Bronchoalveolar Lavage Fluid Cellularity and SolubleIntercellular Adhesion Molecule-1 in Children With Colds

Jonathan Grigg, MD, MRCP,* Josef Riedler, MD, and Colin F. Robertson, FRACP

Summary. Viral colds are an important cause of respiratory symptoms in normal children.Studies in adults suggest that inflammation in the lower respiratory tract is associated with viralcolds, but there are no data regarding inflammation and viral infection in the lower airway ofnormal children with colds. We, therefore, studied the lower airway of two groups of children:Group I, those with active coryzal symptoms and a respiratory virus isolated from bronchoal-veolar lavage fluid (BALF); and Group II: asymptomatic children who had had a clinical coldwithin the previous 2 weeks and no respiratory virus in BALF. Both groups were compared toage- and weight-matched normal noninfected controls, who had had no coryzal symptoms for atleast 8 weeks.

Viruses isolated from BALF of Group I (n = 7) were: respiratory syncytial virus (n = 2),rhinovirus (n = 3), parainfluenza I (n = 1), and echovirus 11 (n = 1). Compared to normal controls,Group I had an increased BALF lymphocyte and neutrophil differential count (P < 0.05), aconcomitant depressed alveolar macrophage differential count (P < 0.05), and increased BALFconcentrations of soluble intercellular adhesion molecule-1 (sICAM-1) (P < 0.05, n = 6), totalprotein (P < 0.05, n = 6) and albumin (P < 0.05, n = 7). Similar changes were seen in Group II(n = 22), with an increased BALF neutrophil (P < 0.05) and lymphocyte (P < 0.01) differentialcount, and increased concentrations of sICAM-1 (P < 0.01, n = 15), total protein (P < 0.0001, n= 9) and albumin (P = 0.05, n = 17).

Our results suggest that inflammation and viral infection in the lower airway are present duringactive colds, and that inflammation is also present during the convalescent period. PediatrPulmonol. 1999; 28:109–116. © 1999 Wiley-Liss, Inc.

Key words: colds; virus; soluble intercellular adhesion molecule-1; bronchoalveolar.

INTRODUCTION

Rhinitis, sneezing, cough, and mild pyrexia are clinicalsigns of viral infection of the upper respiratory tract (thecommon cold). Despite the high prevalence of coryzalsymptoms in children, there are no data on how coldsinfluence their lower airway. A description of changes inthe pediatric lower airway during colds is of potentialinterest, since it might provide clues into how virusesmodulate immune function in the developing lung. Viralinfection and virally induced inflammation in the lowerairway are one way whereby colds could influence thepulmonary immune milieu. In adults, there is a link be-tween coryzal symptoms and neutrophilic inflammationin the lower respiratory tract.1,2 However, it is not knownwhether lower airway inflammation during colds is ini-tiated by direct infection of the lower airway. Since re-spiratory viruses in vitro are able to infect bronchial cellsand upregulate expression of proinflammatory cyto-kines,3 we hypothesized that direct infection of the lowerrespiratory tract and inflammation occur during trivialpediatric colds.

The pattern of inflammatory cells and mediators in thelower respiratory tract of children during the convales-cent phase of a cold is also unknown, but studies of the

upper airway suggest that virally induced inflammationmay persist beyond the resolution of coryzal symptoms.For example, concentrations of tumor necrosis factor(TNF)-a in the nose remain elevated for up to 3 weeksafter the resolution of coryzal symptoms.4 Furthermore,lower airway cells are primed to release increased levelsof TNF-a to antigen challenge during the convalescentphase of experimental colds in adults.5

We sought to determine in this study 1) whether vi-ruses associated with the common cold infect the lowerrespiratory tract; 2) whether viral infection of the lowerairway is associated with an inflammatory response; and3) whether lower airway inflammation persists after theresolution of coryzal symptoms. To define the presence

Department of Thoracic Medicine, Royal Children’s Hospital, Mel-bourne, Victoria, Australia.

