Research Article Assessment of Tachykinin Receptor 3 Gene...

Research ArticleAssessment of Tachykinin Receptor 31015840 Gene Polymorphismrs3733631 in Rosacea

Anthony Karpouzis1 Paschalis Avgeridis2 Gregory Tripsianis3 Elisavet Gatzidou2

Niki Kourmouli2 and Stavroula Veletza2

1Department of Dermatology Faculty of Medicine Democritus University of Thrace 68100 Alexandroupolis Greece2Department of Medical Biology Faculty of Medicine Democritus University of Thrace 68100 Alexandroupolis Greece3Department of Medical Statistics Faculty of Medicine Democritus University of Thrace 68100 Alexandroupolis Greece

Correspondence should be addressed to Stavroula Veletza sveletzamedduthgr

Received 28 May 2015 Revised 31 August 2015 Accepted 3 September 2015

Academic Editor Monika Dmitrzak-Weglarz

Copyright copy 2015 Anthony Karpouzis et al This is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work is properlycited

Background Rosacea is a chronic skin disease possibly following the neurogenic skin inflammationmodel Neurokinin B involvedin the pathogenesis of Parkinsonrsquos disease frequently coexisting with subsequent onset of rosacea is an endogenous ligand of thetachykinin receptor 3 (TACR3)Methods 128 rosacea patients and 121matched controls were genotyped for rs3733631 by PCR-RFLPand analyzed by chi-square testResultsWeobserved statistically significant predominance of the CG orGG genotype (119901 = 0006)and of the G allele (119901 = 0004) in the papulopustular (PP) form of rosacea and statistically marginal significance of the CG or GGgenotype in erythematotelangiectatic (ET) rosacea (119901 = 0052) Significantly higher frequency of the CG or GG genotype and Gallele in PP rosacea (119901 = 0021 and 119901 = 0008 resp) was ascertained within male patients Conclusion TACR3 rs3733631 G allelepossibly predisposes the evolution of the initial phase of rosacea to the PP and not the ET form in male patients

1 Introduction

Rosacea is a persistent and recurrent inflammatory conditionof themiddle face as a rule Classical morphology rosacea fre-quent subtypes are the erythematotelangiectatic (ET) rosaceaand the papulopustular (PP) rosacea [1] while the otheranatomoclinical forms are the granulomatous (or lupoid)rosacea the phymatous rosacea or rhinophyma [2] androsacea fulminans or pyoderma faciale [1] In general a per-sistent stasis erythemamay precede by years the developmentand establishment of either an ET form or a PP form Rosaceamay appear in any immunocompetent patient and has to bedifferentiated from demodicosis of either immunocompetentor immunodeficient patients [2]

Semeiology symptomatology and pathologic features ofrosacea suggest that its pathogenesis is mediated by sensorynerves via antidromically released neuromediators thereforeconstituting a model of neurogenic inflammation [3]

After emotional stimulation or ingestion of alcohol cate-cholamine is released fromgastricmucosa or adrenalmedullaand kallikrein is secreted by salivary glands increasingbradykinin levels in blood Bradykinin is associated withcentrofacial vasodilatation [4] Substance P (SP) is a neuro-peptide secreted by sensory neurons of human epider-mis and dermal papillae SP biologic functions consist inlocal axon reflex vasodilatation acceleration of epidermaland endothelial cell proliferation mast cell degranulationantidromic vasodilatation and liberation of inflammationmediators from macrophages and T lymphocytes Biopsieson papular lesion and healthy skin of volunteer rosaceapatients have shown that in genetically predisposed personsSP vasodilatory activity may be involved in the inductionandor maintenance of PP rosacea [5]

Vessel dilatation in rosacea is not due to yielding todamaged connective tissue but to fusion of vessels by agradual breakdown of adjacent vascular walls [6] Vascular

Hindawi Publishing CorporationInternational Scholarly Research NoticesVolume 2015 Article ID 469402 6 pageshttpdxdoiorg1011552015469402

2 International Scholarly Research Notices

endothelial growth factor (VEGF) staining on skin sectionshas detected no differences between PP rosacea and ETrosacea [7] Laser Doppler Imager has detected elevatedblood flow in affected skin compared with nonaffected skinof the same patient in the PP form but not in the ET form ofrosacea [8]

Normal skin Demodex folliculorum (a head sebaceousfollicle lumen mite) usually located at the upper part of theinfundibulum of the follicle [9] has been indirectly involvedin rosacea although a possible causal correlation has notbeen established up to this day The majority of rosaceamedications do not affect Demodex but do improve rosaceawhile treatment with Lindane which is effective on the mitedoes not improve the condition [1] PossiblyDemodexmay bean exclusively exacerbating factor in rosacea patients Densityof Demodex was demonstrated to be significantly higher inthe PP form of rosacea although no correlation was ascer-tained between Demodex intrafollicular inflammation andperifollicular inflammation Possibly increased blood flow inrosacea skin dilated vessels constitutes an ideal conditionfor dermis Demodex multiplication Demodex folliculorumantigens reacting with rosacea patientsrsquo sera have beenconsidered to be responsible for stimulating mononuclearcell proliferation [10] High vascular density was significantlymore frequent in the PP form skin sections The numberof mast cells (which promote vasodilatation angiogenesisand fibrosis) was significantly higher in lesional than innonlesional skin of patients with rosacea [10]

