics Right ear Left ear

I–III III–V I–V I–III III–V I–V

ICU: no (n ¼ 38) � yes (n ¼ 76) 0.003 0.004 0.907 0.022 0.117 0.514

PIVH: no (n ¼ 101) � yes (n ¼ 13) 0.459 0.077 0.070 0.297 0.028 0.026

MV: no (n ¼ 53) � yes (n ¼ 61) 0.006 0.558 0.079 0.162 0.282 0.041

GA: <31 (n ¼ 29) � 32–36 (n ¼ 49) � � 37 (n ¼ 36) 0.066 0.006a 0.984 0.434 0.307 0.767

BW < 1,500 g: no (n ¼ 79) � yes (n ¼ 35) 0.252 0.324 0.842 0.468 0.580 0.144



Infection: no (n ¼ 71) � yes (n ¼ 43) 0.944 0.309 0.781 0.617 0.420 0.276

Hyperbilirubinemia: no (n ¼ 109) � yes (n ¼ 5) 0.410 0.609 0.171 0.934 0.678 0.552

Ototoxic drugs: no (n ¼ 74) � Yes (n ¼ 40) 0.891 0.059 0.311 0.854 0.152 0.381

Sex: F (n ¼ 51) � M (n ¼ 63) 0.180 0.514 0.067 0.268 0.227 0.065

Abbreviations: ABR, auditory brainstem response; BW, birth weight; GA, gestational age; ICU, intensive care unit; MV, mechanical ventilation; PIVH,peri-intraventricular hemorrhage.a < 31 6¼ 32–36; <31 6¼ � 37 (Dunn test for multiple comparisons).


ABR in Term and Preterm Infants with Complications da Silva et al.164

hyperbilirubinemia with serum level requiring transfusion,and use of ototoxic drugs, showed no significance in the ABRparameters with increasing age.

Results showed that the waveform I is less sensitive to theinjuries that occurred during pregnancy and/or neonatalcomplications. However, waves III and V, which characterizethe maturation of axons and synaptic mechanisms in thebrainstem level, are more likely to be affected by the riskanalyzed in this study.28 Therefore, the differences in the ABRresponses between full-term and preterm newborns andwith some risk indicators for hearing loss cannot only bedue to nervous maturation, but might as well be caused byinjuries occurred during pregnancy and/or neonatalcomplications.17

We believe that these findings bring an important contri-bution to clinical practice. Neonates with complicationsdeserve more attention paid to auditory evaluation, becauseprematurity and neonatal complications may interfere in theABR findings. The interpretation and/or standardization ofthe results may be uncertain if only one assessment isperformed.

Conclusion

The complications thatmost impacted the ABR findingswere:Apgar scores less than 6 at 5 minutes of birth, gestational age,ICU stay, peri-intraventricular hemorrhage, and mechanicalventilation.

The sequential auditory evaluation is necessary in pretermand full-term newborns with risk indicators for hearing lossto correctly identify injuries in the auditory pathway.

References1 Joint Committee on Infant Hearing. Year 2000 position statement:

principles and guidelines for early hearing detection and inter-vention programs. Am J Audiol 2000;9(1):9–29

2 Stephens D, Hétu R. Impairment, disability and handicap inaudiology: towards a consensus. Audiology 1991;30(4):185–200

3 Roopakala MS, Dayananda G, Manjula P, et al. A comparative studyof brainstem auditory evoked potentials in preterm and full-terminfants. Indian J Physiol Pharmacol 2011;55(1):44–52

4 Lin HC, Shu MT, Lee KS, Lin HY, Lin G. reducing false positives innewborn hearing screening program: how andwhy. Otol Neurotol2007;28(6):788–792

5 Benito-Orejas JI, Ramírez B, Morais D, Almaraz A, Fernández-CalvoJL. Comparison of two-step transient evoked otoacoustic emis-sions (TEOAE) and automated auditory brainstem response (AABR)for universal newborn hearing screening programs. Int J PediatrOtorhinolaryngol 2008;72(8):1193–1201

6 Granell J, Gavilanes J, Herrero J, Sánchez-Jara JL, VelascoMJ, MartínG. [Is universal newborn hearing screening more efficient withauditory evoked potentials compared to otoacoustic emissions?]Acta Otorrinolaringol Esp 2008;59(4):170–175

7 Pedersen L, Møller TR, Wetke R, Ovesen T. [Neonatal hearingscreening. A comparison of automatic auditory brainstem audi-ometry and otoacoustic emissions]. Ugeskr Laeger 2008;170(8):642–646

8 Morgan DE, Zimmerman MC, Dubno JR. Auditory brain stemevoked response characteristics in the full-term newborn infant.Ann Otol Rhinol Laryngol 1987;96(2 Pt 1):142–151

9 Zimmerman MC, Morgan DE, Dubno JR. Auditory brain stemevoked response characteristics in developing infants. Ann OtolRhinol Laryngol 1987;96(3 Pt 1):291–299

10 Ponton CW,Moore JK, Eggermont JJ. Auditory brain stem responsegeneration by parallel pathways: differential maturation of axonalconduction time and synaptic transmission. Ear Hear 1996;17(5):402–410

11 Jiang ZD, Xiu X, Brosi DM, Shao XM, Wilkinson AR. Sub-optimalfunction of the auditory brainstem in term infants with transientlow Apgar scores. Clin Neurophysiol 2007;118(5):1088–1096

12 Jiang ZD, Wilkinson AR. Brainstem auditory evoked response atterm in preterm infants after perinatal hypoxia-ischaemia. ActaPaediatr 2006;95(11):1400–1404

13 Wilkinson AR, Brosi DM, Jiang ZD. Functional impairment of thebrainstem in infants with bronchopulmonary dysplasia. Pediatrics2007;120(2):362–371

14 Jiang ZD, Yin R, Wilkinson AR. Brainstem auditory evoked re-sponses in very low birthweight infants with chronic lung disease.Eur J Paediatr Neurol 2007;11(3):153–159

15 Kohelet D, Arbel E, Goldberg M, Arlazoroff A. Brainstem auditoryevoked response in newborns and infants. J Child Neurol 2000;15(1):33–35

16 Sleifer P, da Costa SS, Cóser PL, Goldani MZ, Dornelles C, Weiss K.Auditory brainstem response in premature and full-term children.Int J Pediatr Otorhinolaryngol 2007;71(9):1449–1456

17 Jiang ZD, Brosi DM, Li ZH, Chen C, Wilkinson AR. Brainstemauditory function at term in preterm babies with and withoutperinatal complications. Pediatr Res 2005;58(6):1164–1169

18 American Academy of Pediatrics, Joint Committee on Infant Hear-ing. Year 2007 position statement: principles and guidelines forearly hearing detection and intervention programs. Pediatrics2007;120(4):898–921

19 Lewis DR, Marone SAM, Mendes BCA, Cruz OLM, de Nóbrega M.Comitê Multiprofissional em Saúde Auditiva COMUSA. Braz JOtorhinolaryngol 2010;76:121–128

20 Turchetta R, Orlando MP, Cammeresi MG, et al. Modifications ofauditory brainstem responses (ABR): observations in full-term andpre-term newborns. J Matern Fetal Neonatal Med 2012;25(8):1342–1347

21 Chalak S, Kale A, Deshpande VK, Biswas DA. Establishment ofnormative data for monoaural recordings of auditory brainstemresponse and its application in screening patients with hearingloss: a cohort study. J Clin Diagn Res 2013;7(12):2677–2679

22 Casali RL, Santos MFC. Auditory brainstem evoked response:response patterns of full-term and premature infants. Braz JOtorhinolaryngol 2010;76(6):729–738

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ABR in Term and Preterm Infants with Complications da Silva et al. 165

Health Promotion in Obstructive Sleep ApneaSyndromeCamila de Castro Corrêa1 Wanderléia Quinhoneiro Blasca1 Giédre Berretin-Felix1

1Department of Speech-Language Pathology and Audiology,Bauru School of Dentistry, Universidade de São Paulo, Bauru,São Paulo, Brazil


Address for correspondence Camila de Castro Corrêa, BS, MS,Department of Speech-Language Pathology and Audiology, BauruSchool of Dentistry, University of São Paulo, Octávio Pinheiro BrisolaStreet, Bauru, Sao Paulo 17012-901, Brazil(e-mail: [email protected]; [email protected]).

Introduction

Sleep disorders are characterized by alterations in sleep thataffect the quality of life and the execution of the individual’sdaily activities.1 Such disorders include sleeplessness, noc-turnal bruxism, narcolepsy, somniloquy, nocturnal enuresis,parasomnias, night terror, restless legs syndrome, sleepwalk-ing, and obstructive sleep apnea syndrome (OSAS).2,3

OSAS has demonstrated increased occurrence in theworldwide population. Studies on the prevalence of OSAShave been performed in several places of the world, consid-ering child and adult populations, although without stan-dardization of methods or procedures. In children, OSASprevalence was found to be between 0.8 and 2.8%4; however,the prevalence was higher in oral breathing and obesechildren.5–7 In the adult population, a prevalence of 2 to26% was observed,8–13 increasing from 39 to 71% in obeseadults.14

In Brazil, only one study on the prevalence of OSAS usedpolysomnography, considered the highest standard for thediagnosis of such condition. The sample of 1,042 volunteers,aged 20 to 80 years, resulted in 32.8% of the participantsdiagnosed with OSAS. This study presents findings of higherOSAS prevalence as compared with the above-mentionedstudies.15

Besides the OSAS effect during sleep (i.e., intermittenthypoxemia, transient hypercapnia, and frequent waking),16

other implications such as attention deficit in the learningprocess and in auditorymemory and alteration of the tonicityof orofacialmyofunctionalmusclesmay be observed.17–20 It isworth mentioning the consequences affecting the popula-tion, such as decreased job productivity and increased workand traffic accidents.21–23

Studies demonstrated low rates of OSAS diagnosis,24 not-ing that 95% of the individuals were not identified due to thelack of general knowledge and specific knowledge on sleep

Keywords

► health education► sleep apnea► obstructive► sleep disorders► speech► language and hearing

sciences

Abstract Introduction Obstructive sleep apnea syndrome (OSAS), which is commonly under-diagnosed, has a high occurrence in the world population. Health education concerningsleep disorders and OSAS should be implemented.Objectives The objective was to identify studies related to preventive actions on sleepdisorders, with emphasis on OSAS.Data Synthesis A literature review was conducted using Lilacs, Medline, PubMed, andScopus by combining the following keywords: “Health Promotion,” “Sleep Disorders,”“Primary Prevention,” “Health Education,” and “Obstructive Sleep Apnea Syndromes.”Initially, 1,055 papers, from 1968 to 2013, were located, with the majority from theScopus database. The inclusion criteria were applied, and four articles publishedbetween 2006 and 2012 were included in the present study.Conclusions The studies on preventive actions in sleep disorders, with emphasis onOSAS, involved the general population and professionals and students in the health fieldand led to increased knowledge on sleep disorders and more appropriate practices.

receivedMay 9, 2014accepted after revisionAugust 25, 2014published onlineJanuary 26, 2015



Systematic ReviewTHIEME

166

disorders by health care professionals.25–31 Thus, healtheducation is needed for the identification of OSAS symptomsdirected to the general population, as well as to health careprofessionals, aiming at better diagnosis and earlier detectionthrough specific protocols.

Therefore, the purpose of the present work was to identifystudies involving preventive health care to sleep disorders,with emphasis on OSAS.

Review of Literature

The literature review was done by searching the databasesLilacs, Medline, PubMed, and Scopus, using the followingDeCS/MeSH keywords: (1) “Sleep Disorders”; (2) “SleepApnea Syndromes”; (3) “Health Promotion”; (4) “HealthEducation”; (5) “Primary Prevention.” Their respective termsin Portuguese were also used for the database Lilacs.

Six matches of two keywords were used each time in all thedatabases selected. The keywords were separated by the word“AND” in blank search. The combinations were: 1, 3 (combina-tion 1); 1, 4 (combination 2); 1, 5 (combination 3); 2, 3(combination 4); 2, 4 (combination 5); and 2, 5 (combination 6).

The following inclusion criteria were elected: specific stud-ies concerning preventive actions for health promotion onsleep disorders; emphasis on OSAS; maximum amount ofinformation to people about such clinical condition. Studiesabout health promotion on sleep disorders were includedbecause general information implies favorable attitudes re-garding the quality of sleep, such as sleep hygiene, which is away of preventing OSAS. The exclusion criteria involved litera-ture review and the approach of only the diagnostic evalua-tion/treatment of subjects with OSAS or other sleep disorders.

For this analysis, titles and abstracts of the works foundwere read, and, when they met the inclusion criteria, theworks were read in full and analyzed as to their objectives,methods, results, and conclusion. The search was carried outwithout time limitation.

Results

In all, 1,055 articles from 1968 to 2013 were found, of which159 articles were located in Medline, 7 in Lilacs, 173 inPubMed, and 716 in the Scopus databases. ►Fig. 1 showsthe percentages related to the results found in these databases.

The results of the articles located through the combinationof the keywords DeCS/MeSH, considering all the databases,are shown in ►Table 1.