*Correspondence to: Dr. Jonathan Grigg, currently at the LeicesterChildren’s Asthma Centre, University of Leicester School of Medi-cine, Robert Kilpatrick Clinical Sciences Building, Leicester RoyalInfirmary, PO Box 65, Leicester LE2 7LX, United Kingdom.E-mail: jg33@le.ac.uk

Received 4 January 1999; Accepted 14 April 1999.

Pediatric Pulmonology 28:109–116 (1999)

© 1999 Wiley-Liss, Inc.

of viral infection and inflammation, we used bronchoal-veolar lavage (BAL) to sample cells and protein fromchildren with active and recent colds. Concentrations ofsoluble intercellular adhesion molecule-1 (sICAM-1)were also measured in bronchoalveolar lavage fluid(BALF), since it is a marker of upregulation of the ad-hesion molecule ICAM-1 on pulmonary cells,6 and is anonspecific marker of pulmonary immune cell activa-tion.7,8

MATERIALS AND METHODSSubjects

Children between 2 months and 18 years of age intu-bated for elective orthopedic or abdominal surgery wereeligible for this study. The exclusion criteria were: pre-mature birth, chronic respiratory pathology (includingasthma and chronic rhinitis), acute or chronic systemicdisease, and active atopic disease. The study was ap-proved by the Ethics in Human Research Committee ofthe Royal Children’s Hospital and required written in-formed parental consent.

Assessment of Symptoms

On the day of the BAL, children were examined andthe parents interviewed by a clinician. Respiratory signsand symptoms were recorded, and the parents and childwere asked about previous acute respiratory symptoms.Exposure to parental cigarette smoke was assessed byasking the parents, “Do either you or your partnersmoke?” Recent coryzal symptoms were defined as aself-limiting illness associated with new onset of rhinitis,cough, or both, that lasted less than 7 days, was notassociated with a systemic illness requiring medical at-tention, and that was described as a typical “cold” by theparents. The policy of the anesthetic unit was to allowelective surgery on children with colds, if the symptomswere clinically trivial. Surgery was postponed in childrenwith clinically significant respiratory symptoms.

Study Groups

Two groups of children were studied: Group I (activecold), children with coryzal symptoms and a respiratoryvirus isolated from the BALF; and Group II (convales-

cent), children with a recent coryzal illness, who wereasymptomatic for 1 to 14 days before BAL, and in whomno bacteria or respiratory virus was detected in theBALF. Since we previously showed that the BAL leu-kocyte differential counts change with age,9,10 the studygroups were compared to age-matched normal controlswho had been asymptomatic in the preceding 8 weeksbefore BAL, and who did not have viruses or bacteriaidentified in their BALF.

The age-matched controls were from a group of nor-mal children who underwent BAL during the period ofthis study. Controls were recruited to describe qualitativeand quantitative changes in leukocytes during normaldevelopment, and had had no chronic, acute, or recent(within 8 weeks) respiratory symptoms, and no evidenceof atopic inflammation (hay fever, asthma, eczema). Twoweeks after BAL, all parents were asked about the de-velopment of new respiratory symptoms and the resolu-tion of existing symptoms.

Bronchoalveolar Lavage

BAL was performed before elective surgery by asingle operator, using a nonbronchoscopic BAL tech-nique.11 Children did not receive anticholinergic pre-medication. Anesthesia was achieved with isofluorane,intravenous thiopentone, and pancuronium. Prior to in-tubation, any secretions in the upper airway were aspi-rated. Immediately after intubation, and with the child’shead turned to the left, a straight 60-cm end-hole suctioncatheter (Vygon S.A., E´ couen, France) was inserted intothe endotracheal tube (ET) through a right-angle swivelbronchoscope connector (Mallinckrodt Medical Pty.Ltd., Mount Waverly, Australia). Catheter size was ad-justed to the ET diameter (size 6 French catheter for size<3.5 ET; 7 French catheter for size 3.5 ET; 8 Frenchcatheter for size >3.5 to 5.0 ET; 10 French catheter forsize$5.5 ET). The catheter was advanced until wedged,1 mL/kg body weight of saline at room temperature wasinstilled, and BALF was immediately aspirated into asuction trap, using 150 mmHg negative pressure and athree-way stopcock. With the catheter remaining wedgedin situ, two further aliquots of 1 mL/kg saline were in-stilled and aspirated (i.e., a total instilled volume of 3mL/kg). Immediately after collection, the concentrationof leukocytes was measured on unfiltered BALF by he-mocytometer, and viability of cells was determined byexclusion of a 0.04% solution of Trypan blue. Ciliatedepithelial cells in the hemocytometer chamber were notcounted. Unfiltered BALF was cytocentrifuged at 9/g for5 min (Cytospin 2, Shandon Products Ltd., Runcorn,UK) and stained with Wilson’s reagents, and slides wereexamined under oil immersion by microscopy (×1,000magnification). BALF leukocyte differential counts weredetermined by counting$300 leukocytes. The BALF

