【摘要】 目的:探讨circ_SEC23A通过miR-30a/Beclin-1在心肌细胞损伤中的作用及机制。方法:通过双荧光素酶报告基因实验验证circ_SEC23A和miR-30a的结合情况,分别将circ_SEC23A siRNA、miR-30a mimics、circ_SEC23A siRNA和miR-30a inhibitors、circ_SEC23A siRNA和Beclin-1过表达载体转染心肌细胞后,使用血管紧张素Ⅱ(AngⅡ)诱导心肌细胞损伤,用荧光定量聚合酶链式反应(qPCR)检测circ_SEC23A、miR-30a和Beclin-1的表达,用Western blot检测Beclin-1和LC3-I/II的表达;使用CCK-8法检测细胞增殖活性;使用生化分析仪检测细胞培养上清液中乳酸脱氢酶的浓度以验证细胞损伤程度。结果:circ_SEC23A能靶向结合于miR-30a。AngⅡ诱导心肌细胞损伤后,circ_SEC23A和Beclin-1的表达上调,miR-30a表达下调;下调circ_SEC23A表达后,miR-30a表达增加而Beclin-1表达抑制;LC3-II/LC3-I的比值显著增加,细胞增殖活力下降,同时乳酸脱氢酶浓度增加。结论:circ_SEC23A通过miR-30a/Beclin-1影响自噬,并促使心肌细胞发生损伤。更多还原

【Abstract】 Objective:To explore the mechanisms of circ_SEC23 A in damaging cardiomyocytes through miR-30 a/Beclin-1.Methods:The dual luciferase reporter gene method was used to verify the binding of circ_SEC23 A and miR-30 a.Firstly,the cardiomyocytes were respectively transfected with circ_SEC23 A siRNA,miR-30 a mimics,circ_SEC23 A siRNA and miR-30 a inhibitors,circ_SEC23 A siRNA and Beclin-1 overexpression vector.Then,the angiotensin Ⅱ(Ang Ⅱ) was used to induce the damage of cardiomyocytes.Lastly,the expression of circ_SEC23 A,miR-30 a and Beclin-1 was detected by fluorescent quantitative Polymerase Chain Reaction(PCR) and the expression of Beclin-1 and LC3-I/II was assessed through western blot.Also,the cell proliferation activity was tested by CCK-8 method and the concentration of lactate dehydrogenase(LDH) in cell culture supernatant was evaluated using a biochemical analyzer to verify the degree of cell damage.Results:The circ_SEC23 A could bind to miR-30 a through a specific target site.Besides,Ang Ⅱ significantly promoted the expression of circ_SEC23 A and Beclin-1,while it also inhibited that of miR-30 a.But after down-regulating the expression of circ_SEC23 A,the expression of miR-30 a increased and that of Beclin-1 decreased.Meanwhile,the ratio of LC3-II/LC3-I increased significantly,cell proliferation decreased,and the concentration of LDH increased.Conclusion:The circ_SEC23 A affected autophagy and promoted the damage of cardiomyocytes through miR-30 a/Beclin-1.更多还原