Effects And Mechanisms Of IgA1Isolated From Henoch-schonlein Purpura Patients On Apoptosis In Human Umbilical Vein Endothelial Cells

Objective Observe the effect of IgA1which from Henoch-Sch nlein purpura patientssera on apoptosis in Human umbilical venous endothelial cells(HUVECs) andexpression of P53Bax Bcl-2and Caspase-3to explore possible mechanism and offerthe experimental evidences for its clinical application.Methods1.Ten children with HSP, admitted in the first Affiliated Hospital of Anhui MedicalUniversity from2010Sep to2011Jan, were enrolled in the Study. Serum sampleswere obtained from peripheral vein of all subjects in the morning. Serum IgAl levelswas measured by using enzyme linked immunosorbent assay (ELISA). Serum IgA1was purified by jacalin affinity chromatography.2.HUVECs were cultured in3different conditional media without fetal calf serum,①HSP group: cultured with0.05mg/ml、0.1mg/ml、0.25mg/ml IgA1from HSPpatients;②normal group: cultured with0.05mg/ml、0.1mg/ml、0.25mg/ml IgA1from health chlidren;③control group: cultured with RPMI-l640.3. Morphology change of HUVECs treated with IgA1observed by optical microscopy.Rates of apoptosis in HUVEC cells at different concentration and different times afterinfection with IgA1were by flow cytometry and TUNEL.4.The expression of P53、Bax、Bcl-2、Caspase-3were assayed by RT-RCR andWestern-blot. Results1. IgA1inhibits HUVEC growth. After infection with IgA124h, optical microscopyobserved that the cells smaller and rounded, detached with surrounding cells. Thechange in HSP group is more obviously.2.The apoptotic rate of normal group and HSP group after infection with IgA1atconcentrations of0.25mg/ml (detection by FCM) and0.1mg/ml (detection by TUNEL)12h was significantly higher than that in control group(P＜0.01), but no significantlydifferences between HSP group and normal group(P＞0.05). The apoptotic rate ofnormal group and HSP group after infection with IgA1at concentrations of0.1mg/ml24h was significantly higher than that in control group(P＜0.01), and havesignificantly differences between HSP group and normal group(P＜0.01). Theapoptotic rate was dose-and time-dependence(P＜0.01).3.The results showed that the expression level of P53、Bax、Caspase-3mRNA werehigher and the expression of Bcl-2mRNA were lower after IgA1at concentrations of0.25mg/ml treated HUVECs24h in HSP group than normal group and controlgroup(P＜0.05).4.The results showed that the expression level of P53、Bax、Caspase-3protein werehigher and the expression of Bcl-2protein were lower after IgA1at concentrations of0.25mg/ml treated HUVECs24h in HSP group than normal group and controlgroup(P＜0.01).Conclusion1.IgA1could induce apoptosis on HUVECs. The apoptotic rate was dose-andtime-dependence. At same time and concentrations the effect of apoptosis by IgA1from HSP patients was strongest among3groups. 2. The possible mechanism may be associated with up-regulated P53、Bax、Caspase-3and down-regulated of Bcl-2expression.