All things considered it is evident that neuropeptidessuch as SP andor bradykinin may play a decisive role inincreasing blood flow andor vascular density in rosaceaConsequently it looked reasonable to explore the involve-ment of single nucleotide polymorphisms (SNPs) withinneuromediators and their receptors in rosacea

ldquoKininrdquo is considered as hypotensive polypeptide con-tracting most isolated smooth muscles Tachykinins (TACs)are characterized by pharmacological similarity to theirendogenous inflammatory mediator bradykinin yet TACsare not of similar structure to that of bradykinin TACsare not kinins but the name remains [11] Heparin-initiatedbradykinin formation plays an important role in mast cell-mediated diseases [12] such as rosacea

TACs are a family of closely related peptides such as SPneurokinin A and neurokinin B which act as neuropep-tides in nonneuronal cells and noninnervated tissues [13]Biological actions of TAC1 and TAC3 are mediated by threetypes of G protein-coupled receptors (TACR1 TACR2 andTACR3) TACR3 or NK-R3 or neurokinin B receptor belongsto a family of genes which function as receptors for TACscharacterized by interactionwithG proteins variations in the5-end of the sequence and 7-hydrophobic transmembraneregions interactions TAC1 gene is localized on chromosome7q21-q22 TAC3 is localized on 12q13-q21 TACR1 is localizedon 2p12 TACR2 is localized on 10q11-q21 and TACR3 islocalized on 4q25 [14] Circulating neurokinin B is elevatedin women with preeclampsia during the third trimester ofpregnancy [15] The increased placental expression of TAC3gene belongs to the mechanism responsible for the elevatedcirculating neurokinin B levels in preeclampsia [16] In

preeclampsia complicated pregnancies elevated neurokininB plasma levels have been associated with elevated nitricoxide metabolite levels possibly in a compensatory mecha-nism to improve blood flow to the uteroplacental unit [15] Infamilial hypogonadotropic hypogonadismmutations of neu-rokinin B gene and its respective receptor TACR3 suggestedan eventual key role for neurokinin B in the central control ofreproduction [17]

Rosacea is significantly more frequent in patients suffer-ing from preexisting Parkinsonrsquos disease [18] Neurokinin B(endogenous ligand of TACR3) is involved in pathogenesis ofParkinsonrsquos disease by interacting with brain dopaminergictransmission [19]

Kisspeptinneurokinin Bdynorphin-expressing neuronsof hypothalamus promoting skin vasodilatation partici-pate in the estrogen modulation of the body temperatureand contribute to the induction of ensuing hot flushes inmenopausal women such flushes are clinically similar tothose experienced in rosacea [20] The role of mast cells intachykinin neurogenic skin inflammation induction has beenshown and thus their intense presence in PP rosacea [21 22]may be explained

We chose to study the TACR3 gene polymorphismrs3733631 because TACR3 endogenous ligand neurokinin Bby increasing blood flow (anatomoclinical feature of rosacea)is involved in the pathophysiology of (a) Parkinsonism(frequently coexists with rosacea) and (b) hot flushes inmenopausal women (clinically similar to rosacea vasomotorcrises)

Possible involvement of the TACR3 gene in rosaceawould most probably be associated with either upregulationor downregulation of this gene SNPs such as rs3733631localizedwithin the promoter area upstream from the codingregion make prime candidates to reflect any associationbetween gene and disease Certain additional TACR3 poly-morphisms such as A29V G59E S455G A449S W275Xand rs4580655 have been associated with idiopathic centralpubertal disorders [23 24] predicting and improving learn-ing and memory in aged organism [25] alcohol and cocainedependence [26] and pediatric slow transit constipation [27]

2 Materials and Methods

21 Patientsrsquo Characteristics One hundred and twenty-eightpatients examined at the Outpatient Clinic of the UniversityDepartment of Dermatology at the University Hospital ofAlexandroupolis 53 males (414) and 75 females (586)were included in this study Patientsrsquo age ranged from 14to 86 years with a median age of 5625 plusmn 1635 years Allpatients were clinically diagnosed by the same physician andsuffered from either the ET form (119899 = 67) or the PP form(119899 = 61) of central face rosacea [1] Rosacea patients withaccompanying disease were excluded from the study toavoid possible interactions with potentially coexisting genet-ically determined disorders One hundred and twenty-oneunrelated subjects free from rosacea not suffering fromany potentially genetically determined disorder individuallymatched to patients by both gender and age were recruitedas controls [54 (446) males and 67 (554) females] There

International Scholarly Research Notices 3

Table 1 Characteristics of patients with rosacea and healthy con-trols

Patients Controls 119901 valueNumber 128 121Age (years mean plusmn SD) 5625 plusmn 1635 5681 plusmn 1596 0783Gender 0608

Male 53 (414) 54 (446)Female 75 (586) 67 (554)

Type of diseasePP 61 (477) mdashET 67 (523) mdash

were no significant differences in gender (119901 = 0608) and age(119901 = 0783) between patients and controls (Table 1) Workprocedures conform with the Helsinki Declaration Princi-ples

