www.bjorl.org
Brazilian
Journal
of
OTORHINOLARYNGOLOGY
ORIGINAL
ARTICLE
Nasal
lavage
cytology
and
mucosal
histopathological
alterations
in
patients
with
rhinitis
夽
Loreni
C.S.
Kovalhuk
a,
Ederaldo
Queiroz
Telles
b,
Monica
Nunes
Lima
b,
Nelson
A.
Rosario
Filho
a,b,∗aUniversidadeFederaldoParaná(UFPR),ProgramadePós-Graduac¸ãoemSaúdedaCrianc¸aedoAdolescente,Curitiba,PR,Brazil bUniversidadeFederaldoParaná(UFPR),Curitiba,PR,Brazil
Received15August2018;accepted8January2019 Availableonline22February2019
KEYWORDS Allergicrhinitis; Eosinophil; Basementmembrane; Airwayremodeling Abstract
Introduction:Theextentofepitheliallesioninallergicandnon-allergicrhinitisandits associ-ationwithinflammatorychangesinnasallavagehasnotbeenclarified.
Objective:Toverifytheassociationbetweentheinflammatorycellsinthenasallavage, epithe-liallesion extent andbasement membranethickness, inthenasal mucosa ofpatients with rhinitis;todetermine thecutoff point ofthepercentageofeosinophils inthenasallavage associatedwiththeatopicpatients.
Methods:Patients with rhinitis and indication for septoplasty and (or) turbinectomy for turbinatehypertrophywereselected,andweresubmittedtoallergyskintests,nasallavage withmeasurementofalbuminandinterleukin-8levels,totalanddifferentialcountingofcells, andmucosalhistopathologicalanalysistodeterminetheextentofepitheliallesion,anddegree ofbasementmembranethickening.
Results:Fifty-sixpatients withamedianageof24.5yearsandadiagnosisofallergic rhini-tis(n=36)andnon-allergicrhinitis(n=20)werestudied.Inatopicsubjects,allergyskintests were positive for Dermatophagoides pteronyssinus in35 (97.0%)andLolium perennein 18 (50.0%).Atopicsubjectsshowedahigherclinicalscoreindexofrhinitiscomparedtonon-atopic ones. Thetotal countofcells,neutrophils,andlevelsofalbuminandIL-8 werenot differ-ent inthe nasal lavageof atopicand non-atopic subjects. The cutoff point for eosinophil count in nasal fluid for the distinction between allergic rhinitis and non-allergic rhinitis
夽 Pleasecitethisarticleas:KovalhukLC,TellesEQ,LimaMN,FilhoNA.Nasallavagecytologyandmucosalhistopathologicalalterations
inpatientswithrhinitis.BrazJOtorhinolaryngol.2020;86:434---42.
∗Correspondingauthor.
E-mail:nelson.rosario@ufpr.br(N.A.RosarioFilho).
PeerReviewundertheresponsibilityofAssociac¸ãoBrasileiradeOtorrinolaringologiaeCirurgiaCérvico-Facial.
https://doi.org/10.1016/j.bjorl.2019.01.005
1808-8694/©2019Associac¸˜aoBrasileiradeOtorrinolaringologiaeCirurgiaC´ervico-Facial.PublishedbyElsevierEditoraLtda.Thisisanopen accessarticleundertheCCBYlicense(http://creativecommons.org/licenses/by/4.0/).
was4%.Somedegreeofepitheliallesionwasmorefrequentinallergicrhinitis(94%)thanin non-allergicrhinitis(65%)patients.Inthepresenceofbasementmembranethickness,asamarker ofremodeling,therewasnodifferenceinthenasallavageofpatientswithallergicrhinitisand non-allergicrhinitis.
Conclusion: Inthisseries,4%wasthecutoffpointforthenumberofeosinophilsinthenasal lavage,foratopydifferentiation.Upperairwayremodelingaccessedbybasementmembrane thickness showed similar inflammatorycell infiltrate in thenasal lavage, regardless ofthe presenceofatopy.