Abbreviations

BAL Bronchoalveolar lavageBALF Bronchoalveolar lavage fluidET EndotrachealRSV Respiratory syncytial virussICAM-1 Soluble intercellular adhesion molecule-1TNF-a Tumor necrosis factor-a

110 Grigg et al.

epithelial cell differential count was determined sepa-rately by counting 200 nucleated cells. Particulate-freeBALF supernatant was obtained by centrifugation ofBALF at 1,000g for 10 min (4°C) and 8,000g for a fur-ther 3 min (4°C). Aliquots were stored at −70°C for totalprotein, albumin, and sICAM-1 analysis.

BALF Microbiology and Virology

Aliquots of 0.5 mL BALF were analyzed for respira-tory viruses, and aerobic and anaerobic bacteria. Nasalsamples were not obtained. Viruses were detected bycytopathic effect during culture with human fibroblasts,MDCK, LLC, and Ohio HeLa cell lines. All cell lineswere cultured in roller tubes at 33°C with 5% carbondioxide, and examined every other day for cytopathiceffect for a total of 21 days. The cytopathic effect of anisolate was confirmed by acid stability testing (rhinovi-rus),12 immunofluorescence (RSV, parainfluenza virus 1,2, and 3, and adenovirus), or hemadsorption (influenzavirus).4 Direct immunofluorescence for RSV, influenzaA and B, parainfluenza, and coronavirus was performedon BAL cells concentrated by centrifugation.13 Primary,secondary, and tertiary aerobic and anaerobic bacterio-logical cultures were performed on 10mL BALF. Allpotential bacterial pathogens were identified and re-ported, using a semiquantitative analysis that had previ-ously been validated using quantitative cultures.11

BALF Total Protein, Albumin, and sICAM-1

Total protein and albumin concentrations were mea-sured on duplicate BALF samples, using a Roche Cobascentrifugal analyzer (Roche Diagnostics, Basel, Switzer-land). Total protein in 50mL BALF was measured usingbenzethonium chloride. The linear dynamic range for theassay was from 3.0–400 mg/L. Albumin was determinedby immunoturbidometry with an assay linear dynamicrange from 5.0–800 mg/L. sICAM-1 was measured onduplicate BALF samples using an ELISA kit (PredictatICAM-1, Genzyme Diagnostics, Cambridge, MA) with1:10 and 1:50 dilutions of particulate-free BALF. Theassay sensitivity was 0.09 ng/mL, with a linear dynamicrange from 0.5–9.0 ng/mL. The intra-assay coefficient ofvariation was 6.9%.

Statistical Analysis

Age- and weight-matched groups were analyzed aspaired data by Wilcoxon’s signed ranks test. Data weresummarized as medians (25th and 75th percentiles). Avalue of P < 0.05 was considered significant. Analysiswas performed using a statistical package for microcom-puters (SPSSt for Windowsy, SPSS, Inc., Chicago, IL).