22 Methods Genomic DNA from whole blood was isolatedby salting out as previously described [28] We genotyped theTACR3 gene polymorphism rs3733631 CG by PCR-RFLP asfollows primers F1015840 51015840-CTTGCAGCGAATGAATGAAA-31015840and R1015840 51015840-GGGTAATCGAGTCACATCAGG-31015840 generated a216p long amplicon after 40 cycles of PCR at 95∘ 3010158401015840 55∘3010158401015840 and 72∘ 3010158401015840ThePCRproductwas subsequently digestedwith restriction enzyme HaeIII which generates three bands107 86 and 21 bp for allele G and two bands 128 and 86 bp forallele C Restriction digests were analyzed by electrophoresisin 25 agarose gel

Statistical analysis of the data was performed usingthe statistical package for the Social Sciences (SPSS) (INCChicago IL USA)The chi-square test was used to assess dif-ferences in genotype and allele frequencies between patientssuffering from rosacea and matched controls It was alsoused to compare the observed frequency of each genotypewith that expected for a population in the Hardy-Weinbergequilibrium Multivariate unconditional logistic regressionanalysis was used to estimate age-adjusted odd rations (OR)and 95 confidence intervals (CI) as the measure of associ-ation of the studied polymorphism with the appearance ofrosacea All tests were two-tailed and statistical significancewas considered for values less than 005

3 Results

TACR3 gene polymorphism rs3733631 CG is located atthe promoter of TACR3 The distribution of genotypes andalleles in patients and healthy controls is shown in Table 2CC CG and GG genotypes were found in 844 133and 23 of patients and in 876 116 and 08 ofhealthy controls respectively The genotype distribution wasin Hardy-Weinberg equilibrium in controls (1205942 = 0481 df =1 and 119901 = 0488) but not in patients (1205942 = 4512 df = 1and 119901 = 0034) No statistically significant differences weredetected in either genotype or allele frequencies between thetwo groups (119901 = 0433 and119901 = 0295 resp)However a trendof higher frequency of G carriers (GG or CG genotype) andG allele was substantiated within males (227 versus 130

Table 2 Distribution of genotypes among patients with rosacea andhealthy controls

Controlslowast Patientslowast ORdagger 95 CI 119901 valueTotalGenotypeCC 106 (877) 108 (844) RefCG 14 (116) 17 (133) 121 057ndash258 0621GG 1 (08) 3 (23) 316 032ndash3107 0325

Recessive modelCC 106 (877) 108 (844) RefCG or GG 15 (124) 20 (156) 134 065ndash276 0433

Allele contrastC 226 (934) 233 (910) RefG 16 (66) 23 (90) 143 073ndash278 0295

MalesGenotypeCC 47 (870) 41 (773) RefCG 7 (130) 9 (170) 148 050ndash434 0478GG 0 (00) 3 (57) mdash mdash mdash



FemalesGenotypeCC 59 (881) 67 (893) RefCG 7 (104) 8 (107) 102 035ndash299 0973GG 1 (15) 0 (00) mdash mdash mdash



Note statistical significance for differences in genotype G-containinggenotype and allelic frequencies between patients with rosacea and healthycontrols (i) 1205942 = 1113 df = 2 and 119901 = 0573 1205942 = 0537 df = 1 and 119901 = 04641205942 = 0970 df = 1 and 119901 = 0325 among the entire cohort (ii) 1205942 = 3650

df = 1 and 119901 = 0161 1205942 = 1716 df = 1 and 119901 = 0190 1205942 = 3412 df = 1 and119901 = 0065 among males (iii) 1205942 = 1127 df = 1 and 119901 = 0569 1205942 = 0057 df =1 and 119901 = 0811 1205942 = 0241 df = 1 and 119901 = 0624 among females lowastData arenumber of subjects and percentage () daggeradjusted for age and gender

119901 = 0139 for GG GC genotypes and 142 versus 65119901 = 0071 for theG allele) (Table 2)More specifically inmalepatients an increased (but not statistically significant) riskof rosacea incidence was associated with G allele (OR 23895 CI 093ndash611) Further stratification for the clinical formof rosacea in each patient suggested a statistically significantpredominance ofG carriers suffering fromPP formof rosaceaversus controls (705 CC 262 CG and 33 GG inpatients versus 877 CC 116 CG and 08 GG incontrols) as well as a higher frequency of the allele G (836C and 164 G in patients versus 934 C and 66 G in


Table 3 (a) Distribution of genotypes among patients with ETrosacea and healthy controls (b) Distribution of genotypes amongpatients with PP rosacea and healthy controls

(a)