© 2019 Associac¸˜ao Brasileira de Otorrinolaringologia e Cirurgia C´ervico-Facial. Published by Elsevier Editora Ltda. This is an open access article under the CC BY license (http:// creativecommons.org/licenses/by/4.0/). PALAVRAS-CHAVE Rinitealérgica; Eosinófilo; Membranabasal; Remodelamentoda viaaérea
Citologiadolavadonasalealterac¸õeshistopatológicasnamucosanasaldepacientes comrinite
Resumo
Introduc¸ão: Aextensãodalesãoepitelialnarinitealérgicaenãoalérgicaesuaassociac¸ãocom alterac¸õesinflamatóriasnolavadonasalaindanãoestãoesclarecidas.
Objetivo: Verificararelac¸ãoentreoprocessoinflamatórionolavadonasal,extensãodalesão epitelialeespessamentodamembranabasalnamucosanasaldepacientescomrinite; deter-minaropontodecortedopercentualdeeosinófilosnolavadonasalassociadoàpresenc¸ade atopia.
Método: Foramselecionadospacientescomriniteeindicac¸ãodeseptoplastiae(ou) turbinecto-miaporhipertrofiadeconchasnasais,osquaisforamsubmetidosaostestescutâneosalérgicos, lavadonasalcomdeterminac¸ãodasconcentrac¸õesdealbumina,interleucina-8(IL-8),contagem totalediferencialdecélulas,análisedaextensãodalesãoepitelial,egraudeespessamento damembranabasalpormeiodehistopatologiadamucosa.
Resultado: Foramestudados56 pacientes, mediana de24,5 anoscomdiagnóstico derinite alérgica(n=36)erinitenãoalérgica(n=20).Nosatópicosostestescutâneosalérgicosforam positivosparaD.pteronyssinusem35(97,0%)eL.perenneem18(50,0%).Atópicos apresen-tarammaioríndicedeescoreclínicoparariniteemcomparac¸ãoanãoatópicos.Acontagem totaldecélulas,neutrófiloseníveisdealbuminaeIL-8nãofoidiferenteentreolavadonasal deatópicosenãoatópicos.Opontodecortedacontagemdeeosinófilosnofluidonasalparaa distinc¸ãoderinitealérgicaerinitenãoalérgicafoide4%.Algumgraudelesãoepitelialfoimais frequenteempacientescomrinitealérgica(94%)doqueempacientescomrinitenãoalérgica (65%).Napresenc¸adeespessamentodamembranabasal,comomarcadorderemodelamento, nãohouvediferenc¸anolavadonasalentrepacientescomrinitealérgicaenãoalérgica. Conclusão:Nestacasuística,4%foiopontodecortedonúmerodeeosinófilosnolavadonasal, para diferenciac¸ão deatopia.Remodelamento davia aéreasuperior pelo espessamento da membranabasalrevelouinfiltradosemelhantedecélulasinflamatóriasnolavadonasal, inde-pendentementedapresenc¸adeatopia.
© 2019 Associac¸˜ao Brasileira de Otorrinolaringologia e Cirurgia C´ervico-Facial. Publicado por Elsevier Editora Ltda. Este ´e um artigo Open Access sob uma licenc¸a CC BY (http:// creativecommons.org/licenses/by/4.0/).