RESULTSGroup I

Seven children, ranging in age from 3 months to 40months had a combination of trivial coryzal symptoms

TABLE 1—Viral Isolates and BALF Parameters in Children With an Active Cold (Group I) 1

Patient age(months) BALF virus BALF bacteria

Respiratorysymptoms

Symptomduration(days)

AM(%)

LYMPH(%)

NEUT(%)

EO(%)

sICAM-1(ng/mL BALF)

Albumin(mg/LBALF)

3.3 Parainfluenza I S. pneumoniae Cough 1 73.8 26.0 0.3 0.0 98.4 896.8 RSV Nil Rhinitis, cough 2 11.3 3.5 85.3 0.0 143.4 37.9 Rhinovirus Nil Rhinitis 2 60.3 4.0 35.7 0.0 102.4 278.7 Rhinovirus Nil Rhinitis, cough 7 73.3 25.7 1.0 0.0 159.8 96

10.1 RSV Nil Sneezing, cough 1 93.0 6.7 0.3 0.0 74.6 3512.5 Echovirus 11 Nil Cough 1 25.9 2.0 72.1 0.0 Not done 8740.1 Rhinovirus Nil Rhinitis, cough 1 29.7 5.0 63.3 2.0 39.4 75

1RSV, respiratory syncytial virus; AM, alveolar macrophages; LYMPH, lymphocytes; NEUT, neutrophils; EO, eosinophils; sICAM-1, solubleintercellular adhesion molecule-1.

TABLE 2—Demographic Data and Lavage Dynamics in Groups I and II Compared With Normal Controls 1

Group I(n 4 7)

Group I controls(n 4 7)

Group II(n 4 22)

Group II controls(n 4 22)

Gender (M/F) 4/3 3/4 14/8 15/7Parental smoking (n) 3 3 6 10Age (months)2 8.6 (6.7, 12.5) 8.2 (6.4, 16.1) 20.0 (7.3, 55) 22.6 (7.8, 52)Weight (kg)2 8.1 (6.3, 11.2) 8.6 (7.4, 12.0) 12.4 (9.6, 16.4) 13.8 (8.4, 18.3)Volume of saline instilled (mL)2 24.2 (19, 33) 26.1 (22, 36) 37.3 (28, 49) 38.4 (23, 54)Bronchoalveolar lavage fluid volume (mL)2 8.6 (7.6, 15) 15.0 (6.2, 24) 18.4 (11.3, 24) 19.4 (8.6, 29)Bronchoalveolar lavage fluid return (%)2 42.4 (30, 64) 47.7 (33, 59) 46.9 (36, 56) 45.9 (36, 55)

1Values are expressed as medians (25th, 75th percentiles).2No difference in Group I compared with controls, and in Group II compared with controls, Wilcoxon’s signed rank test.

Colds and Lower Airway Inflammation 111

and a respiratory virus in BALF (Table 1). Six childrenhad a trivial cough at the time of BAL, but none hadcrackles or wheeze on clinical examination. In one child,viral infection was associated with the isolation ofStrep-tococcus pneumoniae(104 colony-forming units/mLBALF) (Table 1). Six children had no bacterial pathogenin the BALF. Respiratory symptoms in Group I remainedclinically trivial, and all children had become asymptom-atic within 5 days of BAL. Age, weight, and volume ofBALF obtained was similar in Group I and controls(Table 2). Leukocyte viability was >95% in subjects andcontrols. The isolation of a virus was associated with areduced alveolar macrophage differential count (P <0.05, Fig. 1A), and an increased neutrophil and lympho-cyte differential count (P < 0.05, Fig. 1B,C, Table 3).There was no difference in the ciliated epithelial celldifferential count, or the concentration of alveolar mac-rophages and eosinophils (Table 3). However, the BALFlymphocyte concentration was increased (P < 0.05), andthere was a trend for an increased concentration of neu-trophils (P 4 0.07, Table 3). The isolation of a virusfrom the BALF was also associated with an increasedconcentration of sICAM-1 (n4 6), total protein (n4 6),and albumin (n4 7, P < 0.05, Table 4).

