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miR-122-5p通过靶向Circ_MTM1在HBV相关肝纤维化中的分子机制研究
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Abstract:
目的：研究微小核糖核酸microRNA-122-5p (miR-122-5p)对HBV感染相关肝纤维化的作用及分子机制。方法：分别用采用HBsAg、HBeAg酶联免疫吸附剂测定(ELISA)试剂盒和实时荧光定量聚合酶链反应(RT-qPCR)检测HBV表面抗原(HBsAg)、抗原(HBeAg)水平以及HBV DNA、HBV共价闭合环状DNA (cccDNA)水平，并采用蛋白质印迹法(Western blot)检测相应蛋白水平。RT-qPCR检测人肝星状细胞(LX-2细胞)和HBV相关肝纤维化组织细胞中Circ_MTM1、miR-122-5p的表达情况。应用生物信息学软件StarBaseV3.0预测miR-122-5p的可能靶基因，并应用双荧光素酶检测方法判定miR-122-5p对其靶基因Circ_MTM1表达的调节作用。结果：miR-122-5p在HBV相关性肝纤维化组织和细胞中低表达，而Circ_MTM1在HBV相关性肝纤维化组织和细胞中高表达。miR-122-5p靶向下调Circ_MTM1的表达，通过下调Circ_MTM1可抑制HBV相关肝纤维化的进程。结论：miR-122-5p与Circ_MTM1在HBV相关肝纤维化中存在靶向关系，Circ_MTM1通过调控肝纤维化相关蛋白的水平在HBV相关肝纤维化进程中起到启动子作用。
Objective: To study the effect of microRNA-122-5p (miR-122-5p) on infection-related liver fibrosis and its molecular mechanism. Methods: HBsAg and HBeAg enzyme-linked immunosorbent assay (ELISA) kit and real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) were used to detect HBV surface antigen (HBsAg), antigen (HBeAg) levels, HBV DNA and HBV covalent closed circular DNA (cccDNA) levels, respectively. The corresponding protein levels were detected by Western blot. The expression of Circ_MTM1 and miR-122-5p in human hepatic stellate cells (LX-2 cells) and HBV-related hepatic fibrosis tissue was detected by RT-qPCR. Bioinformatics software StarBaseV3.0 was used to predict the possible target genes of miR-122-5p, and dual luciferase assay was used to determine the regulatory effect of miR-122-5p on the expression of its target gene Circ_MTM1. Results: The expression of miR-122-5p was low in HBV-associated liver fibrosis tissues and cells, while the expression of Circ_MTM1 was high in HBV-associated liver fibrosis tissues and cells. The expression of Circ_MTM1 is down-regulated by miR-122-5p, and the process of HBV-related liver fibrosis can be inhibited by down-regulating Circ_MTM1. Conclusion: There is a targeting relationship between miR-122-5p and Circ_MTM1 in HBV-related liver fibrosis, and Circ_MTM1 plays a promoter role in the process of HBV-related liver fibrosis by regulating the level of liver fibrosis-related proteins.
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