Introduction
Rhinitis is a chronic, prevalent disease with a complex integration between multiple genetic and environmen-tal factors, interconnected by mechanisms associated or not withIgE. The association withother allergic diseases and phenotypes related to multiple allergen sensitiza-tioninfluencesthe intensity,frequency andpersistenceof symptoms.1Thechronicinflammatoryprocessinthe respi-ratorymucosacanleadtostructuralalterationswithairway remodeling,wellcharacterizedinasthmaticpatients,2---4but
toalesserextentinthosewithrhinitis.5---11Allinflammatory diseasesresultinremodeling,whichcanprogresstoa nor-malorpathologicalreconstructionprocess.4Inrhinitis,itis characterizedbyincreasedthicknessandepithelial detach-mentandpseudofibrosisofthebasementmembrane.10
The inflammatory reaction and the remodeling of the nasal turbinate mucosa results in turbinate edema and (or) hypertrophy, of which clinical consequence is nasal obstruction.9,12 Additionally, the variations in the engorgement of the complex arterial vasculature and of cavernousvenoussinusoidsalsocontributetoseverenasal
obstruction.3,13 An increase in the number of eosinophils inthenasal mucosaisthe parameterthat showsthe best correlationwiththenasalobstructionsymptom.12
Theextentoftheepitheliallesioninthedifferenttypes ofrhinitisandthecorrelationwithinflammatorycellsand mediatorsisyettobeclarified.Thepresenceofeosinophils isassociatedwithlossofepithelialintegrityinpatientswith allergic or non-allergic rhinitis.14 However, there are dis-agreements regarding the cutoffpoint for the number of eosinophilsconsideredhighinnasalsecretionsamples,since thenasalsecretioncollectionprocedurecaninterferewith therecoveredcellularity.15---17Thenasalmucosa,duetothe easilyobtainedsamples,allowsthestudyofcellalterations duringthe allergic reaction.15 Nasal lavageis a relatively noninvasiveandeasytoperformtechniqueforthe quanti-tative measurement of cell distributionand inflammatory mediators.3,18---20Thesimultaneousanalysisofthecell infil-trate andthe degree of cell activation of nasal secretion andnasalmucosabiopsysamplesshowsdifferencesbetween thetwocompartments.21 Thus,thestudyofthe inflamma-tory process in samples simultaneously obtained by nasal lavage(NL)andfromthenasalmucosamayhelpto under-standthe mechanisms involved in allergic or non-allergic nasalinflammatoryreaction.
The aim of this study was toevaluate the association betweentheinflammatoryprocessinthenasalcavitylumen and the extent of the nasal mucosa lesion in atopic and nonatopicpatientswithrhinitis.
Methods
Patientswithallergicornonallergicobstructiverhinitis,with surgicalindicationfor septaldeviation correctionand(or) turbinectomytoalleviatenasalobstruction,wereselected attheOtorhinolaryngologyServiceofHospitaldeClínicasof UniversidadeFederal doParaná.Patientswithahistoryof upperandlowerairwayinfection,respectively,at2and4 weekspriortoclinicalevaluationwereexcluded;inaddition topatientswithahistoryofrecentuseofmedicationssuch assystemic and/ornasal topicalcorticosteroids,disodium cromoglycateandoraland/ornasaltopicalantihistamines. The clinicaldiagnosis andrhinitis severityclassification were attained using a symptom-adjusted (pruritus sneez-ing,nasal obstruction, nasalsecretion/sniffing, post-nasal secretion/snorting)and sign-adjustedscore (mucosacolor andturbinatevolumeincrease,aspectandvolumeofnasal secretion,presenceofalterationsattheoroscopy),graded ona scale goingfrom0to3(absent,mild, moderateand severe),withamaximumscoreof24.3,22,23
The patients underwent skin prick tests,3,24 using the extracts of the most relevant allergens in the city of Curitiba, state of Parana, Brazil,25---27 such as Der-matophagoides pteronyssinus 5.000 BAU (bioequivalent allergen unit)/mL and Lolium perenne (10.000BAU/mL), obtainedfromHollister-Stier®,Spokane,USA.
Thetestwasconsideredpositiveifthemeandiameterof thepapulewas≥3mmwithanerythemahalo,inrelationto thenegativecontrol.3,24
Thepatientsweresubmittedtonasalinstillationof5mL of isotonicsaline solution(0.9%) in both nasal cavities to obtain NL fluid,19---21,28 which washomogenized by shaking
andcentrifugedtoobtainthecellpellet (1000rpm/5min) inarefrigeratedcentrifuge,sothatonly0.01%ofthecells remaininthesupernatant.18
The supernatant was stored at −80◦C for the mea-surementof mediators. The albumin concentrationin the nasal lavage sample was determined by a turbidimetric immunoassay (MicrobuminMULTIGENT --- Abbott Laborato-riesofBrazilLtda®,detectionlimitsof1---500g/mL);the level of interleukin-8 (IL-8) was measured by quantita-tiveenzyme-linkedimmunosorbentassay(ELISAMAXTM Set
Deluxe---Biolegend®,SanDiego,California,detectionlimits of31.2---2000pg/mL).