Group II

Twenty-two asymptomatic children ranging in agefrom 3–159 months had had a recent coryzal illness andno BALF virus. No bacterial pathogens were isolatedfrom BALF of this group. The median duration from thelast coryzal symptoms was 3.5 days (range, 1–7 days),and all children continued to be asymptomatic for 7 daysafter BAL. Age, weight, BALF volume, and percentageof BALF return in Group II did not differ significantlyfrom controls (Table 2). Leukocyte viability was >95%in all children. Group II had a reduced alveolar macro-phage differential count (P < 0.0001, Fig. 2A, Table 3),and an increased neutrophil (P < 0.05, Fig. 2B) and lym-phocyte differential count (P < 0.01, Fig. 2C). The BALFalveolar macrophage concentration was not different, butthe concentration of neutrophils and lymphocytes wasincreased compared to controls (P < 0.05, Table 3). Mea-surement of protein and sICAM-1 was not available forall age-matched controls for technical reasons (insuffi-cient sample volume stored). Where pairing was pos-sible, Group II had an increased BALF concentration ofsICAM-1 (P < 0.01, n4 15), total protein (P < 0.0001,n 4 9), and albumin (P 4 0.05, n4 17, Table 4).

DISCUSSION

We have shown that inflammation and viral infectionsof the lower respiratory tract can occur with clinicallytrivial colds, and that inflammatory changes persist for

up to 1 week after complete resolution of coryzal symp-toms. In children with a respiratory virus in the BALF(Group I), we found a reduced alveolar macrophagesdifferential count, and an increased neutrophil and lym-phocyte differential count. A similar pattern of lowerairway inflammation has been reported in normal adultswith community-acquired colds. Trigg et al.2 found thatactive colds are associated with an increased neutrophil

Fig. 1. Comparison of bronchoalveolar fluid leukocyte differen-tial counts between Group I (lower respiratory tract virus) andnormal controls for (A) alveolar macrophages, (B) neutrophils,and (C) lymphocytes. * P < 0.05, Wilcoxon’s signed rank test. Barindicates median.

112 Grigg et al.

differential count in bronchial washings when comparedto preinfection levels. By contrast, Pizzichini et al.1 innonasthmatic adults found no significant change in theneutrophil differential count or concentrations betweenthe acute and convalescent phase of community-acquiredcolds. However, subject numbers were small (n4 6) inthat study, and the authors reported a “moderate neutro-philia” in induced sputum obtained during the acute phase.In children with congenital heart disease, we previouslyreported that a common cold virus in BALF is associatedwith a reduced alveolar macrophage differential count,and an increased lymphocyte differential count.11 Thismirrors the pattern seen in the present study, although thenormal children with respiratory virus in BALF also hadan increased neutrophil differential count.

A change in the differential count for any leukocyte isdue to either a fall in its number, or a quantitative in-crease of another leukocyte subpopulation. Since the al-veolar macrophage concentration did not change duringviral infection, the lower alveolar macrophage differen-tial count in Group I must be due to an increased numberof lymphocytes, and to a lesser extent, increased concen-trations of neutrophils. Inflammatory changes in Group Iwere not due to concomitant bacterial infection, since abacterial pathogen was isolated with a respiratory virusin only one child. This contrasts with our findings inchildren with congenital heart disease, where bacterialpathogens were isolated from the majority of virus-infected children.11 Could the quantitative changes in

leukocytes be caused by variations in lavage dynamicsbetween subjects and controls? In order to avoid thispossibility, and to control for the known developmentalchanges in leukocyte subpopulations,9,10 we used age-and weight-matched normal controls. No difference wasfound in the volume instilled, percentage of BALF re-covered, and leukocyte viability between study groupsand controls. This suggests that the area of lung sampledwas equally matched in patients and controls, and thatany differences in cell and solute concentrations were notartifactual.

Study Groups I and II do not represent the whole clini-cal spectrum of coryzal illness, and the prevalence oflower airway viral infection and inflammation with co-ryzal symptoms cannot be estimated from our study.However, infection of the lower respiratory tract mustoccur in at least a proportion of children with trivialcolds, since the viruses isolated from BALF were iden-tical to those isolated from nasal samples during com-munity-acquired coryzal illness,14,15and the spectrum ofrespiratory symptoms was similar to that reported foradults with viral colds.2 Could the BALF viral isolates bedue to contamination from infected upper respiratorytract secretions? This is unlikely. Firstly, the suctioncatheter was passed through an endotracheal tube, whichin turn had been inserted after thorough suctioning of theoropharynx. Secondly, using the same BAL technique,we previously found that the isolation of a respiratoryvirus from the nasopharynx was not associated with its