Thus, the titles of the 1,055 works found were read, and991 were discarded through the exclusion criteria. Of the 64abstracts read, 47 were not considered; 17 seemed to meetthe inclusion criteria were read in full, resulting in theexclusion of another 13 additional articles. This way, 4 articleswere effectively included in the present study, and the resultsobtained through the analysis of the selected articles, con-cerning the purpose, methods, results, and conclusion, areshown in ►Table 2. One article was located in Lilacs, one inPubMed, and two in Scorpus; two were published in 2006,one in 2009, and one in 2012.

Discussion

Considering the high prevalence of OSAS and the low rate ofdiagnosis, it is important to highlight the value of the studiesdirected to health education intended for the population ingeneral and for health care professionals, enlarging the accessto reliable and current information, focusing on assertiveevaluation, and, mainly, guiding measures of prevention tothis condition.

Based on the articles found in the search for this study,more results were observed in the Scopus database and fewerin the Lilacs database. Most publications were found in theSciVerse Scopus, as its scope is international, with more than18,000 titles of 5,000 international publishing companies,and it provides author and institution profiles, citation track-ing, h-index, and journal analyzer. Moreover, articles in pressof more than 3,000 periodicals are available—that is, thosearticles that were accepted but have not yet appeared in aregular edition of the journal.36 On the other hand, the Lilacsdatabase, through which fewer works were obtained, re-stricts its scope to Latin America and the Caribbean and has

Fig. 1 Percentages of the results obtained in the databases consulted.

Table 1 Articles found through the combination of keywords inthe databases searched in this study

Combination of the keywordsDeCS/MeSH

Number of articlesfound

Combination 1 363

Combination 2 354

Combination 3 114

Combination 4 71

Combination 5 65

Combination 6 88

Total 1,055


Health Promotion in Obstructive Sleep Apnea Syndrome Corrêa et al. 167

contributed for 27 years to the increase of the visibility,access, and quality of health information.37

A difference in the results obtained in relation to theamount of articles found, when using six different combina-tions, was verified. When combination 3 was used, in theEnglish and Portuguese languages, more results were veri-fied, whereas with combination 6, fewer results were found.Such findings can be justified by the fact that “Health Promo-tion” represents awider concept, including the objective of anexcellent level of life and health, and “Primary Prevention”presents the objective for illness prevention,38 according tomodern concepts of health promotion and primary care,initiated after the Ottawa Charter for Health Promotion.39

After the 1,055 articles were read, 93.9% of the articleswere excluded in this first stage, for even using keywords andspecific combinations related to health promotion, sleepdisorders, and OSAS, many of the studies were aimed at thediagnosis, assessment, treatment, and characterization ofOSAS cases and other sleep disorders. This finding suggeststhat the scientific research has prioritized investigationsdirected to diagnosis and intervention. On the other hand,in the area of health, the importance of health promotion hasbeen described as well, although unplanned actions havebeen observed or restricted to a certain number of peo-ple,40,41 thus limiting the results achieved.

Only recent articles, dating from 2006 to 2012, composedthe results of this work, thus the theme is quite current.Moreover, the actions taken were directed not only to healthprofessionals, but also to doctors,32 medical students,35 post-partum mothers,33 and adolescents.34 The value of the ac-tions directed to professionals/students in the area of health,such as pharmacists42 and pediatricians,43 is highlighted, dueto their qualification on sleep disorders, so as to improve thediagnosis of OSAS. Actions are also important for the generalpopulation, including children and adolescents, because oftheir increased risk of OSAS caused by behaviors that aredetrimental to a good quality of sleep, such as insufficientamount of sleep, television sets in the bedroom, and late andvariable bedtimes.44

Thestudiespresented severalmethods in the transmissionofknowledge, such as lessons in the face-to-face format,34,35

online modules, book discussions,33 and media and exhibitresources.32 IThe possibility of doing an online screening ofthose at risk of OSAS was noted,45 as well as guidance to assistparents in dealing with the diagnosis of insomnia, by means ofthe Mini-Kiss online.46 The positive results obtained throughlocalized actions support the need for specific methods forhealth education according to the profile of the target public.

In the four selected studies, increased sleep knowledge byadolescents and medical students after the actions was

Table 2 Analysis of the articles included, concerning the objective, methods, results, and conclusion

Author, year Purpose Methods Results Conclusion

Conway et al.32 To promote a campaignto increase the aware-ness on sleep disorders

Resources of media,exhibits, and lessonswere used to reach2,000,000 people and55,000 health profes-sionals, evaluating thereferrals made after theinformation was sup-plied in a hospital.

A slight increase in thediagnosis of obstructivesleep apnea syndromeswas observed, amongother sleep disorders,through the referralsmade.

Health professionalsseemed to have a higherunderstanding of sleepdisorders.

Stremler et al.33 To assess the viabilityand acceptability ofinformation about sleepby postpartum mothers

Behavioral-educationalintervention withfirst-timemothers in thepostpartum period wasperformed by nurses,on sleep information, bymeans of a book.

Children from the sleepintervention group hadless nocturnal wakingand more total sleeptime.

A more adequate sleepwas provided for bothmother and child.

Moseley andGradisar34

To investigate the effec-tiveness of an interven-tion in the increase ofknowledge about sleepdirected to adolescents

Adolescents receivedfour lessons on promo-tion andmaintenance ofa healthy lifestyle,reflecting on the qualityof sleep.

The program increasedthe adolescents’ knowl-edge about sleep.

The interventionsproposed for the ado-lescents, emphasizingthe reduction of practi-ces that are detrimentalto sleep, were valid.

Bandla et al.35 To compare the level ofsatisfaction andeconomic factor onsleep medicine,presented virtually andin a face-to-face format

Medical students wereinstructed on sleepmedicine in one of thetwo formats: face-to-face or onlinemodules.

The pupils who partici-pated in the face-to-faceformat were moresatisfied than those whoparticipated online. Thelearning and its costswere approximatelyequivalent for bothformats.

The results between thetwo methodologieswere similar, with theonline platform beingeconomical andeducationally feasible.


Health Promotion in Obstructive Sleep Apnea Syndrome Corrêa et al.168

verified,34,35 as well as increased rate of diagnosis of mildOSAS and other nocturnal sleep disorders,32 reduction ofwaking, and increased nocturnal sleep in babies.33 Suchfindings show that planning actions for the diagnosis ofOSAS directed to the largest number of individuals is para-mount, as in the study of Conway et al.32 Teaching aboutsuitable treatments and incentives for the appropriate pro-cedures in each case is also important,47–49 demonstratingthat an assertive diagnosis should be associated with asuccessful treatment.

Only articles in Portuguese, English, and Spanish wereselected after the headings of the articles were read. Thus,the language of the works was not a limiting factor for theresearch. Moreover, the search in the Virtual Private Networksystem, which provides a myriad of journals, enabled thereading in full of all selected articles, a contributing factor forthe development of the study.

In addition to the investigation of articles published onpreventive actions to health care, future studies be performedinvestigating Web sites related to the subject, as well asactions by official scientific organizations in the field of sleepmedicine. Hence, reliable information regarding education onhealth aimed at OSAS can be found, such as the actionspromoted in World Sleep Day in the official Web site andorganized by theWorld Association of Sleep Medicine, whichaims at emphasizing the importance of quality of sleep.50

The lack of scientific studies focusing on health promotionin OSAS demonstrates that further research using this ap-proach, directed to both professionals and the population,is necessary, aiming at minimizing the consequences thatsuch clinical condition brings to those affected.

Final Comments

A literature reviewof studies on health promotion specificallyinvolving preventive actions of health care in sleep disorders,with emphasis on OSAS, was presented. Four studies thatincreased the knowledge of the population, professionals, andhealth care students on health and sleep disorders werefound, promoting assertive referrals by health care profes-sionals and the adherence of the population tomore favorablebehaviors, aiming at suitable sleep.

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The Study of Otoacoustic Emissions and theSuppression of Otoacoustic Emissions inSubjects with Tinnitus and Normal Hearing:An Insight to Tinnitus EtiologyLucieny Serra1 Gabriela Novanta1 Andre Lopes Sampaio1 Carlos Augusto Oliveira1

Ronaldo Granjeiro2 Silvia Cristina Braga2

1Department of Otolaryngology, Universidade de Brasília, Brasília,Distrito Federal, Brazil

2 Secretaria de Saúde–Hospital de Base, Brasília, Distrito Federal, Brazil


Address for correspondence Lucieny Serra, MSc, Department ofOtolaryngology, Universidade de Brasilia, SGAS 604 AV L2 Norte Setorde Saude Auditiva Asa Norte, Brasilia, Distrito Federal 70830200, Brazil(e-mail: [email protected]).

Introduction

Tinnitus is a condition in which the individual perceives asound in the ears or in the head in the absence of outsidesounds. A multidisciplinary approach should be used for thetreatment of tinnitus as its physiopathology is not fullyunderstood and it seems to be related to disorders of theouter, middle, or inner ears; brain stem; and cerebral cortex.1

Individuals with tinnitus frequently report sleep disor-ders, concentration and attention problems, and symptoms of

anxiety and/or depression. These problems can cause diffi-culties at work and can compromise socialization with familyand leisure activities, interfering with social habits and dailydynamics. Each patient is affected differently, irrespective ofpresence of hearing impairment, and the condition has majornegative consequences on quality of life and psychologicalstate.2

Tinnitus may be associatedwith abnormalities in any levelof the auditory pathways. However, it starts very often in the

Keywords

► tinnitus► organ of Corti/

physiology► otoacoustic emissions

Abstract Introduction Analysis of the suppression effect is a simple method to evaluatecochlear status and central auditory mechanisms and, more specifically, the medialolivocochlear system. This structure may be involved in the generation of mechanismsthat cause tinnitus and in the pathophysiology of tinnitus in patients with tinnitus andnormal hearing.Objective To review the literature of the etiology of tinnitus on the lights ofotoacoustic emissions in patients with normal hearing.Data Synthesis Individuals with tinnitus and normal hearing have a higher prevalenceof alterations in transient-evoked otoacoustic emissions and distortion-product otoa-coustic emissions than normal subjects. This fact suggests that dysfunctions of theouter hair cells (OHCs) might be important in the generation of the tinnitus; however,this feature is not always present in those who have the symptoms of tinnitus.Final Comments These findings suggest that OHC dysfunction is not necessary fortinnitus development—that is, there might be mechanisms other than OHC damage inthe tinnitus development. On the other hand, OHC dysfunction alone is not sufficient tocause the symptom, because a great many individuals with OHC dysfunction did notcomplain about tinnitus.

receivedDecember 12, 2013acceptedMarch 6, 2014published onlineJune 18, 2014



THIEME

Systematic Review 171

cochlea. Jastreboff considers that tinnitus usually starts in thecochlea and later generates abnormal activity in the centralpathway, which perpetuates the symptom.3

Several studies have investigated the relationship betweentinnitus and dysfunction of the efferent auditory system bymeasuring the suppression of otoacoustic emissions (OAEs).Evidence indicates that subtle changes in cochlear functioncan be detected by OAE testing even before the occurrence ofsignificant changes in the patient’s audiogram.4 Therefore,the study of OAEs and its suppression are extremely impor-tant and have become increasingly necessary for the differ-ential diagnosis of diseases that might cause tinnitus.

This article reviews the literature on OAE testing andsuppression of OAEs in patients with normal hearing andtinnitus.

Review of the Literature

Otoacoustic EmissionsAt the end of the 1970s, Kemp defined OAEs as the release ofsound energy produced in the cochlea and its propagation tothe middle ear and external acoustic meatus.5 The authorsuggested that these emissions are produced by the outer haircells (OHCs) of the cochlea as a result of active and nonlinearmechanical feedback processes, which can be spontaneous orevoked by sounds of low and medium intensity.5 OAEs areclassified as spontaneous or evoked. The latter are dividedinto transient, distortion-product, or stimulus-frequencyOAEs.