Thenasalcytologyanalysisoftheinitiallyobtainedpellet (cell pellet) allowed the determinationof the total num-ber of cells/mL, inflammatory and epithelial cell counts; an aliquot was cytocentrifuged (LABHO® CT-12 Cytocen-trifuge)for thepreparationof slidesthat werestainedby theMay---Grünwald---Giemsamethodfordifferentialcounting ofeosinophils,neutrophilsandepithelialcells.18---20,29
The histopathological analysisof mucosa samplesfrom theinferiornasalturbinates,obtainedbypartial turbinec-tomy procedure of the inferior nasal turbinate30 or by mucosal biopsy of the anteroinferior tip of the inferior turbinates,wasevaluatedbylightmicroscopy.21
The anterior portion of the inferior turbinate, more exposedtoairflow andtothegreaterimpactof aeroaller-gensandirritants,ismorerepresentativefortheevaluation of the nasalmucosa inflammatory process31 if the largest amountoftissueandmucusiscollectedtominimizedamage to the sample.30,31 The sample was fixed in formalde-hydeandthetissueblockwasparaffin-embeddedforlater cutting and slide preparation.8,32 The stains used were: Hematoxylin-Eosin (HE) to identify leukocytes, Periodic Acid-Schiff(PAS)tohelpidentifygobletcellsandtrichrome of Gomori tohelp identify andmeasure the subepithelial collagen thickness,according tothe routineof the Anato-mopathologicalServiceofHospitaldeClínicas-UFPR.
The areas of epithelium preferentially covered with mucus were evaluated, excluding fields withevidence of iatrogenicdetachment of theepithelium and presence of extravasatedredblood cells.Aimingtoavoidfalseresults causedbythecuttangency,theimageandpositionofthe cellnucleiwereevaluated.33
ThestagingoftheEpithelialLesion(EL)andthedegree of basement membrane thickening were based on the score proposed by Ponikau et al.32 The epithelial lesion degree staging ranged from 0 to 3 (0: intact epithe-lium, 1:absenceof ciliated cells,2: erosionof theupper layerofcells withintactbasementcelllayer,3:complete epitheliumerosion),whereasthe stagingof thebasement membrane thickening ranged from 0 to 2 (0: basement membrane not visualized; 1: visible basement membrane withthickness≤20m,2:visiblebasementmembranewith thickness>20m).32
Atthestatisticalanalysis,theestimateofthedifference of asymmetric distribution variables was performed by a non-parametric test (Mann---Whitney), whereascategorical variables werecompared by Fisher’sexact test and Pear-son’sChi-squaretest.Aminimumsignificancelevelof5%and aminimumtestpowerof90%wereconsideredforalltests.A receiveroperatingcurve(ROC)wasconstructedtoestimate thecutoffpointwithgreatersensitivityandspecificity.
Uni-Table1 Totalanddifferentialcellcount,albuminandIL-8levelsinnasallavageofatopicandnon-atopicsubjects.
Nasallavage Atopic(n=36) Non-atopic(n=20) p
Totalcellularity 127,000(10×103---6.134×103) 128,000(24×103---682×103) 0.90 Epithelialcells(%) 48(8---98) 76(10---100) 0.07 Eosinophils(%) 3(0---66) 1(0---21) <0.01 Neutrophils(%) 41.5(0---87) 17.5(0---83) 0.24 Mononuclearcells(%) 1(0---12) 1(0---7) 0.96 Albumin(g/mL) 16(5---338) 16.5(5---105) 0.67 IL-8(pg/mL) 80(30---1300) 81.5(30---604) 0.45
Median(limits);Mann---Whitneytest.
variate logistic regression wasperformed toestimate the probabilityofatopyaccordingtotheeosinophilcount.
This study was submitted to and approved by the
HumanResearchEthics CommitteeofHospital deClínicas ofUniversidadeFederaldoParaná(UFPR),registrynumber 755.174/2003-11.
Results
A total of 56 patient samples were analyzed, grouped accordingtothepresenceofatopy.Allergyskintestswere negative in 20/56 (36.0%) and positive in 36/56 (64.0%). Amongtheatopicsubjects,theskinpricktestwaspositive
forD.pteronyssinusin35/36(97.0%)and/orforL.perenne
in 18/36 (50.0%).The test wasexclusively positivefor D.
pteronyssinusin18/36(50%)andforL.perenneinonly1/36
(3%).