TABLE 4—Bronchoalveolar Lavage Fluid Solutes in Groups I and II Compared With Normal Controls 1

BALF solute Group I Group I controls Group II Group II controls

Total protein (mg/L) 152 (90, 223),* n4 6 59 (16, 29), n4 6 88.0 (74, 108),* n4 9 63.0 (30, 64), n4 9Albumin (mg/L) 87 (35, 96),* n4 7 19 (16, 29), n4 7 37.0 (28, 53),** n4 17 30.0 (22, 39), n4 17sICAM-1 (ng/mL) 100 (65, 147),* n4 6 48 (27, 59), n4 6 49.0 (33, 66),** n4 15 31.1 (22, 48), n4 15

1Values are expressed as medians (25th, 75th percentiles).*P < 0.05 compared with controls, Wilcoxon’s signed rank test.** P < 0.01 compared with controls, Wilcoxon’s signed rank test.

TABLE 3—Bronchoalveolar Lavage Fluid Cellularity in Groups I and II Compared With Normal Controls 1

BALF parameterGroup I(n 4 7)

Group I controls(n 4 7)

Group II(n 4 22)

Group II controls(n 4 22)

LEUK (× 103/mL) 210 (180, 890)* 130.0 (100, 190) 140.0 (87, 190) 80.0 (50, 115)AM (%) 60 (25, 73)* 96.0 (95, 97) 83.0 (73, 91)*** 93.0 (90, 96)AM (× 103/mL) 147 (139, 263) 122.0 (95, 184) 121.0 (65, 157) 73.0 (46, 112)LYMPH (%) 5 (3.5, 25)* 2.5 (1.6, 3.0) 9.6 (6, 13)** 5.5 (2.3, 7.0)LYMPH (× 103/mL) 44 (10, 54)* 3.0 (2.5, 4.4) 11.2 (7.1, 24)* 4.7 (2.4, 5.7)NEUT (%) 35 (0.3, 72)* 1.0 (0.3, 2.2) 3.4 (1.5, 7.4)* 1.0 (0.3, 2.4)NEUT (× 103/mL) 64 (0.5, 563) 1.0 (0.6, 2.2) 4.7 (1.9, 11.2)* 1.0 (0.2, 2.2)EOS (%) 0 (0, 0) 0.0 (0, 0) 0.0 (0, 0.3) 0.0 (0, 0)EPI (% nucleated) 2 (1.5, 7.2) 3.0 (1.0, 9.0) 2.5 (0.7, 5.2) 4.0 (3, 7.5)

1Values are expressed as medians (25th, 75th percentiles). BALF, bronchoalveolar lavage fluid; LEUK, total leukocytes; AM, alveolarmacrophage; LYMPH, lymphocytes; NEUT, neutrophils; EOS, eosinophils; EPI, ciliated epithelial cells (as a percentage of all nucleated cells).*P < 0.05 compared with controls, Wilcoxon’s signed rank test.** P < 0.01 compared with controls, Wilcoxon’s signed rank test.*** P < 0.0001 compared with controls, Wilcoxon’s signed rank test.

Colds and Lower Airway Inflammation 113

presence in the BALF.11 Thirdly, experimental studieshave shown that viruses that cause the common cold caninfect lower airway cells. For example, rhinovirus repli-cates in pulmonary epithelial cell lines, and their RNAhave been demonstrated in lower airway cells fromadults after experimental inoculation.16 The proinflam-

matory potential of rhinovirus infection is suggested bythe observation that bronchial epithelial cells infectedwith rhinovirus in vitro release interleukin-8,17 and ex-press increased levels of ICAM-1.18 The present studyvalidates these experimental observations by demonstrat-ing that rhinovirus can indeed infect the lower airwayduring community-acquired infection, and initiate in-flammation in vivo. Our data also suggest that this is truefor other common respiratory viruses.