Controlslowast ETlowast ORdagger 95 CI 119901 value

Total

Genotype

CC 106 (877) 65 (970) Ref

CG 14 (116) 1 (15) 012 002ndash093 0043

GG 1 (08) 1 (15) 172 011ndash2825 0704

Recessive model

CC 106 (877) 65 (970) Ref

CG or GG 15 (124) 2 (30) 022 005ndash101 0052

Allele contrast

C 226 (934) 131 (978) Ref

G 16 (66) 3 (22) 033 010ndash117 0087

Males

Genotype

CC 47 (870) 23 (920) Ref

CG 7 (130) 1 (40) 031 004ndash268 0285

GG 0 (00) 1 (40) mdash mdash mdash

Recessive model

CC 47 (870) 23 (920) Ref

CG or GG 7 (130) 2 (80) 062 012ndash326 057

Allele contrast

C 101 (935) 47 (940) Ref

G 7 (65) 3 (60) 097 024ndash395 0966

Females

Genotype

CC 59 (881) 42 (1000) mdash

CG 7 (104) 0 (00) mdash mdash mdash


Recessive model

CC 59 (881) 42 (1000) mdash

CG or GG 8 (119) 0 (00) mdash mdash mdash

Allele contrast

C 125 (933) 84 (1000) mdash

G 9 (67) 0 (00) mdash mdash mdashNote statistical significance for differences in genotype G-containinggenotype and allelic frequencies between patients with ET rosacea andhealthy controls (i) 1205942 = 6089 df = 2 and 119901 = 0048 1205942 = 4644 df = 1 and119901 = 0031 1205942 = 3437 df = 1 and 119901 = 0064 among the entire cohort (ii) 1205942 =3563 df = 2 and 119901 = 0168 1205942 = 0417 df = 1 and 119901 = 0518 1205942 = 0013 df = 1and 119901 = 0908 amongmales (iii) 1205942 = 5412 df = 2 and 119901 = 0067 1205942 = 5412df = 1 and 119901 = 0020 1205942 = 5885 df = 1 and 119901 = 0014 among females lowastDataare number of subjects and percentage () daggeradjusted for age and gender

(b)

Controlslowast PPlowast ORdagger 95 CI 119901 valueTotalGenotypeCC 106 (877) 43 (705) RefCG 14 (116) 16 (262) 281 126ndash627 0011GG 1 (08) 2 (33) 5 044ndash5698 0195









Note statistical significance for differences in genotype G-containinggenotype and allelic frequencies between patients with PP rosacea andhealthy controls (i) 1205942 = 8217 df = 2 and 119901 = 0016 1205942 = 7999 df = 1 and119901 = 0005 1205942 = 8709 df = 1 and 119901 = 0003 among the entire cohort (ii) 1205942 =7517 df = 2 and119901= 00231205942 = 5808 df = 1 and119901= 00161205942 = 8044 df = 1and119901= 0005 amongmales (iii)1205942 = 3696 df = 2 and119901= 01581205942 = 2490df = 1 and 119901 = 0115 1205942 = 1661 df = 1 and 119901 = 0197 among females lowastDataare number of subjects and percentage () daggeradjusted for age and gender

controls) (Tables 3(a) and 3(b)) Therefore the frequency ofCG or GG genotype on one hand is statistically significantlyhigher in PP rosacea patients (119901 = 0006) and G allele isalso more frequent in PP patients (119901 = 0004 Table 3(b))Consequently a statistically significant increased risk of PProsacea incidence was associated with G allele (OR 277 95CI 138ndash557) In patients with ET rosacea the predominance


of G allele was not significant (119901 = 0087) but the higherfrequency of CG or GG genotype in patients was closerto being statistically significant (119901 = 0052) (Table 3(a))Regarding the distribution of genotypes in PP rosacea malesthe higher frequency of CG or GG genotype in patients wasstatistically significant (119901 = 0021) and the frequency of Gallele was significantly higher (119901 = 0008 Tables 3(a) and3(b)) No statistical significance was observed within females

4 Discussion

Genetic polymorphisms have not been previously studiedextensively in rosacea Specifically in one study of Bsm1 poly-morphism of the vitaminD receptor (VDR) the frequency ofallele 1 was increased in patients with rosacea fulminans butthat increase was not statistically significant No differenceswere detected in PP or ET rosacea [29] Another studyshowed a statistically significant relationship between bothnull combination of glutathione-S-transferase M1 (GCTM1)and GSTT1 genotype polymorphisms and rosacea in thesetting of a comparative genetic study in a group of rosaceapatients and a group of matched healthy controls [30]

A recently published genome-wide association studyreported correlation of rosacea with HLA-DRB1lowast HLA-DQB1lowast and HLA-DQA1lowast as well as rs763035 providingevidence regarding contribution of genetic component topathogenesis of rosacea [31]

In our study we observed a trend (119901 = 0071) withinmalepatients to carry G allele of rs3733631 with higher frequencythan controls regardless of the type of rosacea No such trendwas observed within females

There have been no previous studies in the literaturereporting association or not of rs3733631 TACR3 gene poly-morphism with rosacea however it has been studied inalcohol and cocaine dependence [26]

The Toll-like receptor 2 gene (TLR2) is located at 4q32[32] adjacent to TACR3 at 4q25 In PP rosacea serineprotease kallikrein-5 (KLK5) and two abnormal cathelicidinpeptides (LL-37 and FA-29) are upregulated in the skin andinduce erythema and vascular dilatation when injected inmouse skin LL-37 is characterized by lower antimicrobialactivity than normal smaller cathelicidin in healthy skinand it promotes angiogenesis and consequently secondaryinflammation [33]