Themedianagewas24.5years(14---58years),withequal distributionbygender.Thetotalsymptomscorewashigher in atopic subjects (9 [1---18]) than in nonatopicones (6.5 [0---12])(p=0.01).
Atopicsubjectstendedtoshowahigherfrequencyof pru-ritusandsneezingofmoderatetosevereintensity(47.2%), althoughatthelimitofsignificance(p=0.05).Thefrequency of the moderate to severe nasal obstruction symptom, presentin25/36(69.0%)ofatopicsubjects,wascomparable tononatopicones(p=0.16).
Totalanddifferentialcellcounts,aswellasalbuminand IL-8levels,areshown inTable 1.The totalnumber ofNL cellsandthedifferentialcountofepithelial,neutrophiland mononuclearcellswereequallydistributedbetweenatopic andnonatopicsubjects.Onlythepercentageofeosinophils washigheramongatopicones(p<0.01).
The frequency of cases by ranges of percentage of eosinophilsintheNL(Fig.1)showedthatmostnon-atopic subjects were concentrated within the range of 0%---2%, 14/20(70%) ofthecasesinrelation to13/36(36%)of the atopicones(p=0.01).Ontheotherhand,thefrequencyof caseswitheosinophilcounts>4%amongatopicsubjectswas higher,observed in 16/36(45.0%) of thesepatients, com-paredtoonly2(10%)ofnon-atopicones.
The cutoffpointwiththehighest sensitivityand speci-ficity indexes was 4% of eosinophils in the NL, which differentiated atopic from non-atopic subjects; with this criterion the sensitivity was 44% and the specificity was 90%.Alsowitha4%cutoff,theprobabilityofatopywas80% (p<0.001),increasingto100%witha10%eosinophilcount.
AT x NAT (p=0,01) Atopic (n=36) Non-atopic (n=20) 0 to 2 % 80 70 60 50 40 30 20 36 19 45 70 20 10 10 % > 2 to 4 % > 4%
Figure1 Frequencyofcasesdistributedbypercentageranges ofeosinophilnumbersinnasallavage.Chi-squaretestwith lin-eartrends(p=0.01).
The ciliated columnar pseudostratified epithelium was observed in most samples (Fig. 2). In some samples, the presence in some areas of dysplastic epithelium (n=1), transitional to non-keratinized squamous type (n=7) was observed. The infiltrate in the chorion/lamina propria waspredominantlyofthemononuclear,lymphoplasmacytic type.
The presence of epithelial lesion and basement mem-branethickeningareshowninFig.2.
Amongtheatopicsubjects,theprevalenceofepithelial lesionwas94.4% andtheepitheliallesion prevalencerate was1.4timeshigher(p<0.01).Theprevalenceofbasement membranelesionwas67.0%,indicating,in theatopic sub-jects,thenon-significantchanceofhavingsomedegreeof basementmembranethickening1.2-foldhigherinrelation tonon-atopicones(p=0.40).
The association between the inflammatory process of thenasalcavitylumen(nasallavage)andthenasalmucosa (histopathologicalanalysis) showed that regardless of the atopy,inthepresenceofsomedegreeofepitheliallesion,
HC08 B 8159 HC09 2401 HC09 B 1915 HC09 B 1915 HC08 B 10 1554 HC08 8158 100x 100x 100x 400x
Destaque esinófilos
400x 100x
A
C
D
E
F
B
Figure2 StagingofEpithelialLesion(EL)andBMthickening:(A)ELgrade0(intactepithelium)andBM0(notvisualized),PAS staining;(B)ELgrade1(absenceofciliatedcells)andBM0(notvisualized),HEstaining;(C)ELgrade1(absenceofciliatedcells)and BM1(visibleBM,thickness≤20m),PASstaining;(D)ELgrade1(absenceofciliatedcells)andBM2(visibleBM,thickness>20m), HEstaining;(E) ELgrade2(epithelialerosion,intactbasement celllayer)andBM 1(BM≤20m);(F)Eosinophilinfiltrationis highlighted,HEstaining.
thetotalcellcountofNL(p=0.18);ofeosinophils(p=0.17) andneutrophils(p=0.75);levelsof albumin(p=0.50)and IL-8 was similar (p=0.09) (Figs. 3 and 4). However, only twoatopicsubjectsdidnothave histopathological epithe-liallesion.On theother hand, therewasnodifferencein cellularityandmediators innon-atopic individualswithor without epithelial lesion.Therefore, inthe study sample, the presence of epithelial lesion wasnot associated with changesintheNL.