Spontaneous otoacoustic emissions (SOAE) are narrow-band signals recorded in the ear canal in the absence of anacoustic stimulus, which can be detected in �50% of individ-uals with normal hearing. SOAEs disappear in the frequencyrange associated with hearing losses that exceed 30-dB HL(hearing level) and are present at normal auditory thresh-olds.6,7 The clinical applicability of SOAEs is restricted be-cause this screening test is of low specificity. In addition,these OAEs may be absent even in individuals with normalmiddle and inner ear function.8

Transient-evoked otoacoustic emissions (TEOAEs) are low-intensity responses produced by the cochlea. These emissionsare sound waves emitted after a short acoustic stimulus(clicks or tone bursts) that occur over a wide range offrequencies, thus permitting broad stimulation of the co-chlea.7,8 TEOAEs can be detected in individuals with normalOHC function at the frequency analyzed, or in individualswith auditory thresholds below 30-dB HL. TEOAE screening isable to detect impaired hearing but is unable to identify thetype and degree of impairment.9,10

Like TEOAE, distortion-product otoacoustic emissions(DPOAEs) are sounds generated by OHCs in response toconcomitant stimulation with two pure tones (f1 and f2)with closely similar frequencies (f2/f1 ¼ 1.22). By convention,the lower-frequency pure tone is referred to as primary f1 andits intensity level is L1. The higher-frequency tone is called f2and its intensity level is L2. The parameters analyzed inDPOAE testing are amplitude of the signal and signal-to-noiseratio.5,11,12 Studies using DPOAEs showed that this type of

emission is present in individualswithmild hearing loss of upto 50 dB, depending on the intensity levels used.11,13 Theadvantage of this type of OAE is the greater specificity of thefrequency, which permits evaluation of cochlear functionfrom the basal (high frequencies) to the apical (low frequen-cies) turns by varying the primary frequencies of the evokingstimuli.4,14

Effect of Otoacoustic Emission SuppressionAnalysis of the suppression effect is a simple method thatpermits evaluation of cochlear status and central auditorymechanisms, more specifically, the medial olivocochlear sys-tem.15 In 1946, Rasmussen16 described the anatomy of theefferent auditory pathway. In the cochlea, efferent controloccurs through the olivocochlear system that originates in thesuperior olivary complex. The system comprises two mainbundles, the lateral and medial bundle. The lateral bundleconsists of unmyelinated fibers that project ipsilaterally fromthe lateral region of the superior olivary complex to the innerhair cells. The medial bundle is composed mainly of crossed(�80%)myelinated fibers that originate in the area around themedial superior olivewhere they directly innervate the OHCs.The olivocochlear system can be activated by electrical orchemical stimulation or noise. Once activated, the systeminhibits OHC contraction, thus reducing the amplitude ofOAEs.7,15,17 As a consequence, the effect of suppression ofevoked OAEs is characterized by a reduction in the responseamplitude or by latency and phase changes when a contra-lateral noise is introduced simultaneously to the recording.18

The suppression value is calculated as the differencebetween the values obtained in the presence and absenceof the stimulus. In this respect, a suppression effect isconsidered to be present when there is a reduction of atleast 0.5-dB SPL (sound pressure level) in OAE amplitudes inthe presence of a contralateral noise. A suppression effect of0.5 to 1.0 indicates integrity of the medial olivocochlearsystem.19,20

The presence of a suppression effect in individuals withnormal hearing thresholds is a phenomenon reported in alarge number of studies. The suppressor stimulus frequentlyemployed is a contralateral broadband noise. However, ipsi-lateral or simultaneous noise stimuli have been used indifferent studies.15,21,22

Tinnitus Generation MechanismsMany theories describe the generation of tinnitus. In thisstudy, only those that possess direct relationship with theexams of EOA and suppression will be described.

One of the theories for the tinnitus is that it might occur byvirtue of the SOAEs (although nowadays this theory is notwell accepted). The incidence of tinnitus originated by theSOAEs is frequently uncommon and occurs in less than 10% ofthe population with tinnitus.23

Another hypothesis for tinnitus refers to the existingdisproportional damage between the hair cells. Traumaticsituations such as exposure to intense noise or use of ototoxicdrugs may cause cochlear damage. These injuries initiallyoccur in the basal portion of the basilar membrane (region of


Otoacoustic Emissions in Subjects with Tinnitus Serra et al.172

high frequencies) and in the outer hair cells (OHCs), followingthe inner hair cells (IHCs). In this way, it is possible to observecochlear regions with total lesion of the outer and inner haircells and other areas of the cochlea with lesion on the OHCsbut with unharmed IHCs. The alterations of the mechanicproperties of the organ of Corti with lesions on the OHCsproduce a tonic depolarization (blocking by depolarization)of the IHCs and the appearance of an irregular activity on theafferent fibers.24,25

When there is a lesion of a group of OHCs, a reduction ofentrance of the auditory information is observed, provokingdiminished efferent activity and inhibition of rapid contrac-tions of the OHCs in this region. Considering that the inner-vation of the efferent fibers is very diffuse, it is possiblethat this inhibition affects adjacent healthy regions, causingthe hyperactivity of the healthy OHCs perceived as beingtinnitus.1,26

Granjeiro at the University of Brasilia stated that probablythe less efferent inhibition in the ears with tinnitus resultsfrom functional characteristics of the efferent system withpossible origin on the cochlea ormedial olivocochlear system,on the activation of the medial olivocochlear system (MOCsystem), or with an association of many possibilities.6

The medial efferent system influenced by the autonomicnervous system also is another hypothesis, possibly causingoscillations of the symptoms of tinnitus as a consequence ofstress, suggesting the presence of parasympathetic activity inthe efferent disinhibition.27

Discussion

OAE testing has been used as part of the protocol of audiologicevaluations and is currently an important tool in studies ontinnitus. Tinnitus is a frequently reported symptom, even inpatients with normal hearing. A study conducted at theUniversity Hospital of Santa Maria analyzed 480 audiologicevaluations performed between 2005 and 2008 and foundnormal hearing in 17.08% and tinnitus in 7.5%.28 A significantdifference was observed between genders, with a higherprevalence of tinnitus in women.29

Several studies have demonstrated a relationship betweentinnitus and abnormal OAEs. In a study involving normal-hearing patients, TEOAEs and DPOAEs were abnormal in 70.2and 68.4% of the groupwith tinnitus, respectively, but only in16.10 and 50% of those without tinnitus.30 Similar resultshave been reported by our research group. We observedabnormal TEOAEs and DPOAEs in 67 and 65.2% of patientswith normal hearing and tinnitus, respectively.31

Tinnitus has been suggested to be related to a reduction inthe amplitude of OAEs. Fernandes and Santos found a loweramplitude in all frequency bands and in both ears of normal-hearing individuals with tinnitus when compared with thosewithout tinnitus.32Mor and Azevedo observed a difference inthe cases of unilateral tinnitus, with the overall amplitudebeing lower in the ear with tinnitus.16

The higher prevalence of abnormal OAEs in patients withtinnitus suggests that cochlear dysfunction is involved in the

development of this condition, especially at higher frequen-cies (6 and 8 kHz).30,31,33

Studies developed by Maurer and Mann reported anothercategory of individuals in which the otoacoustic emissionswere present with greater intensity on the patients withtinnitus and auditory thresholds in the 3,000-, 4,000-, and5,000-Hz frequencies, suggesting that the tinnitus might begenerated by the increase in the motility of the OHCs inducedby the diminished activity of the efferent system and not byfailure of the OHCs.34

According to Azevedo et al,35 the auditory perception oftinnitus might be related to dysfunction of the efferentauditory pathway, which would result in the loss of OHCmodulation, causing abnormal activity of the auditory path-ways that could be wrongly interpreted as sound.

The study of OAE suppression provides reliable informa-tion about efferent system function and the interactionbetween afferent and efferent pathways and also contributesto the differentiation between peripheral and central hearingloss. The absence or reduction in the suppression of OAEsmay occur in cases of retrocochlear diseases, auditory neu-ropathy, acoustic neuroma, impaired auditory processing,and tinnitus.15 Studies investigating suppression in individu-als with normal hearing and tinnitus have increased inrecent years. This fact demonstrates concern regarding theneed for a better understanding of the function of theauditory pathway.

Urnau and Tochetto studied normal-hearing adults withtinnitus and hyperacusis and found no association betweenthe suppression effect of TEOAEs and laterality, degree oftinnitus, or degree of hyperacusis.36 A study conductedinvolving a group of normal-hearing individuals with tinnitusshowed that the absence of OAE suppression at frequencies of1.5, 2.0, 3.0, and 4.0 Hz was strongly associated with thepresence of tinnitus. The same study demonstrated lowersuppression of DPOAEs on the side of perceived tinnituscompared with contralateral suppression.36 Another similarstudy recording suppression of OAEs in individuals withtinnitus and normal hearing concluded that the resultswere less reproducible in tinnitus patients compared withthe control group, but no significant differences in latency orsuppression effectswere observed between the two groups.33

In a recent study conducted in The Netherlands, theauthors found no significant differences in the overall analysisof OAE suppression between tinnitus patients and the controlgroup. The suppression value was lower in the right ear oftinnitus patients for the 2.0- and 2.8-Hz frequency bands butwas similar to the control group at the other frequencies.37

Azevedo et al investigated the effect of acupuncture onOAEs in tinnitus patients and found an increase in theamplitude and suppression of OAEs after acupuncture treat-ment,35 suggesting that this method exerts an effect oncochlear function. Lower suppression values were also ob-served in children with impaired central auditory processingwhen compared with the control group. This finding agreeswith the literature showing low or absent suppression ofOAEs in cases of cochlear diseases, auditory neuropathies,central auditory processing disorders, and tinnitus.38


Otoacoustic Emissions in Subjects with Tinnitus Serra et al. 173

In conclusion, the efferent auditory pathways have beenextensively investigated in normal-hearing individuals withtinnitus. However, these studies have not been conclusive anda better understanding of this topic is needed. For this reason,our group has started a new protocol to investigate the effectof OAE suppression in the genesis of tinnitus at our University.

Final Comments

Based on the previous studies, it is possible to observe thatindividuals with tinnitus and normal hearing have a higherprevalence of alterations in the exams of TEOAEs and DPOAEsthan normal subjects. This fact suggests that dysfunction ofthe OHCs might be important in the generation of thetinnitus. However, we studied a group of patients complain-ing about tinnitus and having normal OAE tests. These find-ings suggest that the OHC dysfunction is not necessary fortinnitus development—that is, it might be possible that amechanism other than OHCs is involved in tinnitus develop-ment.30,31 On the other hand, the OHC dysfunction is notsufficient in itself to cause the symptom, because mostindividuals with OHC dysfunction did not complain abouttinnitus.30,31 In this way, it is important to consider thehypothesis of the involvement of the efferent system inthe generation of tinnitus, as well as its analysis by meansof the test of suppression of the EOA.

Despite the growing number of publications in recentyears, the results of studies investigating tinnitus in nor-mal-hearing patients using OAEs are still inconclusive andfurther research in this area is needed.

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34 Gouveris H,Maurer J,MannW.DPOAE-grams inpatientswith acutetonal tinnitus. Otolaryngol Head Neck Surg 2005;132(4):550–553

35 Azevedo RF, Chiari BM, Okada DM, Onishi ET. Efeito da acumpul-tura sobre as emissões otoacústicas de pacientes com zumbido.Rev Bras Otorrinolaringol 2007;73(5):599–607

36 Urnau D, Tochetto TM. Occurrence and suppression effect ofotoacoustic emissions in normal hearing adults with tinnitusand hyperacusis. Braz J Otorhinolaryngol 2012;78(1):87–94

37 Fávero ML, Sanchez TG, Bento RF, Nascimento AF. Supressãocontralateral das emissões otoacústicas nos indivíduos com zum-bido. Rev Bras Otorrinolaringol 2006;72(2):223–226

38 Geven LI, de Kleine E, Free RH, vanDijk P. Contralateral suppressionof otoacoustic emissions in tinnitus patients. Otol Neurotol 2011;32(2):315–321

39 Ribeiro PS, Torres TL, Starling ALP, Iório MCM, Mancini PC.Crianças com fenilcetonúria: avaliação audiológica básica e su-pressão das otoemissões. Rev Soc Bras Fonoaudiol 2012;17(3):248–253


Otoacoustic Emissions in Subjects with Tinnitus Serra et al. 175

Olfaction in Neurologic and NeurodegenerativeDiseases: A Literature ReviewMaria Dantas Costa Lima Godoy1 Richard Louis Voegels1 Fábio de Rezende Pinna1 Rui Imamura1

José Marcelo Farfel2

1Department of Otorhinolaryngology, Faculdade de Medicina daUniversidade de São Paulo, São Paulo, São Paulo, Brazil

2Department of Geriatrics, Faculdade de Medicina da Universidade deSão Paulo, São Paulo, Brazil


Address for correspondence Maria Dantas Costa Lima Godoy, MD,Department of Otorrinolaringologia, Faculdade de Medicina daUniversidade de São Paulo, Rua Dr. Eneas de Carvalho Aguiar, 255, 6°andar, 6167 Cerqueira Cesar, São Paulo 05403000, Brazil(e-mail: [email protected]).

Introduction

Olfaction is usually neglected by physicians at large, eventhough it monitors the intake of airborne agents into thehuman respiratory system, including dangerous substances,and warns of spoiled foods. Furthermore, this primary sen-sory system enhances quality of life by adding to flavor andpalatability of foods and beverages.1 Most complaints ofdecreased “taste” function actually reflect decreased smellfunction.1,2 In addiction, loss of smell is involved in variousneurologic and neurodegenerative diseases, such as Parkin-son disease (PD), Alzheimer disease (AD), multiple sclerosis,and Huntington disease.3

The aim of this study was to review the current literatureabout the relationship between olfactory dysfunction andneurologic and neurodegenerative diseases.