It is possible that the polymorphism at the adjacentTACR3 may be somehow involved in increasing expressionof TLR2 in rosacea and therefore in upregulation of KLK5 ina calcium-dependent manner possibly PP rosacea patientsoverreact resulting in the histogenesis of rosacea papules andpustules even though bacterial diversity and quantities aresimilar in rosacea lesions and in normal skin [34]

TLR2 stimulation occurs only as a response to specifictriggers such as emotional stimulation or ingestion of alco-hol and kallikrein secreted by salivary glands increasingvasodilatory bradykinin blood levels [5 33] The vitamin D-dependent amplification mechanism might induce increaseof TLR2 susceptibility as 125 (OH)

2vitamin D gene poly-

morphism has been correlated with the intensity of inflam-mation [29]

Polymorphism R702W in NOD2CARD15 is specifi-cally associated with childhood granulomatous rosacea [35]This gene participates in the N-terminal caspase recruit-ment (NACT) protein domain family being involved inthe response of Toll-like receptors transduced inflammatorystimuli [36]

Our study suggested a statistically significant predomi-nance of CG or GG genotype (and G allele) in patientssuffering from PP rosacea versus matched healthy controlsThe number of patients included in our study constitutes arelatively small size sample additional studies are needed toconfirm our reported results PP rosacea is characterized byelevated blood flow and increased SP vasodilatory activityin affected skin Genotypes carrying G allele may be accom-panied by alterations of kisspeptinneurokinin Bdynorphin(KNDy) neurons of the hypothalamus and these alteredneurons (via projections to rostral hypothalamic structuressuch as the medial preoptic area and medial preoptic nucleuswhich control thermoregulatory effectors) could maximizethe vasodilatation of the skin as well as the inflammation(induced by vasodilatation) resulting in development of PProsacea [6 20 33]

5 Conclusion

Genotypes carrying G allele may provide a genetic predis-position to PP rosacea and not to the ET form ThereforePP rosacea may be characterized by a genetic uniquenessposing the possibility of its nosological distraction from theET form

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

References

[1] G F Webster ldquoRosaceardquoMedical Clinics of North America vol93 no 6 pp 1183ndash1194 2009

[2] B Cribier ldquoRosacea under the microscope characteristic histo-logical findingsrdquo Journal of the European Academy of Dermatol-ogy and Venereology vol 27 no 11 pp 1336ndash1343 2013

[3] A A Aubdool and S D Brain ldquoNeurovascular aspects of skinneurogenic inflammationrdquo Journal of Investigative DermatologySymposium Proceedings vol 15 no 1 pp 33ndash39 2011

[4] M Guarrera A Parodi C Cipriani C Divano and A ReboraldquoFlushing in rosacea a possible mechanismrdquo Archives of Der-matological Research vol 272 no 3-4 pp 311ndash316 1982

[5] N Kurkcuoglu and F Alaybeyi ldquoSubstance P immunoreactivityin rosaceardquo Journal of the American Academy of Dermatologyvol 25 no 4 pp 725ndash726 1991

[6] E Neumann and A Frithz ldquoCapillaropathy and capillaroneo-genesis in the pathogenesis of rosaceardquo International Journal ofDermatology vol 37 no 4 pp 263ndash266 1998

[7] A H A GomaaM YaarMM K Eyada and J Bhawan ldquoLym-phangiogenesis and angiogenesis in non-phymatous rosaceardquoJournal of Cutaneous Pathology vol 34 no 10 pp 748ndash7532007


[8] D A Guzman-Sanchez Y Ishiuji T Patel J Fountain Y HChan and G Yosipovitch ldquoEnhanced skin blood flow andsensitivity to noxious heat stimuli in papulopustular rosaceardquoJournal of the American Academy of Dermatology vol 57 no 5pp 800ndash805 2007

[9] E C Sattler T Maier V S Hoffmann J Hegyi T Ruzicka andC Berking ldquoNoninvasive in vivo detection andquantification ofDemodexmites by confocal laser scanning microscopyrdquo BritishJournal of Dermatology vol 167 no 5 pp 1042ndash1047 2012

[10] K Aroni E Tsagroni N Kavantzas E Patsouris and E Ioanni-dis ldquoA study of the pathogenesis of Rosacea how angiogenesisand mast cells may participate in a complex multifactorialprocessrdquoArchives of Dermatological Research vol 300 no 3 pp125ndash131 2008

[11] A M Khawaja and D F Rogers ldquoTachykinins receptor toeffectorrdquo International Journal of Biochemistry and Cell Biologyvol 28 no 7 pp 721ndash738 1996

[12] C Oschatz C Maas B Lecher et al ldquoMast cells increase vascu-lar permeability by heparin-initiated bradykinin formation invivordquo Immunity vol 34 no 2 pp 258ndash268 2011

[13] J N Pennefather A Lecci M L Candenas E Patak F MPinto and C A Maggi ldquoTachykinins and tachykinin receptorsa growing familyrdquo Life Sciences vol 74 no 12 pp 1445ndash14632004