ThedifferenceineosinophilcountintheNLofatopicand non-atopicsubjects,withoutBMT,wasborderline(p=0.05). Inthe presence of BMT,there wereno differencesin the subgroupsof patientsregardingthenumberof eosinophils intheNL.Ontheotherhand,theBMTwasassociatedwith anincreaseofneutrophilsinthenonatopic(NAT)subjects’ NL(p<0.01).ThenumberofneutrophilsintheNLofatopic (AT)subjectswithoutBMTwashigherthanintheNATones (p=0.04).AnalysisoftheotherNLparametersdidnotshow significantdifferencesinthepresenceofBMT(Figs.3and4).
Discussion
Rhinitisis theconsequence ofan allergicandnon-allergic nasal mucosa inflammatory process, associated with an accumulationofinflammatorycellsinthenasalcavitylumen andstructuralchangesinthenasalmucosa.Quantifyingthe extentanddegreeofepitheliallesion,aswellasthe associ-ationwithdifferent celltypes in thenasal mucosain the different types of rhinitis may help in the assessment of nasalmucosaremodeling.
Studies on the occurrence and extent of remodeling in rhinitis are conflicting, because the nasal mucosa is moreexposedtoexternalstimuli,resultinginsomedegree of adaptive inflammatory process or due to the different remodelingcriteriausedinthestudies.3,5,9,10,14,15,31---33Nasal obstructionisoneofthemostcommonanduncomfortable symptomsofrhinitis,34 anditisanimportantcomplaintin thisgroupofpatients,whetherallergicornon-allergicones. However, cases were selected at the Otorhinolaryngology
Mediana dos valores
(n=7) (n=34) (n=13) (n=12) (n=9) (n=24) (n=11)
(n=2)
NAT NAT NAT
NAT AT AT p=0.05 p=0.17 p=0.13 % 100 90 30 80 70 60 50 20 10 40 p=0.21 p=0.10 p=0.58 AT AT Epithelial lesion 0 Epithelial lesion 1-3 Lesion BM 1-2 Lesion BM 0
Figure3 Eosinophils(%)innasallavageofatopicandnon-atopicsubjectsaccordingtoepitheliallesionsandbasementmembrane thickening.---,medianvalues(limits);Mann---Whitneytest.
outpatient clinic due to the surgical indication of septal deviationcorrectionand/orturbinectomyduetoturbinate hypertrophy,whichare anatomicalchangesthatalso con-tributetonasalobstruction.
The total scoreof the rhinitis scorewas higher in the atopic patients and, for atopy investigation, the extracts usedintheskin pricktestwereselected accordingtothe mostrelevantregionalaeroallergens,25---27witha sensitiza-tionfrequency of97% for D.pteronyssinus and50% for L. perenne. Nasalprovocation testswithallergens could cir-cumvent thebias of including patients withlocal allergic rhinitisorsensitizedtootherallergensinadditiontohouse dustmitesandgrasspolleninthenon-atopicgroup.35
In thisstudy,thesixpatientssensitizedtopollen were alsosensitizedtoD.pteronyssinus andtheirrhinitis score wasnotdifferentinrelationtotheotheratopicsubjects.
The inflammatory process evaluation of the nasal mucosa, through nasal lavage and analysis by quantita-tivecytologyofthenasallavage,allowstheassessmentof thecell infiltrate andof mediators,3,18---20 withthe collec-tion being more representative when carried out in both nostrils,15 whichis whysampleswerecollectedfromboth nostrilsinthepresentstudy.