Review of the Olfactory System

Olfactory Mucosa Cellular Composition andNeurogenesisHuman olfactory mucosa is a pseudostratified columnarepithelium resting on a highly cellular lamina propria. Fourtypes of cells are present at the epithelium: ciliated bipolarolfactory receptors, sustentacular cells, microvillar cells, andbasal cells, beyond the Bowman gland.1,4–6

The olfactory system is composed by peripheral structuresof �6 million bipolar receptor cells located within the olfac-tory neuroepithelium.4,7 This ciliated bipolar olfactory recep-tor cell is a true bipolar neuron, projecting a single dendrite tothe surface of the olfactory neuroepitheliumand a single axonto the olfactory bulb. The dendrite extends to the epithelialsurface and has nonmotile cilia with membrane receptors,

Keywords

► olfactory mucosa► cerebrum► dementia► olfaction disorders► aging

Abstract Introduction Loss of smell is involved in various neurologic and neurodegenerativediseases, such as Parkinson disease and Alzheimer disease. However, the olfactory test isusually neglected by physicians at large.Objective The aim of this study was to review the current literature about therelationship between olfactory dysfunction and neurologic and neurodegenerativediseases.Data Synthesis Twenty-seven studies were selected for analysis, and the olfactorysystem, olfaction, and the association between the olfactory dysfunction and dementiaswere reviewed. Furthermore, is described an up to date in olfaction.Conclusion Otolaryngologist should remember the importance of olfaction evalua-tion in daily practice. Furthermore, neurologists and physicians in general shouldinclude olfactory tests in the screening of those at higher risk of dementia.

receivedOctober 16, 2013accepted after revisionJuly 27, 2014published onlineNovember 14, 2014



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where odor molecules bind.4 The axons form into bundles,called the olfactory fila, and project through the cribriformplate of the ethmoid bone and synapse at the olfactory bulb.In humans, the surface area of the cilia is �25 mm.1,8

Olfactory receptor neurons are surrounded by the susten-tacular cells, which probably contribute to regulation andconservation of the appropriate ionic environment aroundthe receptor neurons for adequate olfactory transduction.1,4

Microvillar cells presumably act as a second morphologi-cally distinct class of chemoreceptor, first described in 1982.However, their precise role in olfaction has not yet beenclearly demonstrated.4,9

The Bowman gland is a serous-producing tubuloalveolargland, another component of the lamina propria, composed ofsecretory acini with a duct that passes through the olfactoryepithelium.4,10 This secretion probably is essential for olfac-tory transduction.4

Recent studies have shown that the olfactory system dis-plays robust and functional neurogenesis throughout life,6 bycontaining cells with very broad developmental potency.Human olfactory mucosa yield neurospheres that could bepropagated as secondary and tertiary neurospheres.11 Theyare the basal cells, a well-recognized distinct population ofstem cells of the olfactory epithelium, capable of continuouslyregenerating olfactory receptor neurons throughout the lifespan.4,11,12 The basal cells havemultipotency and self-renew-al characteristics and are able to give rise experimentally toother neural and non-neural cells.4,6,13

Olfactory Bulb and Primary Olfactory CortexAs described previously, the single axons of the 10 to 20million olfactory receptor neurons join together into fasciclesand nerves, which pass through the 15 to 20 foramina of eachcribriform plate to synapse within the olfactory bulb.4,14

The olfactory bulb continues to the olfactory tract andarrives at the primary olfactory cortex areas, such as anteriorolfactory nucleus, entorhinal cortex, and amygdale.15,16

Olfactory Mucosa LocalizationTheoretically, the human olfactory mucosa constitutes 1.25%of the nasal mucosa, corresponding to an area of �2 cm2. Thenumber and density of bipolar olfactory neurons are probably�6 � 106 and 30,000/mm2, respectively.4,17

The olfactory epithelium in the human fetus is uniformlydistributedwithout interruption by respiratory epithelium ina continuous pattern.18 Féron and other authors have ob-served that the distribution of the olfactory epithelium inadult humans is frequently disrupted with interspersedpatches of respiratory epithelium.18–20 Comparison of fetaland adult samples suggests that invasion of respiratory tissueinto olfactory epithelium increaseswith age, as suggested in aprevious study of adult olfactory epithelium.18 This probablyoccurs because this mucosa is a dynamic structure withfeatures reflecting innate and environmental as well asdevelopmental influences. Thus, it can be assumed that theprecise location and the overall dimension of the neuro-epithelium may be different among individuals and maychange over time.4

Recent anatomical studies modified the previous conceptof distribution area of the olfactory mucosa. It was assumedthat the neuroepithelium was restricted to the area of thecribriform plate, superior turbinate, and the opposite supe-rior nasal septum.4 Leopold and other authors demonstratedthat the olfactory mucosa extends within the medial andanterior surface of the middle turbinate, either in the lateralor in themedialwall of the nasal cavity.21,22 Thiswould not bein conflict with the work of Biedlingmaier and Whelan23,since they looked at much more inferior middle turbinatetissue and did not find olfactory tissue.21

Discussion

Olfactory Deficit in Elderly PeopleA decay of the smell function occurs in old age. In fact, age isthe strongest correlate of olfactory decline in healthy adulthumans and has a much larger impact than even cigarettesmoking.1 These data are confirmed by cross-sectional andlongitudinal studies.24

Generally, olfactory impairment related to age is moresevere for men than for women, although individual differ-ences are present. This deficit often goes unnoticed and israrely investigated by physicians, unlike alterations in hear-ing and vision. About 2% of the population under 65 years ofage has olfactory impairment. This rises dramatically be-tween 65 and 80 years, with about half of the populationcomplaining of loss of smell. Over 80 years, smelling problemsare noticed by 75% of elderly.1

Possible reasons for smell changes related to age includeossification and closure of the foramina of the cribriformplate, age-related degenerative processes occurring in thebrain,1,24 and cumulative damage to the olfactory receptorsfrom different types of insults throughout life.1

Olfactory Deficit and DementiaRecently, the olfactory neuroepithelium has attracted therenewed interest of scientists, because the olfactory mucosahas the potential to be an early marker of neurodegenerativeconditions, such as schizophrenia, AD, multiple sclerosis, andPD.4

There is considerable variation in the prevalence andmagnitude of olfactory dysfunction among neurodegenera-tive diseases. In AD, PD, and Parkinson–dementia complex ofGuam, olfactory dysfunction is severe (University of Penn-sylvania Smell Inventory Test [UPSIT] scores under 20),whereas that of Huntington disease, multi-infract dementia,amyotrophic lateral sclerosis, and schizophrenia is moremoderate. Progressive supranuclear palsy is associated withminor changes in olfactory function, even though it sharesmajor clinical features with PD. These data suggest thatolfactory testing could help in differential diagnosis of severalneurodegenerative diseases.1

Olfactory deficit can be noticed in the ability to detect,recognize, and remember odorants in the elderly, particularlyin patients with AD.25 In the case of AD, smell problems occurin the beginning of the disease,26 and this pattern can reflectan “preclinical” period of disease development by preceding


Olfaction in Neurologic and Neurodegenerative Diseases Godoy et al. 177

the onset of classic disease symptoms.1 Moreover, olfactoryimpairment is mostly found in individuals at risk for AD,including subjects with mild cognitive impairment whoeventually develop AD, those with another potential riskfactor for AD (namely subjective memory complaints),25,27

and relatives of AD patients.7,27,28 Olfactory dysfunction inAD is associated with disease progression, can be helpful inthe differential diagnosis of major depression and AD, andmay have clinical value as an early diagnostic marker inpredicting incident AD in high-risk individuals.25 The pres-ence of apolipoprotein E-4 allele in an anosmic normalindividual increases the risk of having cognitive decline inthe future by 4.9-fold.1

Surprisingly, most patients with AD and PD are unaware oftheir olfactory loss before taking the test; 85 to 90% of patientsin the early stages present olfactory impairment, and thisdeficit is associated with decreased activation of central odorprocessing structures (as measured by functional imaging).1,15

In AD, neuropathologic changes within olfaction-relatedbrain regions usually accompany these olfactory changes. Oneof the pathologic hallmarks of AD, the neurofibrillary tangles(NFTs), have been identified within the olfactory bulb, olfac-tory tract, transentorhinal and entorhinal cortex, anteriorolfactory nucleus, and amygdale.25 The number of NFTswithin such regions has been correlated with the severityof dementia.15 Recent neuropathologic studies suggest thatAD-related pathology may begin within olfactory centralcortex and then spread tomultiple areas of the brain.15 Theseneuropathologic changes have been associatedwith impairedolfaction around the time of death in thosewithout dementiaor with mild cognitive impairment.24,27

As seen, olfactory impairment has been significantly as-sociatedwith the AD neuropathology burden in the brain andthe risk of future AD. Some animal models of AD-relatedneuropathologic changes have indicated a strong associationbetween NFT in the olfactory system and cognitive declineand a negative association between amyloid-β burden in thebrain, another hallmark of AD disease, and olfaction. More-over, oxidative damage in the olfactory epithelium is presentin the early stages of AD.27

Functional Magnetic Resonance Imaging in Patientswith ADWang demonstrated that functional magnetic resonanceimaging (fMRI) is sensitive to changes in olfactory functiondue to AD. Patients with AD have reduced BLOD (blood-oxygen-level dependent) signals in the hippocampus andinsula regions when compared with healthy control subjectsof similar age. Such alterations are significantly correlatedwith UPSIT, Mini–Mental State Examination, and CDR (clini-cal dementia rating) scores, proposing the significance ofolfactory fMRI in patients with AD.15

Furthermore,when the odorant concentration is increased10-fold, only slightlymore activation is induced in somebrainregions. These data about additional recruitment of activitysuggest that total anosmia in AD is not the rule but rather theexception, and they indicate that at least some residualcapacity is available.15,26 Experimental results indicate the

feasibility for using olfactory fMRI as a marker for diagnosisand evaluation of AD.15

Olfactory Epithelium BiopsyBecause the olfactorymucosa contains the only surface neuralcells of the body, the olfactory receptor neurons, someauthors call the neuroepithelium the “window to the brain.”Therefore, an olfactory biopsy could potentially help tounderstand what occurs in the brain with neurologic andneurodegenerative disease.4 Olfactory epithelium is accessi-ble for low-risk biopsy, allowing examination of the neuro-genesis process,6,18 as shown by neuropathologic studies onneurodegenerative diseases like AD and PD.18

Gender Differences in Human Olfactory Bulb

Recent studies indicate the existence of differences in humanolfactory bulb related to gender. Oliveira-Pinto et al demon-strate a sex-related difference in the absolute number of total,neuronal and non-neuronal cells, favoring women by 40-50%,which may have olfactory functional impact.29

Conclusions

Olfactory dysfunction can manifest in various degrees anddifficulties in discrimination, odor identification, and olfac-tory memory. Neurologic and neurodegenerative diseases,particularly AD and PD, aremajor causes of dysosmia. There ismuch evidence that olfactory tests can be used to differentiatediagnosis between PD and other types of parkinsonism.

This review shows that olfactory tests can be a useful toolin differential diagnosis of dementia and other diseases, aswell as among the various types of dementia, and indicatesthat impairment in olfactory discrimination can predictfuture cognitive decline.

However, some questions remain unanswered: ‘What isthe boundary between the olfactory changes related to agingand those caused by disease?’; ‘Is it possible to differentiatethem through the olfactory tests?’; ‘In which time the patho-logical changes of AD are installed in the olfactory struc-tures?’. More studies are necessary to clarify these issues..

Based on current knowledge of the association betweensmell and dementias, the authors suggest that the otolaryn-gologist should remember the importance of olfaction evalu-ation in daily practice. Furthermore, neurologists andphysicians in general should all include olfactory tests inthe screening those at higher risk of dementia.

References1 Doty RL. The olfactory system and its disorders. Semin Neurol

2009;29(1):74–812 Deems DA, Doty RL, Settle RG, et al. Smell and taste disorders, a

study of 750 patients from the University of Pennsylvania Smelland Taste Center. Arch Otolaryngol Head Neck Surg 1991;117(5):519–528

3 Doty RL. Clinical studies of olfaction. Chem Senses 2005;30(Suppl 1):i207–i209


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4 Escada PA, Lima C, da Silva JM. The human olfactory mucosa. EurArch Otorhinolaryngol 2009;266(11):1675–1680

5 Hahn CG, Han LY, Rawson NE, et al. In vivo and in vitro neuro-genesis in human olfactory epithelium. J Comp Neurol 2005;483(2):154–163

6 Ronnett GV, Leopold D, Cai X, et al. Olfactory biopsies demonstratea defect in neuronal development in Rett’s syndrome. Ann Neurol2003;54(2):206–218

7 Doty RL, Perl DP, Steele JC, et al. Olfactory dysfunction in threeneurodegenerative diseases. Geriatrics 1991;46(Suppl 1):47–51

8 Graves AB, Bowen JD, Rajaram L, et al. Impaired olfaction as amarker for cognitive decline: interaction with apolipoprotein Eepsilon4 status. Neurology 1999;53(7):1480–1487

9 Montani G, Tonelli S, Elsaesser R, Paysan J, Tirindelli R. Neuropep-tide Y in the olfactory microvillar cells. Eur J Neurosci 2006;24(1):20–24

10 Moulton DG, Beidler LM. Structure and function in the peripheralolfactory system. Physiol Rev 1967;47(1):1–52

11 Murrell W, Féron F, Wetzig A, et al. Multipotent stem cells fromadult olfactory mucosa. Dev Dyn 2005;233(2):496–515

12 Bartolomei JC, Greer CA. Olfactory ensheathing cells: bridging thegap in spinal cord injury. Neurosurgery 2000;47(5):1057–1069

13 Roisen FJ, Klueber KM, Lu CL, et al. Adult human olfactory stemcells. Brain Res 2001;890(1):11–22

14 Hadley K, Orlandi RR, Fong KJ. Basic anatomy and physiology ofolfaction and taste. Otolaryngol Clin North Am 2004;37(6):1115–1126