[14] T Marui I Funatogawa S Koishi et al ldquoTachykinin 1 (TAC1)gene SNPs and haplotypes with autism a case-control studyrdquoBrain amp Development vol 29 no 8 pp 510ndash513 2007

[15] R DrsquoAnna G Baviera F Corrado et al ldquoNeurokinin B andnitric oxide plasma levels in pre-eclampsia and isolated intra-uterine growth restrictionrdquo BJOG An International Journal ofObstetrics and Gynaecology vol 111 no 10 pp 1046ndash1050 2004

[16] N M Page J Dakour and D W Morrish ldquoGene regulation ofneurokinin B and its receptor NK3 in late pregnancy and pre-eclampsiardquo Molecular Human Reproduction vol 12 no 7 pp427ndash433 2006

[17] A K Topaloglu F Reimann M Guclu et al ldquoTAC3 andTACR3mutations in familial hypogonadotropic hypogonadismreveal a key role for Neurokinin B in the central control ofreproductionrdquoNature Genetics vol 41 no 3 pp 354ndash358 2009

[18] M Fischer I Gemende W C Marsch and P A Fischer ldquoSkinfunction and skin disorders in Parkinsonrsquos diseaserdquo Journal ofNeural Transmission vol 108 no 2 pp 205ndash213 2001

[19] V Mesnage J L Houeto A M Bonnet et al ldquoNeurokinin Bneurotensin and cannabinoid receptor antagonists and Parkin-son diseaserdquoClinical Neuropharmacology vol 27 no 3 pp 108ndash110 2004

[20] M A Mittelman-Smith HWilliams S J Krajewski-Hall N TMcMullen and N E Rance ldquoRole for kisspeptinneurokininBdynorphin (KNDy) neurons in cutaneous vasodilatation andthe estrogen modulation of body temperaturerdquo Proceedings ofthe National Academy of Sciences of the United States of Americavol 109 no 48 pp 19846ndash19851 2012

[21] R Paus T C Theoharides and P C Arck ldquoNeuroimmuno-endocrine circuitry of the brain-skin connectionrdquo Trends inImmunology vol 27 no 1 pp 32ndash39 2006

[22] E M J Peters P C Arck and R Paus ldquoHair growth inhibitionby psychoemotional stress a mouse model for neural mecha-nisms in hair growth controlrdquo Experimental Dermatology vol15 no 1 pp 1ndash13 2006

[23] C Tusset S D Noel E B Trarbach et al ldquoMutational analysisof TAC3 and TACR3 genes in patients with idiopathic centralpubertal disordersrdquo Arquivos Brasileiros de Endocrinologia eMetabologia vol 56 no 9 pp 646ndash652 2012

[24] X Xin J Zhang Y Chang and Y Wu ldquoAssociation studyof TAC3 and TACR3 gene polymorphisms with idiopathicprecocious puberty in Chinese girlsrdquo Journal of PediatricEndocrinology and Metabolism vol 28 no 1-2 pp 65ndash71 2015

[25] M A de Souza Silva B Lenz A Rotter et al ldquoNeurokinin3receptor as a target to predict and improve learning andmemory in the aged organismrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 110 no37 pp 15097ndash15102 2013

[26] T Foroud L F Wetherill J Kramer et al ldquoThe tachykininreceptor 3 is associated with alcohol and cocaine dependencerdquoAlcoholism Clinical and Experimental Research vol 32 no 6pp 1023ndash1030 2008

[27] M Garcia-Barcelo S K King X Miao et al ldquoApplication ofHapMap data to the evaluation of 8 candidate genes for pedi-atric slow transit constipationrdquo Journal of Pediatric Surgery vol42 no 4 pp 666ndash671 2007

[28] S A Miller D D Dykes and H F Polesky ldquoA simple saltingout procedure for extracting DNA from human nucleated cellsrdquoNucleic Acids Research vol 16 no 3 article 1215 1988

[29] T Jansen S Krug P Kind G Plewig and G Messer ldquoBsm1polymorphism of the vitamin D receptor gene in patients withthe fulminant course of rosaceardquo The Journal of Dermatologyvol 31 no 3 pp 244ndash246 2004

[30] A C Yazici L Tamer G Ikizoglu et al ldquoGSTM1 and GSTT1null genotypes as possible heritable factors of rosaceardquo Photo-dermatology Photoimmunology and Photomedicine vol 22 no4 pp 208ndash210 2006

[31] A L S Chang I Raber J Xu et al ldquoAssessment of the geneticbasis of rosacea by genome-wide association studyrdquo Journal ofInvestigative Dermatology vol 135 pp 1548ndash1555 2015

[32] E AMisch and T R Hawn ldquoToll-like receptor polymorphismsand susceptibility to human diseaserdquo Clinical Science vol 114no 5-6 pp 347ndash360 2008

[33] F M N Forton ldquoPapulopustular rosacea skin immunity andDemodex pityriasis folliculorum as a missing linkrdquo Journal ofthe European Academy of Dermatology and Venereology vol 26no 1 pp 19ndash28 2012

[34] J Schauber R A Dorschner A B Coda et al ldquoInjury enhancesTLR2 function and antimicrobial peptide expression througha vitamin D-dependent mechanismrdquo The Journal of ClinicalInvestigation vol 117 no 3 pp 803ndash811 2007