The cells recovered by the nasal lavage are derived fromepithelial desquamationand increased vascular
per-meabilityof the epithelium, which allowsthe passage of plasmaproteinsandinflammatorycellsintothenasalcavity lumen.15
In this series, IL-4, IL-5 and IFN-␥ levels were unde-tectable,possiblybecausetheamountsofthesemediators in samples collected without the allergenic stimulus of thenasalprovocation test werelabileandminimal. How-ever, IL-4, IL-5 and IL-13 levels may be elevated in the nasalsecretion ofpatients with persistentsevere allergic rhinitis.36
The wide distribution limit of IL-8 in atopic and non-atopicNLsamplesmayhavemaskedanydifferencebetween thegroups.Thisstudyshowednocorrelationbetween albu-min levels and percentage of nasal lavage eosinophils in both groups of patients with rhinitis. This association is describedinnasalprovocationtestswithallergens,where theincreaseofvascularpermeabilityandtheinflammatory cellinfluxis significantlyhigher,37,38 aswell asinpatients withallergicrhinitisandasthma,whoalsoshowextensive involvement by computed tomography assessment of the paranasalsinuses.20
Cellularinfiltrationinthenasalcavitylumen,reflecting epithelialdesquamationandincreasedmucosal permeabil-ity, was similar in this group of patients with allergic or non-allergic rhinitis; only the number of eosinophils
Mediana dos valores (n=7) (n=34) (n=13) (n=12) (n=9) (n=24) (n=11) (n=2) Epithelial lesion 0 AT p=0.75 p=0.04 p=0.56 p=0.80 p=0.01 p=0.45 % 100 90 80 70 60 40 50 30 10 20 AT AT AT
NAT NAT NAT NAT
Epithelial lesion 1-3 Lesion BM 0 Lesion BM 1-2
Figure4 Neutrophils (%)inthe nasallavageofatopicandnon-atopic subjectsaccording toepithelial lesionsand basement membranethickening.---,medianvalues(limits);Mann---Whitneytest.
in the NL differentiated the group of atopic sub-jects.
The increase in the number of nasal eosinophils show a better correlation with the nasal obstruction symptom in patients withpersistent allergicrhinitis.12 In the study caseswithmoderate tosevere nasalobstruction,the pro-portion of eosinophils in the nasal lavage was higher in the atopic group. Eosinophil infiltration is more evident afternasalprovocationtestswithallergens,18,38 but there is a disagreementregarding the cutoffpoint of the num-berofeosinophilsthatisconsideredhighinnasalsecretion samples.
The difficulty in comparingstudy results is due tothe differentmaterialcollectionmethods, either by scraping, blowedsecretionanddifferenttechniquesofnasallavage, aswellasseveralmethodsofstainingandcriteriafornasal eosinophilquantification.15---17Insamplesfromthe quantita-tivenasallavagecytology,the5%eosinophilcutoffresulted inanaccuracyof82%(sensitivity80%andspecificity83%)in thediagnosisofperennialallergicrhinitis.19
Inadults,nasalfluideosinophilcountwas6%inpatients withallergicrhinitisand2%inthosewithnon-allergic rhini-tis,andthebestcutoffpointwasestablishedasbeing4%.39 Inthepresentstudy,consideringthe4%eosinophil cut-offpoint,theprobabilityofatopywas80%andincreasedto
100%witha10%eosinophilcount.The4%eosinophilcutoff point in thenasal lavagehas bettersensitivity and speci-ficityindexeswhendifferentiatingallergicfromnon-allergic rhinitis.