15 Wang J, Eslinger PJ, Doty RL, et al. Olfactory deficit detected byfMRI in early Alzheimer’s disease. Brain Res 2010;1357:184–194

16 Attems J, Jellinger KA. Olfactory tau pathology in Alzheimerdisease and mild cognitive impairment. Clin Neuropathol 2006;25(6):265–271

17 Moran DT, Rowley JC III, Jafek BW, Lovell MA. The fine structure ofthe olfactory mucosa in man. J Neurocytol 1982;11(5):721–746

18 Féron F, Perry C, McGrath JJ, Mackay-Sim A. New techniques forbiopsy and culture of human olfactory epithelial neurons. ArchOtolaryngol Head Neck Surg 1998;124(8):861–866

19 Morrison EE, Costanzo RM. Morphology of the human olfactoryepithelium. J Comp Neurol 1990;297(1):1–13

20 Kobal G, Van Toller S, Hummel T. Is there directional smelling?Experientia 1989;45(2):130–132

21 Leopold DA, Hummel T, Schwob JE, Hong SC, Knecht M, Kobal G.Anterior distribution of human olfactory epithelium. Laryngo-scope 2000;110(3 Pt 1):417–421

22 Nibu K, Li G, Zhang X, et al. Olfactory neuron-specific expression ofNeuroD in mouse and human nasal mucosa. Cell Tissue Res 1999;298(3):405–414

23 Biedlingmaier JF,Whelan P, Zoarski G, RothmanM. Histopathologyand CT analysis of partially resected middle turbinates. Laryngo-scope 1996;106(1 Pt 1):102–4

24 Wilson RS, Yu L, Schneider JA, Arnold SE, Buchman AS, Bennett DA.Lewy bodies and olfactory dysfunction in old age. Chem Senses2011;36(4):367–373

25 Peters JM, Hummel T, Kratzsch T, Lötsch J, Skarke C, Frölich L.Olfactory function in mild cognitive impairment and Alzheimer’sdisease: an investigation using psychophysical and electrophysio-logical techniques. Am J Psychiatry 2003;160(11):1995–2002

26 Doty RL. Studies of human olfaction from the University ofPennsylvania Smell and Taste Center. Chem Senses 1997;22(5):565–586

27 Sohrabi HR, Bates KA, Weinborn MG, et al. Olfactory discrimina-tion predicts cognitive decline among community-dwelling olderadults. Transl Psychiatr 2012;2:e118

28 Doty RL. The olfactory vector hypothesis of neurodegenerativedisease: is it viable? Ann Neurol 2008;63(1):7–15

29 Oliveira-Pinto AV, Santos RM, Coutinho RA, et al. Sexual dimor-phism in the human olfactory bulb: females have more neuronsand glial cells than male. Plos One (In Press)


Olfaction in Neurologic and Neurodegenerative Diseases Godoy et al. 179

Ortner’s Syndrome: Secondary LaryngealParalysis Caused by a Great Thoracic AortaAneurysmAna Claudia Alves Zangirolami1 Frederico Vieira de Oliveira1 Miguel Soares Tepedino1

1Department of Otorhinolaryngology, Policlinica de Botafogo, Rio deJaneiro, Brazil


Address for correspondence Ana Claudia Alves Zangirolami, MD,Department of Otorrinolaringologia, Policlinica de Botafogo, AvPasteur 72, 1andar, Rio de Janeiro 22290240, Brazil(e-mail: [email protected]).

Introduction

Ortner’s syndrome, or cardiovocal syndrome, was introducedby Norbert Ortner in 1897, who described three cases of leftvocal fold immobility due to the recurrent compression of thelaryngeal nerve, caused by dilatation of the left atrium inpatients with mitral valve stenosis.1 Since then, the termOrtner’s syndrome has been used to describe any nonmalig-nant intrathoracic heart condition that results in recurrentcompression of the laryngeal nerve. The possible causes areleft ventricular failure, atrial septum failure, arterial canal

persistency, primary pulmonary hypertension, pulmonaryartery relapsing embolism, left ventricular aneurysm, andother types of aortic aneurysm that cause vocal fold paralysis.Thoracic aortic artery aneurysm represents 5% of the cases.2,3

The prevalence of the cardiac alterations that characterizeOrtner’s syndrome is not discussed in the literature, becauseitsmanifestation is linked tomany causes and depends on thedirect compression of the nerve. The left recurrent laryngealnerve is the most frequently affected, due to its longer coursebypassing the aortic arch, comparedwith the right one,whichgoes around the subclavian artery.

Keywords

► vocal cord palsy► arch of the aorta► dysphonia

Abstract Introduction Recurrent laryngeal nerve injury caused by cardiovascular disease is arare condition, and often it is the only prominent sign of an imminent break of an aorticartery aneurysm.Objective To report left laryngeal paralysis caused by a great aortic arch aneurysm andto highlight the importance of an otorhinolaryngologic evaluation along with a thoracicradiologic study.Resumed Report A 42-year-old man complained of thickness of his voice anddysphagia for 3 months, but no thoracic pain or other relevant complaints. Videolaryngoscopy revealed immobility of his left vocal fold in the paramedian position.Imaging was obtained for investigation, including magnetic resonance imaging of histhorax, which showed a fusiform aneurysm in the aortic arch, leading to recurrentcompression of the left laryngeal nerve. The patient was successfully treated withendovascular repair of the aneurysm. At 2-month follow-up, there was still no recoveryof the laryngeal mobility.Conclusion An aortic artery aneurysm can suddenly break, requiring emergency heartsurgery, and the results can be fatal inmany cases.We suggest routine exam of the vocalfolds in all patients with a heart condition, and we review the literature and suggest theuse of imaging to reduce the number of emergency procedures.

receivedJune 3, 2014acceptedNovember 3, 2014published onlineJanuary 5, 2015



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Literature Review

The vocal fold immobility in Ortner’s syndrome can beassociated with involvement of the vagal nerve or the recur-rent laryngeal nerve, from the jugular foramen to the en-trance of the larynx. It has many etiologies according to thelesion’s location, including trauma during thoracic (53.1%)and heart surgeries (28.5%), extended intubation (16.7%),neoplasms (32%), central nervous system dysfunction (8%),and metabolic and infectious (3%), idiopathic (16%), andtraumatic (11%) causes.2 Aortic aneurysm is an uncommoncause for vocal cord paralysis, described in only 5% of thepatients with this vascular condition, and the involvement ofthe ascendant aorta and the aortic arch is also infrequent.2–4

Case Report

A 42-year-old man born in Rio de Janeiro presented to theOtorhinolaryngologic Service complaining of dry cough andthickness of his voice of 3 months’ duration, combined withprogressive dysphagia from solid to liquid. Otorhinolaryngologicphysical exam discovered no alterations. Indirect laryngoscopyrevealed left vocal fold paralysis at the paramedian position(►Fig. 1). Thoracic X-ray was requested, which showed a bulgeadjacent to the aortic area; neck and cranium computerizedtomography showed no alterations.Magnetic angioresonance ofthe mediastinum revealed a fusiform aneurysm of the thoracicaorta with laminar mural thrombi, measuring 7 cm in diameterand extending �9.5 cm, with no evidence of dissection. Theheart had normal dimensions (►Figs. 2, 3, and 4). These resultsindicated left recurrent laryngealnerveparalysis bycompressionof the aortic artery. The patient was referred to the cardiovascu-lar surgeon for surgical treatment of the aortic aneurysm, whichwas accomplished by means of endovascular surgery. Twomonths postoperatively, the patient still had dysphonia andvocal fold immobility.

Discussion

The left recurrent laryngeal nerve, due to its intimate ana-tomic relation to the aortic arch, left lung apex, trachea,esophagus, left pulmonary artery, and mediastinal lymphnodes, is particularly vulnerable to lesions. Themost frequentcauses of paralysis are tumors of the lung, esophagus, andthyroid. It has also been reported as a possible complicationfrom other cardiovascular diseases, including pulmonary

Fig. 1 Laryngoscopy image showing paramedian left vocal foldparalysis.

Fig. 2 Magnetic resonance T1-weighted image, coronal section,showing aneurysmal dilatation of the aorta arch.

Fig. 3 MRI examination T1-weighted image, examination in obliqueprofile for rotational rebuild with paramagnetic contrast, showing afusiform aneurysm extending from the post emerging segment of thesubclavian artery and demonstrating mural thrombi and misuse ofadjacent structures. Note also topography of vagus nerve. Abbrevia-tion: A, anterior; F, feet; H, head; L, left; MIP, maximum intensityprojection; P, posterior.


Ortner’s Syndrome Zangirolami et al. 181

hypertension, interatrial communication, and patency of thearterial canal.5,6 The mechanism of left recurrent laryngealnerve paralysis remains unclear. However, it is assigned to thecompression of this nerve, which can get hooked around theligament arteriosus between the pulmonary artery and theaortic aneurysm. Recurrent laryngeal nerve paralysis inpatients with cardiovascular diseases demands a thoroughexamination, because it suggests possible dilatation of theaneurysm.7 For diagnosis, in addition to the anamnesis andthe physical examination, imagingmust be used to delimitatethe location and diameter of the aneurysm, with thoraciccomputed tomography and magnetic resonance able to senselesions nearly 100% of the time. It is also vital that in patientswith laryngeal complaints (vocal, breathing, or swallowingdisturbances), a videolaryngoscopy is used first to identifythe vocal fold position , if there is an intrinsic lesion, imagingis used to investigate the entire neural path of the vagal nerve,from its emergence on the cranium base to the recurrentlaryngeal nerve emission in the thorax.8 Recurrent laryngealnerve paralysis prognosis depends on the level and timeelapsed from the compression of the nerve. Stoob et alreported the first case of curing the laryngeal paralysis of apatient 1 year after surgically correcting the aneurysm byendovascular means.4However, in most of the cases reportedin literature, there was no resolution of the thickness of the

voice. Deaths by surgical complications or breaking of theaneurysm were also reported.2,3,9 Patients with laryngealparalysis in the paramedian position must be submitted tospeech therapy to use the contralateral musculature to com-pensate for the glottal failure. Surgical treatment can beindicated for patients with important refractory vocal symp-toms on speech therapy or for those with imminent risk ofbronchoaspiration.10,11

Final Comments

An aortic artery aneurysm can suddenly break, indicatingemergency heart surgery, which often has fatal results. Wesuggest routine exam of the vocal folds in all patients with aheart condition.

References1 Chan P, Lee CP, Ko JS, et al. Cardiovocal (Ortner’s) syndrome: left

recurrent laryngeal nerve palsy associated with cardiovasculardiseases. Eur J Med 2004;11:69–70

2 Yuan SM. Hoarseness subsequent to cardiovascular surgery, inter-vention, maneuver and endotracheal intubation: the so-callediatrogenic Ortner’s (cardiovocal) syndrome. Cardiol J 2012;19(6):560–566

3 Ohki M. Thoracic saccular aortic aneurysm presenting with recur-rent laryngeal nerve palsy prior to aneurysm rupture: a prodromeof thoracic aneurysm rupture? Case Rep Otolaryngol 2012;2012:367873

4 Camishiou RC, Gibbon JH, Pierucci L, et al. Paralysis of the leftrecurrent laryngeal nerve secondary to mitral value disease:report of two cases and literature review. Ann Surg 1966;163(6):818–827

5 Fennessy BG, Sheahan P, McShane D. Cardiovascular hoarseness:an unusual presentation to otolaryngologists. J Laryngol Otol2008;122(3):327–328

6 Stoob K, Alkadhi H, Lachat M, Wildermuth S, Pfammatter T.Resolution of hoarseness after endovascular repair of thoracicaortic aneurysm: a case of Ortner’s syndrome. Ann Otol RhinolLaryngol 2004;113(1):43–45

7 Gupta P, Sharma S. Ortner’s syndrome secondary to aortic aneu-rysm. Ann Acad Med Singapore 2012;41(1):40–41

8 Gulel O, Elmali M, Demir S, Tascanov B. Ortner’s syndromeassociated with aortic arch aneurysm. Clin Res Cardiol 2007;96(1):49–50

9 Subramaniam V, Herle A, Mohammed N, Thahir M. Ortner’ssyndrome: case series and literature review. Braz J Otorhinolar-yngol 2011;77(5):559–562

10 Mathai J, Swapna UP. Hoarseness -as a presenting feature of aorticarch aneurysm. Indian J Otolaryngol Head Neck Surg 2006;58(3):309–310

11 Veldtman GR, Dearani JA, Warnes CA. Low pressure giantpulmonary artery aneurysms in the adult: natural history andmanagement strategies. Heart 2003;89(9):1067–1070

Fig. 4 MRI examination T1-weighted image, coronal cut for rotationalrebuild with paramagnetic contrast, showing a fusiform aneurysmextending from the post emerging segment of the subclavian arteryand demonstrating mural thrombi and misuse of adjacent structures.Note also topography of vagus nerve. Abbreviation: PL, posterior left;PR, posterior right.