[35] MAM van Steensel S Badeloe VWinnepenninckxMVree-burg P M Steijlen and M van Geel ldquoGranulomatous rosaceaand Crohnrsquos disease in a patient homozygous for the Crohn-associated NOD2CARD15 polymorphism R702Wrdquo Experi-mental Dermatology vol 17 no 12 pp 1057ndash1058 2008

[36] T Watanabe A Kitani P J Murray and W Strober ldquoNOD2 isa negative regulator of Toll-like receptor 2ndashmediated T helpertype 1 responsesrdquoNature Immunology vol 5 no 8 pp 800ndash8082004

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Evidence-Based Complementary and Alternative Medicine

Volume 2014Hindawi Publishing Corporationhttpwwwhindawicom


endothelial growth factor (VEGF) staining on skin sectionshas detected no differences between PP rosacea and ETrosacea [7] Laser Doppler Imager has detected elevatedblood flow in affected skin compared with nonaffected skinof the same patient in the PP form but not in the ET form ofrosacea [8]

Normal skin Demodex folliculorum (a head sebaceousfollicle lumen mite) usually located at the upper part of theinfundibulum of the follicle [9] has been indirectly involvedin rosacea although a possible causal correlation has notbeen established up to this day The majority of rosaceamedications do not affect Demodex but do improve rosaceawhile treatment with Lindane which is effective on the mitedoes not improve the condition [1] PossiblyDemodexmay bean exclusively exacerbating factor in rosacea patients Densityof Demodex was demonstrated to be significantly higher inthe PP form of rosacea although no correlation was ascer-tained between Demodex intrafollicular inflammation andperifollicular inflammation Possibly increased blood flow inrosacea skin dilated vessels constitutes an ideal conditionfor dermis Demodex multiplication Demodex folliculorumantigens reacting with rosacea patientsrsquo sera have beenconsidered to be responsible for stimulating mononuclearcell proliferation [10] High vascular density was significantlymore frequent in the PP form skin sections The numberof mast cells (which promote vasodilatation angiogenesisand fibrosis) was significantly higher in lesional than innonlesional skin of patients with rosacea [10]

All things considered it is evident that neuropeptidessuch as SP andor bradykinin may play a decisive role inincreasing blood flow andor vascular density in rosaceaConsequently it looked reasonable to explore the involve-ment of single nucleotide polymorphisms (SNPs) withinneuromediators and their receptors in rosacea

ldquoKininrdquo is considered as hypotensive polypeptide con-tracting most isolated smooth muscles Tachykinins (TACs)are characterized by pharmacological similarity to theirendogenous inflammatory mediator bradykinin yet TACsare not of similar structure to that of bradykinin TACsare not kinins but the name remains [11] Heparin-initiatedbradykinin formation plays an important role in mast cell-mediated diseases [12] such as rosacea

TACs are a family of closely related peptides such as SPneurokinin A and neurokinin B which act as neuropep-tides in nonneuronal cells and noninnervated tissues [13]Biological actions of TAC1 and TAC3 are mediated by threetypes of G protein-coupled receptors (TACR1 TACR2 andTACR3) TACR3 or NK-R3 or neurokinin B receptor belongsto a family of genes which function as receptors for TACscharacterized by interactionwithG proteins variations in the5-end of the sequence and 7-hydrophobic transmembraneregions interactions TAC1 gene is localized on chromosome7q21-q22 TAC3 is localized on 12q13-q21 TACR1 is localizedon 2p12 TACR2 is localized on 10q11-q21 and TACR3 islocalized on 4q25 [14] Circulating neurokinin B is elevatedin women with preeclampsia during the third trimester ofpregnancy [15] The increased placental expression of TAC3gene belongs to the mechanism responsible for the elevatedcirculating neurokinin B levels in preeclampsia [16] In

preeclampsia complicated pregnancies elevated neurokininB plasma levels have been associated with elevated nitricoxide metabolite levels possibly in a compensatory mecha-nism to improve blood flow to the uteroplacental unit [15] Infamilial hypogonadotropic hypogonadismmutations of neu-rokinin B gene and its respective receptor TACR3 suggestedan eventual key role for neurokinin B in the central control ofreproduction [17]

Rosacea is significantly more frequent in patients suffer-ing from preexisting Parkinsonrsquos disease [18] Neurokinin B(endogenous ligand of TACR3) is involved in pathogenesis ofParkinsonrsquos disease by interacting with brain dopaminergictransmission [19]

Kisspeptinneurokinin Bdynorphin-expressing neuronsof hypothalamus promoting skin vasodilatation partici-pate in the estrogen modulation of the body temperatureand contribute to the induction of ensuing hot flushes inmenopausal women such flushes are clinically similar tothose experienced in rosacea [20] The role of mast cells intachykinin neurogenic skin inflammation induction has beenshown and thus their intense presence in PP rosacea [21 22]may be explained

We chose to study the TACR3 gene polymorphismrs3733631 because TACR3 endogenous ligand neurokinin Bby increasing blood flow (anatomoclinical feature of rosacea)is involved in the pathophysiology of (a) Parkinsonism(frequently coexists with rosacea) and (b) hot flushes inmenopausal women (clinically similar to rosacea vasomotorcrises)