The disadvantage of the nasal lavage technique is the factthattherecoveredcellsoriginateonlyfromthenasal cavity lumen anddo notnecessarily reflect theepithelial tissueinflammatoryprocess,40 inwhichthereticular base-mentmembranethickeningisusedasaparameterforairway remodeling.9
Atopic subjects more frequently had some degree of epithelial lesion, but this finding wasnot associatedwith differences in the inflammatory process in the nasal cavity lumen when compared to non-atopic ones. Amin etal. reportedloss of epithelial integrityassociatedwith increasednumberofeosinophils,butnotofneutrophils,in the nasal mucosa of patients with persistent allergic and non-allergicrhinitis.14
According to the findings by Lim et al., nasal secretion and nasal mucosa represent distinct com-partments with different populations of leukocytes. After provocation tests with allergens, there was no epithelial destruction, basement membrane thickeningor subepithelialcollagendeposition,asdescribedinbronchial asthma.21
Fewstudieshaveinvestigatedthestructuralremodeling inthenasalmucosaofpatientswithallergicrhinitisandthe dataareconflicting.36Thepresenceanddegreeofbasement membrane thickeningmay varyaccording tothe assessed region. In the anterior region of the inferior turbinate, wheretherespiratoryepitheliumpredominates,the propor-tionofbasementmembranethickeningishigher.31Because itismoreexposedtoairflow,wherethereisgreaterimpact ofaeroallergensandirritants,some aspectsofthe inflam-matoryprocessand remodelingfindingsmay representan adaptiveresponseof thenasalmucosa,makingitdifficult tocomparepatientswithoutandwithrhinitis.14,31,33
Epitheliallesion,characterizedbythepresenceof inter-cellular edema, epithelial desquamation and eosinophilic clusters,wasdemonstratedintheanteriornasalmucosaof patients allergic to dust mites.6,14,31 In eight of the sam-plesfromthepresentstudy,areasofdysplasticepithelium andtransitionaltothenon-keratinizedsquamoustypewere observed,alsodescribedbyotherauthors.41,42
Anincreasednumberofeosinophilsinthesubmucosaof patientswithpersistentsevereallergicrhinitisisnotalways accompaniedbychangesinepithelialintegrityand/or base-ment membrane thickening,36 which may be associated withprotectivemechanismsthatminimizeremodelingand potentiateepithelialregeneration.41
However,whenthereisseverenasalobstructiondueto persistentturbinatehypertrophy,thepresenceof subepithe-lial fibrosis has been described, aswell as an association withsymptomchronicityandlittlereversibilityofthenasal airflowrestriction.43 Remodelingwasalsodemonstratedin thenasalmucosaofpatientswithmildandsevereintensity persistent allergic rhinitis,although withrelatively intact epithelium,characterizedbyintensecollagendepositionin thesuperficialandsubmucosallayers,inadditionto signif-icantbasement membranethickening, whichcouldreflect thehighlevelsofTGF-anditspro-fibroticeffect.44
Remodeling in allergic rhinitis, characterized by some degree of basement membrane thickening, without other alterations in the epithelium and submucosa,45 may be relatedto collagendeposition.44,46 The allergic inflamma-tionpotentiatesandacceleratesthisphysiological process of collagen deposition in the basement membrane of the inferiorturbinate.7,47
The frequency of basement membrane thickening was similarbetweenatopic(67%)andnonatopic(55%)subjects inourstudy.Theproportionofeosinophilsandneutrophilsin theNLofatopicandnon-atopicsubjectswassimilaramong thosewithBMthickening.Ontheotherhand,thenumberof neutrophilsinthenasallavageofnon-atopic subjectswith basementmembranethickeningwashigherincomparisonto NATwithoutBMT.Thisfindingcanassociatetheneutrophilic infiltrationtothemucosallesionmechanisminnon-allergic patients.Asimilarnumberofeosinophilsintheatopicand non-atopicNLsamples,withbasementmembrane thicken-ing,mayalsosuggesttheparticipationofeosinophilsinthe mucosallesionofpatientswithallergicornon-allergic rhini-tis.
Eosinophilic infiltration is the main cause of epithelial lesion,associatedwithlossofepithelialintegrityinpatients with allergic or non-allergic rhinitis.6,14,29,33 In chronic neutrophilic and eosinophilic rhinosinusitis, remodeling findingsweresimilar,whichalsocorroboratesthe concept
thattheoccurrenceofmucosalremodelingisindependent ofthetypeofinflammatoryreaction.45
Conclusion
Thesefindingsshowthatinpatientswithrhinitis,allergicor non-allergic,therearesomedifferencesintheinflammatory process,buttheymayresultinsimilarstructuraldamageto the nasalmucosa. If there is an association between the progressionoftheinflammatoryprocessinthenasalcavity lumenandthehistopathologicalmucosallesion(thickening ofthebasementmembrane),thisassociationisindependent ofthepresenceofatopyinpatientswithrhinitis.
Conflicts
of
interest
Theauthorsdeclarenoconflictsofinterest.
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