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Intralabyrinthine Penetrating Ventilation Tubewith Preservation of Hearing: An UnusualClinical SituationTantely Razafimahefa Raoelina1 Maya Elziere1 Justin Michel2 Arnaud Devèze1,3

1Department of Otolaryngology and Skull Base Surgery, UniversityHospital Nord, Marseille, France

2Department of Otolaryngology and Skull Base Surgery, La TimoneUniversity Hospital, Marseille, France

3Laboratory of Applied Biomechanics IFSTTAR, Aix MarseilleUniversity, Bd Pierre Dramard, Marseille, France


Address for correspondence Arnaud Devèze, MD, PhD, Department ofOtolaryngology and Skull Base Surgery, University Hospital Nord,Chemin des bourrely, Marseille 13015, France(e-mail: [email protected]; [email protected]).

Introduction

Traumatic perilymphatic fistula is not a rare event with regardsto sport activities or traffic accident. However, iatrogenic dam-age to the inner ear can occur following the common use ofgrommets and ventilation tube insertion. Here,we describehowearly surgical management may help recover the inner earfunction, despite severe damage to this fragile structure.

Review of Literature with DifferentialDiagnosis

A perilymph fistula is an abnormal communication betweeninner ear perilymphatic spaces to the aerated middle ear. A

wide variety of signs and symptoms as well as numerousetiologies are associated with perilymph fistula, from congen-ital malformations to accidental or iatrogenic trauma to theinner ear.1–3 During the past 30 years, several cases of trau-matic perilymphatic fistula have been reported. Those includecases associated with ear cleansing, surgeries, trauma, andblast.2–11 Traumatic perilymph fistula following penetratingintravestibular ventilation tube has yet to be reported.

Case Report

A 62-year-old woman with a past history of chronic otitisunderwent a tube placement for a right-sided posterior

Keywords

► labyrinth► trauma► tympanic membrane

rupture► perilymph► stapes surgery

Abstract Introduction Traumatic perilymphatic fistula is not a rare event with regards to sportactivities or traffic accident. However, iatrogenic damage to the inner ear can occurfollowing the common use of grommets and ventilation tube insertion.Objectives To report an unusual case of insertion of aeration tube into the vestibuletrough the stapes footplate.Resumed Report A 62-year-old woman experienced iatrogenic penetration into thevestibule from a ventilation tube inserted for retraction pocket management. The eventwas misdiagnosed both by the surgeon and by the emergency room physicians, leadingto delay in the management. However, preservation of the hearing function lasted for2 weeks prior to deafness, thanks to the valve of Bast, which preserved the cochlearfluid.Conclusion This case gives us the opportunity to stress the need for systematic clinicalexamination of traumatic injury to the ear and to recommend performing multiplanarmillimetric computed tomography scan with accurate interpretation. Traumatic injuriesshould be referred to a dedicated traumatic emergency referral center.

receivedMay 5, 2014acceptedJune 7, 2014published onlineDecember 12, 2014



THIEME

Case Report 183

retraction pocket. Immediately after the procedure, shedeveloped severe vertigo associated with nausea and vomit-ing. Shewas dischargedwith antivomitingmedication, whichwas said to be a normal postoperative event. Four days latershe was admitted in emergency for acute vertigo. Clinicalevaluation revealed right index finger deviation and leftbeating nystagmus with hearing loss. Otoscopy showed thetube in place. A temporal bone computed tomography (CT)scan was performed and showed the transtympanic ventila-tion tube associated with middle ear effusion, but bonycomponents within the vestibule were missed (►Fig. 1).She was admitted in the otolaryngology department andmanaged medically. Her condition improved, and she wasdischarged 3 days later.

One week later, she was referred to our tertiary referralcenter complaining of progressive right-sided hearing lossassociated with severe vertigo. She presented with right-sided vestibular impairment and positive Hennebert signon the right ear. On otoscopic examination, she had tympanicmembrane retraction with the ventilation tube inserted inthe posterosuperior quadrant with pulsating waterlikefluid leaking through the tube. The audiometry showed aright-sided, severe mixed hearing loss (►Fig. 2). A new

Fig. 1 High-resolution temporal computed tomography (with 0.6-mmslice thickness) in axial (A) and coronal reconstruction (B). The middleear is filled with fluid; the ossicular chain is not recognizable. There isno pneumolabyrinth, but careful examination may have revealed high-density components within the vestibule (A, arrow). The tube is seen inthe external canal (B, arrowhead).

Fig. 2 Right-ear pure tone audiogram performed 1 week afterdischarge from postemergency hospitalization showing severe mixedhearing loss. Dotted line: bone conduction; plain line: air conduction.

Fig. 3 Second computed tomography scan. The presence of air withinthe inner ear space (arrowhead) permits the visualization of the tubethat penetrates into the vestibule (arrow).


Intralabyrinthine Penetrating Ventilation Tube with Preservation of Hearing Raoelina et al.184

high-resolution temporal bone CTscan clearly showed a rightpneumolabyrinth within the vestibule and the semicircularcanals, sparing the cochlea, associated with the ventilationtube penetrating the vestibule through the oval window(►Figs. 3 and 4). She was diagnosed with right iatrogenicstapes trauma with perilymph fistula and urgently taken tothe operating room. The long process of the incus as well asthe stapes posterior crura and the posterior and inferior part

of the footplate were fractured. The rest of the footplate wasintact. The tubewas penetrating the vestibule in the posteriorand superior part of the stapes footplate. The tube was gentlyremoved, and the vestibule was refilled with saline solution.The vestibulewas sealedwith perichondriumandmaintainedwith fibrin glue; the whole montage was stabilized using aSilastic (Dow Corning, USA) sheet positioned from the facialcanal to the promontory. The tympanic membrane wasrepaired using underlay tympanoplasty with cartilage. Theossicular chain was not reconstructed.

In the immediate postoperative period, the patient stillpresented positional dizziness as well as a left-sided beatingnystagmus. Bone conduction auditory testing at postopera-tive day 2 showed marked improvement of the right-sidethresholds (►Fig. 5). The patient was discharged 5 days aftersurgery. On day 12 after surgery, she suddenly complained ofworsening hearing loss and severe tinnitus. The pure toneaverage (PTA) showed complete anacusis on the right ear(►Fig. 5). No revision surgery was indicated. Treatment withvestibular rehabilitation was introduced and helped dispelthe dizziness within a 6-month period of time.

Discussion

Our case is singular for the three following points: theerroneous placement of a tube under general anesthesia;the missed diagnosis in the emergency setting; and therelative conservation of the cochlear function despite severetrauma to the labyrinth. This is, to our knowledge, the firstdirect penetration of an aeration tube into the vestibule. Wefound another case of labyrinthine injury following tubeinsertion, due to gradual bony erosion of the osseous laby-rinth as a long-term complication.12

Fig. 4 Second computed tomography scan, coronal view. The pneu-molabyrinth extended into the vestibule and the superior and lateralcanals (arrows) but did not extend into the cochlea (arrowhead) andmight have explained both the conservation of the auditory functionup to the 2 weeks after injury as well as the initial recuperation ofhearing function after the surgery.

Fig. 5 Right-ear pure tone audiogram done 4 days (A) and 12 days (B) after surgical intervention. (A) Note the hearing improvement in boneconduction. (B) Complete anacusis occurred 12 days after the surgery. Dotted line: bone conduction; plain line: air conduction. Abbreviations: RE,right ear; LE, left ear.


Intralabyrinthine Penetrating Ventilation Tube with Preservation of Hearing Raoelina et al. 185

First, the trauma undoubtedly followed immediate mis-placement of the tube, in the posterosuperior quadrant of thedrum. This is clearly a wrong procedure, demonstrating aninsufficient and inappropriate education. Second, the workupperformed in the emergency room was inadequate. Both theclinical history and the CTscan clearly should have stressed thesuspicion of trauma to the inner ear. The presence of apneumolabyrinth on a high-resolution CTscan of the temporalbone is pathognomonic of perilymphatic fistula.8,10–12 Thisinappropriate diagnosis highlights the necessity of appropriateeducation in trauma-related injuries. Education, reconfigura-tion of trauma services, and better provision of neurocriticalcare facilities do improve the quality of care to trauma-relatedpatients.13,14 Although the initial CT scan was difficult tointerpret, the high-density spots within the vestibule shouldhave drawn attention to a possible severe inner ear trauma(►Fig. 1A). High-resolution CT scan of temporal bone ismandatory in such a scenario.5,6,15–17 Finally, it was unexpect-ed that the pneumolabyrinth did not extend into the cochlea(►Fig. 4), but we think that the preservation of the cochlearfluid explains the conservation of the sensorineural compo-nent of thehearing. Theprotective role of the valve of Bast (thatcloses the utricular duct) explains this unusual situation:decreasing pressure in the whole endolymphatic system col-lapses theductus reuniens and causes closure of thevalve.18–21

It is plausible that the reopening of the valve of Bast after therepair of the fistula may have let the remaining air bubbles tdiffuse within the cochlea and induce total auditory loss.

Final Comments

Tympanic tubes should be inserted in all cases in the infero- oranteroinferior quadrant of the drum, and this procedureshould be performed under local anesthesia. In cases ofsuspicion of traumatic damage to the stapes footplate, sur-gery should be performed as fast as possible followingtrauma. Although the outcome of vestibular symptoms isgood, regardless of lesion, severity, or intervention,3,6hearingoutcome is more unpredictable. We agree with Hidaka et alon the following predictive factors of hearing preservation ina scenario of pneumolabyrinth: (1) early intervention within15 days; (2) pneumolabyrinth limited to semicircular canalsand vestibule; (3) existence of stapes lesion.3 According toTsubota et al, the value of the bone conduction thresholdsshould be considered a significant prognostic factor as well.6

References1 Calmels MN, Deguine O. Perilymphatic fistula. Ear Nose Throat J

2004;83(10):6662 Al Felasi M, Pierre G, Mondain M, Uziel A, Venail F. Perilymphatic

fistula of the roundwindow. Eur Ann Otorhinolaryngol Head NeckDis 2011;128(3):139–141

3 Hidaka H, Miyazaki M, Kawase T, Kobayashi T. Traumatic pneumo-labyrinth: air location and hearing outcome. Otol Neurotol 2012;33(2):123–131

4 Silverstein H, Fabian RL, Stoll SE, Hong SW. Penetrating wounds ofthe tympanic membrane and ossicular chain. Trans Am AcadOphthalmol Otolaryngol 1973;77(3):ORL125–ORL135

5 Kojima H, Tanaka Y, Mori E, Uchimizu H, Moriyama H. Penetratingvestibular injury due to a twig entering via the external auditorymeatus. Am J Otolaryngol 2006;27(6):418–421

6 TsubotaM, Shojaku H,Watanabe Y. Prognosis of inner ear functionin pneumolabyrinth: case report and literature review. Am JOtolaryngol 2009;30(6):423–426

7 Hatano A, Rikitake M, Komori M, Irie T, Moriyama H. Traumaticperilymphatic fistula with the luxation of the stapes into thevestibule. Auris Nasus Larynx 2009;36(4):474–478

8 Rother T, Albrecht C, Issing PR. Pneumolabyrinth after cochlearimplantation in large vestibular aqueduct syndrome: a case report.Am J Otolaryngol 2011;32(5):430–432

9 Kaffel N, Jlassi N, Selmi Z, et al. [Traumatic perilymphatic fistulae :about 13 cases]. Tunis Med 2011;89(5):471–475

10 Mandalà M, Colletti L, Carner M, et al. Pneumolabyrinth andpositional vertigo after stapedectomy. Auris Nasus Larynx 2011;38(4):547–550

11 Prisman E, Ramsden JD, Blaser S, Papsin B. Traumatic perilym-phatic fistula with pneumolabyrinth: diagnosis and management.Laryngoscope 2011;121(4):856–859

12 Hajiioannou JK, Bathala S, Marnane CN. Case of perilymphaticfistula caused by medially displaced tympanostomy tube. J Lar-yngol Otol 2009;123(8):928–930

13 Hashmi ZG, Haider AH, Zafar SN, et al. Hospital-based traumaquality improvement initiatives: first step toward improvingtrauma outcomes in the developing world. J Trauma Acute CareSurg 2013;75(1):60–68, discussion 68

14 Smith NC, Findlay GP, Weyman D, Freeth H. The management oftrauma victims with head injury: a study by the National Confi-dential Enquiry into Patient Outcome and Death. Ann R Coll SurgEngl 2013;95(2):101–106

15 Adil EA, Choudhary AK, Moser KW, Ghossaini SN. Vestibularpneumolabyrinth: whyassessment with temporal bone computedtomography utilizingdynamic focal spotmode is important for thediagnosis. Emerg Radiol 2011;18(1):43–45

16 Mafee MF, Valvassori GE, Kumar A, Yannias DA, Marcus RE.Pneumolabyrinth: a new radiologic sign for fracture of the stapesfootplate. Am J Otol 1984;5(5):374–375

17 Achache M, Sanjuan Puchol M, Santini L, et al. Late pneumo-labyrinth after undiagnosed post-traumatic perilymphatic fistula.Case report illustrating the importance of systematic emergencymanagement. Eur Ann Otorhinolaryngol Head Neck Dis 2013;130(5):283–287

18 Bast TH. The utriculo-endolymphatic valve. Anat Rec 1928;40:61–65

19 Bast TH. Function of the utriculo-endolymphatic valve. Two casesof ruptured saccules in children. Arch Otorhinolaryngol 1934;19:537–550

20 Bachor E, Karmody CS. The utriculo-endolymphatic valve inpediatric temporal bones. Eur Arch Otorhinolaryngol 1995;252(3):167–171

21 Hofman R, Segenhout JM, Buytaert JA, Dirckx JJ, Wit HP. Morphol-ogy and function of Bast’s valve: additional insight in its function-ing using 3D-reconstruction. Eur Arch Otorhinolaryngol 2008;265(2):153–157


Intralabyrinthine Penetrating Ventilation Tube with Preservation of Hearing Raoelina et al.186

A Rare Location of Angiofibroma in the InferiorTurbinate in Young WomanAsif Salimov1 Serdar Ozer1

1Department of Otolaryngology, Hacettepe University Hospital,Ankara, Turkey


Address for correspondence Asif Salimov, MD, Department ofOtolaryngology, Hacettepe University Hospital, Sihhiye, Ankara06100, Turkey (e-mail: [email protected];[email protected]).

