The Effect Of Cyslts On Proliferation And Apoptosis Of T Cell Of Adenoid In Children With Osahs And Its Mechanism

PART ⅠTHE CLINICAL EFFECTS OF MONTELUKAST ANDINTRANASAL CORTICOSTEROIDS IN CHILDRENOSAHSObjective: Analysis clinical features in children with OSAHS inhospital. Comparative analysis the clinical effects of a cysteinyl LT (cysLT)receptor1antagonist (montelukast) and intranasal corticosteroid (ICS).Discuss the feasibility of non-surgical treatment in children with mildOSAHS.Methods:(1) Retrospective analysis the data of children withOSAHS at department of otorhinolaryngology in CHCMU from January2011to September2013. The gender, age, diseases history, characteristic oftonsil and adenoid, AHI and nadir SpO2were analyzed;(2) One hundredand eight-nine patients with mild OSAHS were divided into3groups (thegroup A, the group B and the group C) randomly from October2011toOctober2012.(3) Therapeutic schedule:63were treated with oralmontelukast (OM) in group A for12weeks.63were treated with ICS ingroup B for12weeks.63were treated with a combination of OM and ICSin group C for12weeks.(4) Clinic symptoms and polysomnographicchanges in polysomnographic findings were followed with telephoneinquiring.(5) Contrastive analysis was applied to the analysis of improvement of symptoms and effective rate.(6) The group A was devidedinto the group A1(effective therapy group)and the group A2(invalidtherapy group). The differences of clinical features of the two groups wereanalyzed.Results:(1) There was a peak prevalence at3-6years of age inChildren with OSAHS in hospital. The boys were more than girls inpatients. Though the children with moderate OSAHS were the most in allchildren, Children with mild occupiesed thirty-two point three percent.Some patients had the history of allergic disease such as allergichinitis(11.7%);(2) The clinical symptoms and PSG monitoring resultswere obviously improved posttreatment. The clinical symptoms includedsnoring, mouth breathing, restlessness, and sleep apnea;(3) The symptomsdisappeared more quickly in group C than group A or B. The total effectiverate in group C was higher than that of group A or B. significant differencewas not found between Group A and group B.(4) Tonsil size grading ingroup A2was obviously higher than that of group A1.Conclusion:(1) The children with mild OSAHS occupied theproportion about one-third in all hospitalized children with OSAHS.(2)The children with mild OSAHS may treated with OM and/or ICS inperioperative period. Non-surgical treatment can reduce the size of adenoidand Relieve part of clinical symptoms effectively. The total effective rate ofdrug combination is higher than that of single medication.The result showthat the inflammatory response may be involved in the pathologicalmechanism of airway lymphoid tissue hypertrophy.(3) The therapeuticeffect of the children with the larger tonsils is not good. PART ⅡTHE EFFECT OF LTD4ON THE PROLIFERATIONAND APOPTOSIS OF T CELL IN ADENOIDObjective: To observe CysLTR1expression in the adenoid inchildren with OSAHS, and investigate the effect of CysLTs on theproliferation and apoptosis of CD4+T and CD8+T cell in adenoid.Methods:(1) Collect the adenoid of the children undergoingAdenoidectomy.(2) The specimen were devided into3groups according tothe AHI (the control group: AHI＜5, the mild group:5＜AHI＜10and themoderate-to severe group:AHI＞10）. AdMC were separated and culturedusing96-well plates to exposed to ConA for48hours. The cell culturesupernatants were collected. The levels of LTC4in the supernatants of allgroups were determined using a human LTC4ELISA kit following themanufacturer’s instructions.(3) Adenoid were prepared for paraffinsections.Immunohistochemistry detected the expression of CysLTR1andImmunofluorescence assay located CysLTR.(4) AdMC were separated andrandomized to4groups: group A, control group; group B, PHA; group C,PHA+LTD4; and group D, PHA+LTD4+montelukast. FCM assay was usedto detect the CD4+T and CD8+T cell proliferation.(5) AdMC wererandomized to4groups: group A, control group; group B, DEX; group C,DEX+LTD4; and group D, DEX+LTD4+montelukast. FCM assay wasused to detect the CD4+T and CD8+T cell apopsis.Results:(1) Adenoid of children with OSAHS but not of controlsubjects have enhanced expression of CysLTR1. Part of CysLTR1immunoreactivity was detected in adenoid CD3+T cells.(2) moderate tosevere OSAHS show higher LTC4compared to mild OSAHS and controlsubjects.(3) The rate of increase of CD4+T and CD8+T cell in PHA+LTD4 group is higher than PHA group. Montelukast can inhibit the proliferationeffect.(4) The early apoptosis rate of CD4+T and CD8+T cell of DEX+LTD4group is lower than DEX group.Montelukast can inhibit the apopsiseffect.Conclusions:(1) Expression of LTC4is related to OSAHS severityin children.(2) The expression of CysLTR1enhanced markedly in adenoidchildren with OSAHS. Only part of CysLTR1immunoreactivity wasdetected in adenoid CD3+T cells. The result shows that CysLTR1may maybe related with adenoid hypertrophy and participate in the pathogenesis ofOSAHS in children.(3) LTD4can regulate the proliferation and apoptosis ofCD4+T and CD8+T cell in AdMC stimulated by PHA. PART ⅢMECHANISM OF EFFECT LTD4ON ROLIFERATION ANDAPOPTOSIS OF T CELL IN ADENOIDObjective: To explore whether LTD4by MAPKs pathway regulatedthe proliferation and apoptosis of CD4+T and CD8+T cells in AdMC.Methods:(1) AdMC were randomly assigned to2groups: group A,control group; group B: PHA10ug/ml+LTD41×10-4mmol/l. The cellswere grown in an incubator (37℃，5%CO2) and collected according to thetraining time for5min and15min,30min,1h,2h,12h and extracted theprotein.(2) Western Blot detected the expressions of p-p38、p-ERK1/2andp-JNK protein in ADMC treated with LTD4for each point in time.(3)AdMC were randomized to4groups: group A, control group; group B, PHA;group C, PHA+LTD4; and group D, PHA+LTD4+SB203580or PHA+LTD4+PD98059. FCM assay was used to detect change theproliferation of the CD4+T Cell and CD8+T cell.(4) AdMC wererandomized to4groups: group A, control group; group B, DEX; group C,DEX+LTD4; and group D, DEX+LTD4+SB203580orPHA+LTD4+PD98059. FCM assay was used to detect change the apoptosisof the CD4+T Cell and CD8+T cell.Results:(1) p-p38protein increased obviously after LTD4treated for5min （P＜0.05), and continued to the highest level for1hour （P＜0.001). The p38total protein expression had no obvious change. Thephosphorylation reaction was inhibited by the CysLTR1inhibitormontelukast or a selective p38MAP kinase inhibitor SB203580.(2)p-ERK1/2protein increased obviously after LTD4treated for15min（P＜0.01), and continued to the highest level for1hour（P＜0.001). Theexpression was down to the basic level for2hours. The ERK1/2totalprotein expression had no obvious change. The aforementioned three kindsof inhibitor can restrain the reaction also.(3) Obvious effect on p-JNK byLTD4were not observed.(4) The proliferation and apoptosis of CD4+T andCD8+T cell in adenoid mononuclear cells stimulated by PHA and LTD4were restrained by PD98059.Conclusion:(1) ERK1/2and p38MAPK pathway but no JNK wereactivated by LTD4.(2) LTD4promoted the proliferative effect of CD4+Tand CD8+T cell stimulated with PHA through ERK1/2pathway.(3)Effect of activated p38MAPK pathway by LTD4was not watched.