The elevated BALF protein in Group I suggests thatalveolar-capillary permeability to protein was increasedby viral infection. This finding is compatible with studiesof the upper airway during colds. Increased nasal totalprotein and albumin concentrations in lavage fluid havebeen reported for adult volunteers with experimentalcolds, and children with community-acquired coryzalsymptoms.4,19 The increased levels of sICAM-1 inBALF during colds suggest that the protein leak is ac-companied by upregulation of ICAM-1 in bronchoalveo-lar cells. Increased expression of ICAM-1 in bronchialepithelial cells has been reported in atopic adults withcoryzal symptoms.2 Because of the relatively small num-ber of children studied, we could not control for otherconfounding factors in the environment. For example,parental smoking increases BALF sICAM-1, but doesnot affect the BALF leukocyte differential, or albuminconcentrations.20 We could not match all subjects andcontrols for the combination of age, weight, and parentalsmoking, and did not measure a serum marker of envi-ronmental cigarette exposure. However, a similar propor-tion of children exposed to parental smoke were in boththe patient and normal control groups, with slightly moreparental smokers in the normal controls for Group II. Aconfounding effect of parental smoking on sICAM-1 istherefore unlikely, but cannot be completely excluded.sICAM-1 contains most of the extracellular domains ofICAM-1, and is produced when surface ICAM-1 is shedinto the extracellular space.21 Since pulmonary ICAM-1expression is ubiquitous,6 and includes alveolar macro-phages22 and bronchial epithelial cells,23 the precise ori-gin of virally induced alveolar sICAM-1 is not known.Animal models have shown that sICAM-1 blocks theentry of rhinovirus into pulmonary cells,24 but also acti-vates alveolar macrophages and enhances lung injury.25

Thus, virally induced sICAM-1 has the potential to bothattenuate and enhance pulmonary inflammation.

We have shown alterations in the BALF leukocyteprofiles, and elevated protein in BALF of children withactive colds, even after the resolution of coryzal symp-toms. The pattern of lower airway inflammation in GroupII was similar to that in Group I, but the changes were notas profound (Table 3). This indirectly supports the hy-pothesis that trivial coryzal symptoms in children arefrequently associated with infection and inflammation inthe lower respiratory tract. Noah et al.4 reported in chil-

Fig. 2. Comparison of bronchoalveolar fluid leukocyte differen-tial counts between Group II (recent coryzal illness, no bron-choalveolar lavage fluid respiratory virus), and age- and weight-matched normal controls for (A) alveolar macrophages, (B) neu-trophils, and (C) lymphocytes. * P < 0.05, **P < 0.01, ***P < 0.0001,Wilcoxon’s signed rank test. Bar indicates median.

114 Grigg et al.

dren 2–3 weeks after a clinical cold that the mean(±SEM) percentage of neutrophils in nasal lavage fluidwas 79 ± 7%, compared to the preinfection level of 37 ±7%. Similarly, we found that the BALF neutrophil dif-ferential count and concentration were increased duringthe convalescence phase. The lymphocyte differentialcount and concentration were also increased in Group II,and 3 children had lymphocyte differentials significantlyabove the 95th percentile for normal children.26 Tran-sient and very high BALF lymphocyte differential counts(>30%) have been reported in normal adult volunteers, aphenomenon that has been attributed to recent “asymp-tomatic” colds.27 The present study supports this specu-lation by providing direct evidence that the concentrationof BALF lymphocytes is increased for up to 1 week aftera cold. Persistence of inflammation after colds could bedue to very low levels of virus, continued release ofchemoattractants from pulmonary cells, or relatively de-layed clearance of leukocytes from the airway. Viruseswere not isolated from Group II, but it is possible thatpolymerase chain reaction analysis would have detectedlow levels of virus.28

In summary, we have shown that viral infection of thelower respiratory tract can occur in children with trivialcoryzal symptoms. Lower respiratory tract infections areassociated with inflammation, protein leak, and the ac-cumulation of sICAM-1 in the lower airway. Further-more, similar inflammatory changes are present in thelower airway after resolution of coryzal symptoms. Theimportance of virally induced lower airway inflammationon the developing lung remains to be determined.

ACKNOWLEDGMENT

We thank Dr. Eric Uren (Department of Virology,Royal Children’s Hospital, Melbourne) for performingviral cultures.

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