下调miR-616-3p对补体攻击骨髓间充质干细胞的影响  

Down-regulation of mi R-616-3p inhibits attacking of complement system on BMSCs by targeting membrane complement regulatory protein CD46

作　　者：周建国[1] 刘如月[1] 韩枫[1] 刘一亮 ZHOU Jian-guo;LIU Ru-yue;HAN Feng;LIU Yi-liang(The First Department of Orthopaedics,The First People's Hospital of Shangqiu City,Shangqiu476000,China;Department of Microsurgery and Hand-Foot Surgery,Nanhua Hospital,Uni-versity of South China,Hengyang,421002,China)

机构地区：[1]商丘市第一人民医院骨一科,河南商丘476000 [2]南华大学附属南华医院手足显微外科,湖南衡阳421002

出　　处：《中国矫形外科杂志》2021年第1期69-74,共6页Orthopedic Journal of China

摘　　要：[目的]探讨miR-616-3p对补体系统攻击骨髓间充质干细胞(BMSCs)的影响及其机制。[方法]从SD大鼠骨髓腔中分离提取并获得原代BMSCs,将miR-616-3p抑制剂转染到rBMSCs中,再采用大鼠颈静脉血分离的异体血清(ARS)进行干预。采用qRT-PCR法检测各组rBMSCs细胞中miR-616-3p的表达水平,Western Blot法检测膜补体调节蛋白CD46和凋亡蛋白cleaved Caspase-3的表达水平,MTT法检测各组rBMSCs细胞增殖活性,流式细胞术检测细胞凋亡率以及补体攻击复合物(MAC)沉积情况,采用双荧光素酶报告系统验证miR-616-3p与CD46的靶向作用关系。[结果]ARS干预能促进rBMSCs中miR-616-3p表达,抑制CD46蛋白表达,并降低rBMSCs增殖活性,促进rBMSCs凋亡以及增加r BMSCs表面MAC沉积。抑制miR-616-3p能促进ARS干预条件下rBMSCs的存活及CD46蛋白表达,抑制r BMSCs凋亡,减少rBMSCs表面MAC沉积。双荧光素酶报告系统实验证实CD46是miR-616-3p靶基因。[结论]下调miR-616-3p通过靶向结合膜补体调节蛋白CD46可以促进BMSCs细胞增殖,抑制细胞凋亡,进而抑制补体系统对rBMSCs的攻击作用。[Objective]To explore the effect of miR-616-3 p on attacking of complement system on bone marrow mesenchymal stem cells(BMSCs)and its mechanism.[Methods]The primary BMSCs were obtained and isolated from the marrow cavity of SD rats,and transfected with miR-616-3 p inhibitor.Then rBMSCs were intervened with the isolated allogeneic rat serum(ARS).The expression of miR-616-3 p in rBMSCs was detected by qRT-PCR assay;the expression of membrane complement regulatory protein CD46 and apoptosis protein cleaved Caspase-3 were detected by using Western Blot;the proliferation activity was detected by MTT assay;and the apoptosis rate and deposition of membrane attact complex(MAC)of rBMSCs were assessed by flow cytometry.Then the relationship between miR-616-3 p and CD46 was verified by dual luciferase report system.[Results]ARS intervention promoted the expression of miR-616-3 p,inhibited the expression of CD46 protein,decreased the proliferation activity of rBMSCs,promoted the apoptosis rate and MAC deposition.However,inhibition of miR-616-3 p expression promoted the survival of rBMSCs and the expression of CD46 protein,inhibited the apoptosis and reduce the deposition of MAC in the rBMSCs intervened with ARS.Dual Luciferase Report System confirmed that CD46 was the target gene of miR-616-3 p.[Conclusion]Down-regulation of miR-616-3 p does promote BMSCs proliferation,inhibit cell apoptosis and suppress the attack of complement system on rBMSCs by targeting CD46 expression.

关 键 词：大鼠骨髓间充质干细胞 补体系统 miR-616-3p 膜补体调节蛋白 靶向结合 

分 类 号：R318[医药卫生—生物医学工程]