siRNA基因沉默ASIC1a对过敏性紫癜(HSP)患儿血清IgA1诱导的血管内皮细胞损伤的保护作用  

Silencing of ASIC1a protects the vascular endothelial cells damage induced by IgA1 from Henoch-Schonlein syndrome patients

作　　者：闫波[1] 袁丽萍[2] 张琴[2] 华冉 李玉飞[2] 戴寒晶[1] YAN Bo;YUAN Li-ping;ZHANG Qin;HUA Ran;LI Yu-fei;DAI Han-jing(Department of medical technology, Anhui Medical College, Hefei, Anhui 230000);Department of Pediatrics, First Affiliated Hospital of Anhui Medical University, Hefei, Anhui 230000)

机构地区：[1]安徽医学高等专科学校医学技术系,安徽合肥230000 [2]安徽医科大学第一附属医院儿科,安徽合肥230000

出　　处：《赣南医学院学报》2019年第4期331-336,共6页JOURNAL OF GANNAN MEDICAL UNIVERSITY

基　　金：国家自然科学基金项目(81471617);安徽省高校自然科学重点科研项目(KJ2015A360)

摘　　要：目的:探讨siRNA基因沉默ASIC1a对过敏性紫癜患儿血清IgA1诱导的血管内皮细胞的保护作用。方法:体外培养人正常皮肤微血管内皮细胞(HDMVEC),设计针对人ASIC1a基因编码区的 siRNA 序列,构建重组慢病毒LV-sh-ASIC1a转染给HDMVEC细胞,设立空病毒转染对照组(NC组)、LV-h-ASIC1a转染组(si-ASIC1a组),采用RT-PCR检测转入基因ASIC1a的表达。病毒转染72 h后,加入分离的过敏性紫癜患儿血清IgA1(HSP IgA1)和正常患儿血清IgA1,采用ELISA法检测细胞培养上清中TNF-α、IL-8水平,real-time PCR和 Western blotting 分别检测细胞内ASIC1a、细胞骨架蛋白(SM-α、Destrin、Vinculin、ML-CK)mRNA及蛋白表达水平。结果:与NC对照组比较,si-ASIC1a组HDMVEC内TNF-α、IL-8的分泌显著减少,细胞骨架蛋白SM-α、Destrin、Vinculin、ML-CK mRNA和蛋白表达明显增加,差异有统计学意义(P<0.01)。结论:沉默ASIC1a可以显著抑制HSP血管内皮细胞细胞骨架蛋白的下调,改善血管内皮细胞损伤。Objective: To investigate the protective effect of silencing of ASIC1a on the vascular endothelial cells damage induced by IgA1 from Henoch-Schonlein syndrome(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. The levels of TNF-α and IL-8 in cell culture supernatant were detected by ELISA, ASIC1a and cytoskeleton protein(SM-α, Destrin, Vinculin, ML-CK)mRNA and protein expressions were detected by real-time PCR and Western blotting Methods. Results: TNF-α、IL-8 release in the group of si-ASIC1a group was significantly reduced and cytoskeleton protein(SM-α, Destrin, Vinculin, ML-CK)mRNA and protein expressions compared with the NC control group(P<0.01). Conclusions: Silencing ASIC1a can protect the damage of HSP vascular endothelial cells.

关 键 词：过敏性紫癜 血管内皮细胞 ASIC1a 

分 类 号：R554.6[医药卫生—血液循环系统疾病]