Introduction

Juvenile nasopharyngeal angiofibroma (JNA) is a rare benignneoplasm in the nasopharynx. The tumor tends to be locallyaggressive and is typically seen in adolescent boys. It arises fromthesuperiormarginof thesphenopalatine foramenandaccountsfor 0.05% of all head and neck neoplasms. The typical histopath-ologic appearance of JNA is numerous wide, irregular vesselswith a single layer of endothelial cells embedded in fibrousstroma. The abundant vascular component is responsible for theexcessive bleeding during surgery or following biopsies.1

Extranasopharyngeal angiofibromas (ENAs) have beenreported sporadically in the literature. They most commonlyoriginate from the maxillary sinus, and women are affected.Compared with nasopharyngeal fibromas, the lesion is diag-nosed earlier, is less vascularized, and occurs in older pa-tients.2 Similar to JNAs, the treatment of choice for ENAs issurgery. Radiotherapy may be used for unresectable lesions.The surgical approach is tailored to the location and size oftumor.1 In our case, the tumor was small enough for totalexcision with an endonasal endoscopic approach. This is thefirst case report in the English literature of angiofibroma

Keywords

► extranasopharyngealangiofibroma

► epistaxis► inferior turbinate

Abstract Introduction Juvenile nasopharyngeal angiofibroma is a rare benign neoplasm in thenasopharynx. The tumor tends to be locally aggressive and is typically seen inadolescent boys. Extranasopharyngeal angiofibromas have been reported sporadicallyin the literature. They most commonly originate from the maxillary sinus.Objectives A 26-year-old woman was referred to our clinic with intermittent epistaxisfrom the right nasal passage for the previous 2 months. Maxillofacial magneticresonance imaging showed a lobular, contouredmass originating from the right inferiorturbinate and hanging in the right nasal cavity, with dense contrast enhancementdenoting hypervascularity.Resumed Report Vascular feeding of the mass was seen from the right internalmaxillary artery with angiography, and this branch was embolized. On the following day,the patient underwent transnasal endoscopic excision of the mass. An approximately 3-cm-diameter mass was excised by partial turbinectomy, and the posterior edge of theremaining turbinate was cauterized.Conclusion Extranasopharyngeal angiofibromas are rarely seen, and the inferiorturbinate is an extremely rare location for them. This young woman is the first casereported in the English literature of angiofibroma originating from the inferior turbi-nate. We should consider these neoplasms can be found in female, nonadolescentpatients with extranasopharyngeal localization, and we should not perform biopsybecause of its massive bleeding.

receivedMay 29, 2014acceptedNovember 25, 2014published onlineDecember 29, 2014



THIEME

Case Report 187

originating in the inferior turbinate in a young woman.Preoperative embolization makes surgery more feasible.

Review of the Literature with DifferentialDiagnosis

ENAs are rarely seen, and the inferior turbinate is an ex-tremely rare location for them. There are reports in theEnglish literature of inferior turbinate angiofibroma in malepatients.3–6 In female patients, there is only one case report inthe English literature of inferior turbinate angiofibroma in a52-year-old woman.7 Recently, a report was published of a�9-year-old girl with angiofibroma obstructing the nasalcavity and originating from the inferior turbinate.8

Differential diagnosis includes fibrosed antrochoanal andethmoidal polyp and other fibrovascular tumors, such ascapillary hemangioma, hemangiopericytoma, and solitaryfibrous tumor.3

Case Report

A 26-year-old womanwas referred to our clinic with intermit-tent epistaxis from the right nasal passage for the previous2 months. Physical examination revealed a mass originatingfrom the posterior right inferior turbinate. Maxillofacial mag-netic resonance imaging showed a lobular, contoured massoriginating from the right inferior turbinate and hanging to theright of the nasal cavity, with dense contrast enhancementdenoting hypervascularity (►Figs. 1, 2, and 3). We decided toembolize the vascular feeding of the mass to make surgerymore feasible. The patient consultedwith the Invasive VascularRadiology Department and decided on the embolization pro-cess under intravenous sedation. Vascular feeding of the masswas seen from the right internal maxillary artery, and this

branch was embolized. The following day the patient under-went transnasal endoscopic surgery for excision of the mass.The procedure was performed under general anesthesia start-ing with uncinectomy. After that, the maxillary sinus ostiumwas found andwidened.When looking at the posterior wall ofthe sinus and pterygopalatine fossa, nomasslike structurewasevident. The approximately 3-cm-diameter mass originatedfrom theposterior edge of the right inferior turbinate (►Fig. 4).A partial turbinectomywas performed, and the posterior edgeof the remaining turbinate was cauterized. The operationended with a sponge gel filling in the right middle meatus.

Fig. 1 Preoperative maxillofacial magnetic resonance imaging (cor-onal T1-weighted). Arrow shows the mass originating from the rightinferior turbinate.

Fig. 2 Preoperative maxillofacial magnetic resonance imaging (axialT1-weighted). Arrow shows the mass originating from the right inferiorturbinate.

Fig. 3 Preoperative maxillofacial magnetic resonance imaging (axialT1-weighted postcontrast). Arrow shows dense contrast enhancementof the mass.


Angiofibroma in the Inferior Turbinate Salimov, Ozer188

There was no evidence of recurrence in the following13 months.

Discussion

A mean age of 20 to 30 years in ENAs was found in theliterature.1,2,9 Our case was also in this group.

ENAsmostly originate from themaxillary sinus.2 There arealso some reports of tumors located in the ethmoid sinus,nasal cavity, nasal septum, larynx, sphenoid sinus, cheek,conjunctiva, oropharynx, retromolar area, and middleturbinate.3,10–16

The clinical presentation of ENAs depends on tumorlocalization. In our case, the clinical presentation was similarto JNAs. But because of the limited space in the nasal cavity,diagnosis was made at an early tumor stage.

The cause of inferior turbinate angiofibroma is not wellunderstood. The tumor’s location indicated that the originmaybe from ectopic tissues located further from its usual place.17

There is also a reported case of an angiofibroma arisingfrom the inferior turbinate after CO2 laser turbinoplasty.18 Inour case, there was no history of turbinate surgery.

Selective angiography is a useful diagnostic method todemonstrate tumor vascular composition and to confirm thediagnosis. It also allows tumor embolization, which reducesintraoperative bleeding.1

Final Comments

ENA in a young woman is a very rare clinical entity. Endoscopicand radiologic examination is important, butdefinitivediagnosisis made by histopathologic analysis. We should consider theseneoplasms possibly in female patients, at every age, with extra-nasopharyngeal localization, and we should not perform biopsy

because of massive bleeding. Preoperative embolization of thevessels makes surgery more feasible.

References1 Szymańska A, Szymański M, Morshed K, Czekajska-Chehab E,

Szczerbo-Trojanowska M. Extranasopharyngeal angiofibroma:clinical and radiological presentation. Eur Arch Otorhinolaryngol2013;270(2):655–660

2 Windfuhr JP, Remmert S. Extranasopharyngeal angiofibroma:etiology, incidence and management. Acta Otolaryngol 2004;124(8):880–889

3 Nomura K, Shimomura A, Awataguchi T, Murakami K, Kobayashi T.A case of angiofibroma originating from the inferior nasal turbi-nate. Auris Nasus Larynx 2006;33(2):191–193

4 Celik B, Erisen L, Saraydaroglu O, Coskun H. Atypical angiofibro-mas: a report of four cases. Int J Pediatr Otorhinolaryngol 2005;69(3):415–421

5 Taggarshe D, Quraishi MS, Dugar JM. Inferior turbinate angiofi-broma: an atypical presentation [correction of preservation].Rhinology 2004;42(1):45–47

6 Gaffney R, Hui Y, Vojvodich S, Forte V. Extranasopharyngealangiofibroma of the inferior turbinate. Int J Pediatr Otorhinolar-yngol 1997;40(2–3):177–180

7 Lee JH, Jeong HM. Extranasopharyngeal angiofibroma originatingin the inferior turbinate. Ear Nose Throat J 2013;92(9):E31–E32

8 Baptista MAFB, Pinna FR, Voegels RL. Extranasopharyngeal angio-fibroma originating in the inferior turbinate: a distinct clinicalentity at an unusual site. Int Arch Otorhinolaryngol 2014;18:403–405

9 Huang RY, Damrose EJ, Blackwell KE, Cohen AN, Calcaterra TC.Extranasopharyngeal angiofibroma. Int J Pediatr Otorhinolaryngol2000;56(1):59–64

10 Somdas MA, Ketenci I, Unlu Y, Canoz O, Guney E. Extranasophar-yngeal angiofibroma originating from the nasal septum. Otolar-yngol Head Neck Surg 2005;133(4):647

11 Dere H, Ozcan KM, Ergul G, Bahar S, Ozcan I, Kulacoglu S. Extra-nasopharyngeal angiofibroma of the cheek. J Laryngol Otol 2006;120(2):141–144

12 Tasca I, Compadretti GC. Extranasopharyngeal angiofibroma ofnasal septum. A controversial entity. Acta Otorhinolaryngol Ital2008;28(6):312–314

13 Mohindra S, Grover G, Bal AK. Extranasopharyngeal angiofibromaof the nasal septum: a case report. Ear Nose Throat J 2009;88(11):E17–E19

14 Tsunoda A, Kohda H, Ishikawa N, Komatsuzaki A. Juvenile angiofi-broma limited to the sphenoid sinus. J Otolaryngol 1998;27(1):37–39

15 Steele MH, Nuss DW, Faust BF. Angiofibroma of the larynx: reportof a case with clinical and pathologic literature review. Head Neck2002;24(8):805–809

16 Handa KK, Kumar A, Singh MK, Chhabra AH. Extranasopharyngealangiofibroma arising from the nasal septum. Int J Pediatr Otorhi-nolaryngol 2001;58(2):163–166

17 Perić A, Baletić N, Cerović S, Vukomanović-Durdević B. Middleturbinate angiofibroma in an elderly woman. Vojnosanit Pregl2009;66(7):583–586

18 Kang JW, Kim YH, Kim JH. Angiofibroma of inferior turbinate as anunusual complication of CO2 laser turbinoplasty. J Craniofac Surg2013;24(5):e513–e514

Fig. 4 Intraoperative view of the lesion.


Angiofibroma in the Inferior Turbinate Salimov, Ozer 189

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A title page, including the manuscript title and all authors’ full names, academic degrees (no more than three), institutional affi liations, and locations. Designate one author as the corresponding author. Also indicate where the paper was presented, if applicable. A structured abstract of up to 250 words with the headings: Introduction, Objective, Methods, Results, and Conclusion. The Manuscript body should be divided as: introduction with objective(s); method; result; discussion; conclusion; references.

Manuscript length of no more than 24 pages (exclusive of the title page and abstract). Studies involving human beings and animals should include the approval protocol number of the respective Ethics Committee on Research of the institution from which the research is affi liated.

Systematic Reviews (including Meta-analyses)Critical assessments of literature and data sources on important clinical topics in otolaryngology-head and neck surgery. Systematic reviews that reduce bias with explicit procedures to select, appraise, and analyze studies are highly preferred over traditional narrative reviews. The review may include a meta-analysis, or statistical synthesis of data from separate, but similar, studies leading to a quantitative summary of the pooled results. The components of a systematic review are:

A title page, including the manuscript title and all authors’ full names, academic degrees, institutional affi liations, and locations. Designate one author as the corresponding author. Also indicate where the paper was presented, if applicable. A structured abstract of up to 250 words with the head-ings: Introduction, Objectives, Data Synthesis, and Con-clusion. The Manuscript body should be divided as: introduction; review of literature; discussion; fi nal comments; refer-ences. Manuscript length of no more than 24 pages (exclusive of the title page and abstract).

Case ReportsCase Reports will no longer be accepted for submission, starting on 2015. Submitted manuscripts until December 2014 will be reviewed and published, if approved.

Update ManuscriptsThe manuscript is an update that explores a particular subject, developed from current data, based on recently published works.

A title page, including the manuscript title and all authors’ full names, academic degrees, institutional affi liations, and locations. Designate one author as the corresponding author. Also indicate where the paper was presented, if applicable. A structured abstract of up to 250 words with the headings: Introduction, Objectives, Data Synthesis, and Conclusion. The Manuscript body should be divided as: introduction; review of a particular subject; discussion; fi nal comments; references. Manuscript length of no more than 15 pages (exclusive of the title page and abstract).

Letters to the Editor and Opinion articlesOnly by invitation from the Editorial Board. Manuscript length: no more 2 pages.

