Introduction: Henoch-Schönlein purpura (HSP) is a common kind of systematic vasculitis in children characterized by rash, joint pain,abdominal symptoms and renal disease,the detail pathogenesis of HSP has not been elucidated.Acid-sensing ion channels (ASICs) is a proton-gated sodium selective channel that belongs to Degenerin /epithelial Sodium Channel(DEG/ENaC) superfamily,previous research found the expression of ASIC1a in vascular endothellial cells could be stimulated by serum IgA1 from HSP patients under acidic environment.This study aims to investigate the molecular mechanisms of silencing of acid-sensing ion channel 1a(ASIC1a) protects the vascular endothelial cells from Henoch-Schonlein purpura(HSP) patients.
Methods:Human dermal microvascular endothelial cells ( HDMVEC) were cultured in vitro，siRNA sequences were designed for the coding region of human ASIC1a gene and HDMVEC cells were transfected with recombinant lentivirus( LV) -sh-ASIC1a． The control group ( NC group) without virus transfection and LV-sh-ASIC1a transfection group ( si-ASIC1a group) were set up． The expression of transfixed ASIC1a gene was detected by RT-PCR． After virus transfection 72 h，serum IgA1from HSP patients and serum IgA1 of normal children were added into HDMVEC cells． ASIC1a and cytoskeleton protein ( sm-α f-actin，MLCK) mRNA and protein expressions were detected by real-time PCR and Western blotting Methods.The binding activity of NF- κB with DNA in HDMVEC was determined by electrophoretic mobility shift assay (EMSA).The whole-cell patch-clamp technique was used to record the current changes and electrophysiological characteristics of ASICs and calcium in HDMVEC.
Results：Cytoskeleton protein ( sm-α f-actin，MLCK) mRNA and protein expressions in the group of si-ASIC1a group were significantly increased compared with the NC control group( P ＜ 0． 01) ．The HSP serum and silencing of ASIC1a had no significant effect on the binding activity of NF-κB with DNA in HDMVEC. Compared with the NC control group, the ASICS current and calcium overload of the si-ASIC1A group were reduced( P ＜ 0． 01) ．
Conclusion: Silencing ASIC1a can protect HSP vascular endothelial cell injury by inhibiting ASIC1-related calcium influx and reducing the overload of calcium ,not the NF- κB signaling pathway.