Possible involvement of the TACR3 gene in rosaceawould most probably be associated with either upregulationor downregulation of this gene SNPs such as rs3733631localizedwithin the promoter area upstream from the codingregion make prime candidates to reflect any associationbetween gene and disease Certain additional TACR3 poly-morphisms such as A29V G59E S455G A449S W275Xand rs4580655 have been associated with idiopathic centralpubertal disorders [23 24] predicting and improving learn-ing and memory in aged organism [25] alcohol and cocainedependence [26] and pediatric slow transit constipation [27]

2 Materials and Methods

21 Patientsrsquo Characteristics One hundred and twenty-eightpatients examined at the Outpatient Clinic of the UniversityDepartment of Dermatology at the University Hospital ofAlexandroupolis 53 males (414) and 75 females (586)were included in this study Patientsrsquo age ranged from 14to 86 years with a median age of 5625 plusmn 1635 years Allpatients were clinically diagnosed by the same physician andsuffered from either the ET form (119899 = 67) or the PP form(119899 = 61) of central face rosacea [1] Rosacea patients withaccompanying disease were excluded from the study toavoid possible interactions with potentially coexisting genet-ically determined disorders One hundred and twenty-oneunrelated subjects free from rosacea not suffering fromany potentially genetically determined disorder individuallymatched to patients by both gender and age were recruitedas controls [54 (446) males and 67 (554) females] There




Male 53 (414) 54 (446)Female 75 (586) 67 (554)





3 Results

















(a)


Total

Genotype

CC 106 (877) 65 (970) Ref

CG 14 (116) 1 (15) 012 002ndash093 0043

GG 1 (08) 1 (15) 172 011ndash2825 0704

Recessive model

CC 106 (877) 65 (970) Ref

CG or GG 15 (124) 2 (30) 022 005ndash101 0052

Allele contrast

C 226 (934) 131 (978) Ref

G 16 (66) 3 (22) 033 010ndash117 0087

Males

Genotype

CC 47 (870) 23 (920) Ref

CG 7 (130) 1 (40) 031 004ndash268 0285


Recessive model

CC 47 (870) 23 (920) Ref

CG or GG 7 (130) 2 (80) 062 012ndash326 057

Allele contrast

C 101 (935) 47 (940) Ref

G 7 (65) 3 (60) 097 024ndash395 0966

Females

Genotype

CC 59 (881) 42 (1000) mdash



Recessive model

CC 59 (881) 42 (1000) mdash


Allele contrast

C 125 (933) 84 (1000) mdash


(b)














4 Discussion












5 Conclusion




References











































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of



















Male 53 (414) 54 (446)Female 75 (586) 67 (554)





3 Results

















(a)


Total

Genotype

CC 106 (877) 65 (970) Ref

CG 14 (116) 1 (15) 012 002ndash093 0043

GG 1 (08) 1 (15) 172 011ndash2825 0704

Recessive model

CC 106 (877) 65 (970) Ref

CG or GG 15 (124) 2 (30) 022 005ndash101 0052

Allele contrast

C 226 (934) 131 (978) Ref

G 16 (66) 3 (22) 033 010ndash117 0087

Males

Genotype

CC 47 (870) 23 (920) Ref

CG 7 (130) 1 (40) 031 004ndash268 0285


Recessive model

CC 47 (870) 23 (920) Ref

CG or GG 7 (130) 2 (80) 062 012ndash326 057

Allele contrast

C 101 (935) 47 (940) Ref

G 7 (65) 3 (60) 097 024ndash395 0966

Females

Genotype

CC 59 (881) 42 (1000) mdash



Recessive model

CC 59 (881) 42 (1000) mdash


Allele contrast

C 125 (933) 84 (1000) mdash


(b)














4 Discussion












5 Conclusion




References











































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of


















(a)


Total

Genotype

CC 106 (877) 65 (970) Ref

CG 14 (116) 1 (15) 012 002ndash093 0043

GG 1 (08) 1 (15) 172 011ndash2825 0704

Recessive model

CC 106 (877) 65 (970) Ref

CG or GG 15 (124) 2 (30) 022 005ndash101 0052

Allele contrast

C 226 (934) 131 (978) Ref

G 16 (66) 3 (22) 033 010ndash117 0087

Males

Genotype

CC 47 (870) 23 (920) Ref

CG 7 (130) 1 (40) 031 004ndash268 0285


Recessive model

CC 47 (870) 23 (920) Ref

CG or GG 7 (130) 2 (80) 062 012ndash326 057

Allele contrast

C 101 (935) 47 (940) Ref

G 7 (65) 3 (60) 097 024ndash395 0966

Females

Genotype

CC 59 (881) 42 (1000) mdash



Recessive model

CC 59 (881) 42 (1000) mdash


Allele contrast

C 125 (933) 84 (1000) mdash


(b)














4 Discussion












5 Conclusion




References











































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of


















4 Discussion












5 Conclusion




References











































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of



















































of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















Research Article Assessment of Tachykinin Receptor 3 Gene...

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