Instructions to AuthorsA-2

Manuscript Preparation

Correct preparation of the manuscript will expedite the review and publishing process. Manuscripts must conform to acceptable English usage.

Necessary Files for Submission (each topic should start in a new page):

Title Page Abstract Manuscript (main text, references, and fi gure legends) Figure(s) (when appropriate) Table(s) (when appropriate)

In accordance with double-blind review, author/institutional information should be omitted or blinded from the following submission fi les: Manuscript, Figure(s), Table(s), Response to Reviewers.

The Abstract should be followed by three to six keywords in English, selected from the list of Descriptors (Mesh) created by National Library of Medicine and available at http://www.nlm.nih.gov/mesh/2013/mesh_browser/MBrowser.html.

AbbreviationsDo not use abbreviations in the title or abstract. When using ab-breviations in the text, indicate the abbreviation parenthetically after the fi rst occurrence and use the abbreviation alone for all subsequent occurrences.

AuthorshipAuthorship credit should be based on criteria established by the International Committee of Medical Journal Editors: (1) substantial contributions to conception and design, acquisition of data, or analysis and interpretation of data; (2) drafting the article or revising it critically for important intellectual content; and (3) fi nal approval of the version to be published.

ReferencesAuthors are responsible for the completeness, accuracy, and format of their references. References should be numbered consecutively using Arabic numbers in the text. All authors shall be listed in full up to the total number of six; for seven or more authors, list the fi rst three authors followed by “et al.”There should be no more than 90 references for original articles and no more than 120 for systematic reviews or update articles. Refer to the List of Journals Indexed in Index Medicus for abbreviations of journal names, or access the list at http://www.nlm.nih.gov/tsd/serials/lji.html. Sample references are given below. For more information, please check: http://www.ncbi.nlm.nih.gov/books/NBK7256/.

Examples Journals: Author | Article Title | Journal Title | Date of Publication | Volume Number | Issue Number | Pagination.

Huttenhower C, Gevers D, Knight R, et al. Structure, function and diversity of the healthy human microbiome. Nature 2012;486(7402):207-214 Dissertations and Theses: Author | Title | Content Type | Place of Publication | Publisher | Date of Publication | Pagination.

Baldwin KB. An exploratory method of data retrieval from the electronic medical record for the evaluation of quality in healthcare [dissertation]. Chicago: University of Illinois at Chicago, Health Sciences Center; 2004:116

Books: Author/Editor | Title | Edition | Place of Publication | Publisher | Date of Publication.

Valente M, Hosford-Dunn H, Roeser RJ. Audiology Treatment. 2nd ed. New York: Thieme; 2008 Book chapters: Author of the chapter | Title of chapter | In: Editor(s) of book | Title of chapter | Place of Publication | Publisher | Date of Publication | Pagination.

Vilkman E. A survey on the occupational safety and health arrangements for voice and speech professionals in Europe. In: Dejonckere PH, ed. Occupational Voice: Care and Cure. Hague: Kugler Publications; 2001:129-137 Electronic material: for articles taken entirely from the Internet, please follow the rules mentioned above and add at the end the web site address.

Ex: AMA: helping doctors help patients [Internet]. Chicago: American Medical Association; c1995-2007 Available at: http://www.ama-assn.org/. Accessed Feb 22, 2007

FiguresFigures must be uploaded separately. Include the number of the fi gure in the description box.

Figure LegendsProvide a legend for each fi gure. List the legends (double-spaced) on a separate text page, after the reference page. Up to 8 pictures will be published at no cost to the authors; color pictures will be published at the editor’s discretion. Acceptable submissions include the following: JPG, GIF, PNG, PSD, or TIF. The Publication Management System accepts only high defi nition images with the following features:

Width up to 1000 px and DPI equal to or higher than 300; The image formats should be preferentially TIF or JPG; The maximum image size should be 8 MB; If fi gures have multiple parts (e.g., A, B, C, D), each part must be counted as a separate image in the total number allowed.

Tables and GraphsTables should be numbered in Arabic numbers consecutively as they appear in the text, with a concise but self-explicative title, without underlined elements or lines inside it. When tables have too many data, prefer to present graphics (in black and white). If there are abbreviations, an explicative text should be provided on the lower margin of the table or graph.

AppendicesAppendices will only be published online, not in the print journal, and may include additional fi gures or tables that enhance the value of the manuscript. Appendices must be submitted online with the rest of the manuscript and labeled as such. Questionnaires will be considered as Appendices.

Online Manuscript Submission

All manuscripts should be submitted at http://mc.manuscriptcentral.com/iaorl, which gives access to the ScholarOne Manuscripts submission system where the submission of the article is done by the authors and the evaluation process is done by the reviewers of our editorial board in a blinded process where the names of the authors are not displayed in any instance. The system will ask for your user ID and password if you have already registered. If you have not registered, click on the link “Create Account” and make your registration. In case you have forgotten your password, click on the appropriate link and the system will generate an automatic e-mail with the information.

Instructions to Authors A-3

The author(s) should keep a copy of all submitted material for publication because the editor cannot be held responsible for any lost material.

After submission, the system off ers the option of saving a copy of your manuscript in PDF format for your control.

The journal strongly recommends that the authors submit their electronic manuscripts written in Microsoft Word. In the “Preparing Manuscript” step a screen that simulates the word processor will be displayed, where it is possible to “copy and paste”, including tables.

Mandatory Author Forms

Ethics and Financial Disclosure: The manuscript will be assigned to an Editor for solicitation of peer review and editorial evaluation ONLY after this form has been submitted by the corresponding author.

Patient Confi dentiality

For manuscripts containing photographs of a person, submit a written release from the person or guardian, or submit a photograph that will not reveal the person’s identity (eye covers are inadequate to protect patient identity).

Using Previously Published Material and IllustrationsFor manuscripts containing illustrations and/or material reproduced from another source, permission from the copyright holder, medical illustrator, or original publication source must be obtained and submitted to the editorial offi ce.

IRB Policy and Animal Studies

For all manuscripts reporting data from studies involving human participants, formal review and approval, or formal review and waiver (exemption), by an appropriate institutional review board (IRB) or ethics committee is required and should be described in the Methods section with the full name of the reviewing entity. All clinical research requires formal review, including case reports, case series, medical record reviews, and other observational studies. For experiments involving animals, state the animal-handling protocol in the Methods section, including approval by an institutional board.

Duplicate or Redundant Submission

Manuscripts are considered with the understanding that they have not been published previously and are not under consideration by another publication. If the author explicitly wishes the journal to consider duplicate publication, he or she must submit the request, in writing, to the Editor with appropriate justifi cation.

Deadlines

Submissions not in compliance with the following instructions will be returned to the author by the editorial offi ce and a corrected version must be resubmitted within 30 days. Papers not resubmitted within that time will be withdrawn from consideration.

Revised manuscripts must follow the same instructions and should be submitted within 30 days of the revision letter date.

Accepted manuscripts sent to the publisher will be typeset and proofs will then be sent by e-mail to the corresponding author. If proofs are not approved and received within 2 business days, the article will not be published.

The reviewers should send their comments within 20 days.

English Language Assistance

Appropriate use of the English language is a requirement for publication in IAORL. Authors who wish to improve the grammar and spelling in their articles may wish to consult a professional service. Many companies provide substantive editing via the web. A few examples are:

www.journalexperts.comwww.editage.com

Please note that IAORL has no affi liation with these companies and use of the service does not guarantee your manuscript will be accepted.

The International Archives of Otorhinolaryngology Scientifi c Merit Journal Prize

The IAORL Scientifi c Merit Journal Prize is awarded every year for up to three best systematic review (meta-analysis) papers published each year in the journal. The 2015 manuscript awards will be selected from articles published in issues 1-4 of volume 19, based on novelty, impact, data quality, and number of online downloads by the journal readers.

The adjudication committee consists of the editorial board, assisted by comments received through the peer review process. The judgment of the papers will be published after issue number 4 of volume 19. The result will be communicated to the winners and offi cially published in volume 20 of IAORL.

All authors and co-authors will receive certifi cates of award and the fi rst author of each of the three selected manuscripts will receive $1,000 (USD).

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135 Mercury Exposure in a Riverside Amazon Population, Brazil: A Study of the Ototoxicity of MethylmercuryAna Hoshino, Heloisa Pacheco-Ferreira, Seisse Gabriela G. Sanches, Renata Carvallo, Nathália Cardoso, Maurício Perez, and Volney de Magalhães Câmara

141 Lipidomic Profi ling of Mastoid Bone and Tissue from Patients with Chronic OtomastoiditisFarbod Fazlollahi, Kessiri Kongmanas, Nongnuj Tanphaichitr, Jeff rey Suh, Kym Faull, and Quinton Gopen

151 Auditory Neuropathy/Dyssynchrony: A Retrospective Analysis of 15 CasesMurat Unal, and Yusuf Vayisoglu

156 Characterization of Hearing Thresholds from 500 to 16,000 Hz in Dentists: A Comparative StudyClaudia Giglio de Oliveira Gonçalves, Luciana Santos, Diolen Lobato, Angela Ribas, Adriana Bender Moreira Lacerda, and Jair Marques

161 Auditory Brainstem Response in Term and Preterm Infants with Neonatal Complications: The Importance of the Sequential EvaluationDaniela da Silva, Priscila Lopez, and Jair Cortez Mantovani

Systematic Reviews166 Health Promotion in Obstructive Sleep Apnea Syndrome

Camila de Castro Corrêa, Wanderléia Quinhoneiro Blasca, and Giédre Berretin-Felix

171 The Study of Otoacoustic Emissions and the Suppression of Otoacoustic Emissions in Subjects with Tinnitus and Normal Hearing: An Insight to Tinnitus EtiologyLucieny Serra, Gabriela Novanta, Andre Lopes Sampaio, Carlos Augusto Oliveira, Ronaldo Granjeiro, and Silvia Cristina Braga

Update Article176 Olfaction in Neurologic and Neurodegenerative Diseases: A Literature Review

Maria Dantas Costa Lima Godoy, Richard Louis Voegels, Fábio de Rezende Pinna, Rui Imamura, and José Marcelo Farfel

Case Reports180 Ortner’s Syndrome: Secondary Laryngeal Paralysis Caused by a Great Thoracic Aorta Aneurysm

Ana Claudia Alves Zangirolami, Frederico Vieira de Oliveira, and Miguel Soares Tepedino

183 Intralabyrinthine Penetrating Ventilation Tube with Preservation of Hearing: An Unusual Clinical SituationTantely Razafi mahefa Raoelina, Maya Elziere, Justin Michel, and Arnaud Devèze

187 A Rare Location of Angiofi broma in the Inferior Turbinate in Young WomanAsif Salimov, and Serdar Ozer

A-1 Instructions to Authors

Continuation from outside back cover

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Rio de Janeiro • New York • StuttgartThieme Publicações Ltda, Av. Nilo Peçanha, 50, sala 2508, Rio de Janeiro 20020-906, Brazilwww.thieme-connect.com/products

Editorial99 Research Awards 2015

Geraldo Pereira Jotz

Original Research100 Comparing Voice Self-Assessment with Auditory Perceptual Analysis in Patients with Multiple Sclerosis

Vladimir Bauer, Zorica Aleric, and Ervin Jancic

106 Teleducation about Cleft Lip and Palate: An Interdisciplinary Approach in the Promotion of HealthCamila de Castro Corrêa, Thais Freire, Júlia Speranza Zabeu, Aline Martins, Rafael Ferreira, Paulo Afonso Silveira Francisconi, Jeniff er de Cássia Rillo Dutka, and Wanderléia Quinhoeiro Blasca

112 Parotid Incidentaloma Identifi ed by Positron Emission/Computed Tomography: When to Consider Diagnoses Other than Warthin TumorCarolina Bothe, Alejandro Fernandez, Jacinto Garcia, Montserrat Lopez, Xavier León, Miquel Quer, and Joan Lop

116 Preoperative Imaging Modalities to Predict the Risk of Regional Nodal Recurrence in Well-Diff erentiated Thyroid CancersMohammed K. AlNoury, Saad M. Almuhayawi, Khalid B. Alghamdi, and Khaled I. Al-Noury

121 Foreign Bodies in the Ear, Nose and Throat: An Experience in a Tertiary Care Hospital in Central NepalRamesh Parajuli

124 Diff erential Diagnosis and Treatment of Isolated Pathologies of the Sphenoid Sinus: Retrospective Study of 46 CasesThomas Ribeiro Marcolini, Maryane Cristine Safraider, Jan Alessandro Socher, Guilherme Olinto Lucena

130 Surfactant Protein A Expression in Chronic Rhinosinusitis and Atrophic RhinitisMohammad Waheed El-Anwar, Atef A. Hamed, Abd ElRaof Said Mohamed, Ahmad Abdel-Fattah Nofal, Maha A. Mohamed, and Hesham R. Abdel-Aziz

Offi cial Publication of the Otorhinolaryngology Foundation and Societas Oto-Rhino-Laryngologia Latina

OTORHINOLARYNGOLOGYINTERNATIONAL ARCHIVES OF

ISSN 1809-9777Issue 2 Volume 19 April - May - June 2015

International Archives of Otorhinolaryngology

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Transcript of International Archives of Otorhinolaryngology