MiR-103a通过TAK1介导的NF-κB信号通路保护炎症诱导的心肌细胞损伤的研究  

MiR-103a protecting cardiomyocyte damage induced by inflammation through TAK1-mediated nuclear factor-κB signaling pathway

作　　者：李璐斐 金培印[1] 王磊[1] Li Lufei;Jin Peiyin;Wang Lei(Department of Cardiovascular Medicine,People's Hospital of Anyang City,Anyang 455000,China.;不详)

机构地区：[1]安阳市人民医院心血管内科,安阳455000

出　　处：《中国循证心血管医学杂志》2023年第2期193-197,201,共6页Chinese Journal of Evidence-Based Cardiovascular Medicine

基　　金：河南省医学科技攻关计划项目(2020022516)。

摘　　要：目的研究miR-103a表达对炎症诱导的H9C2心肌细胞凋亡的影响,并探讨其分子机制与转化生长因子β活化激酶1(TAK1)及其介导的NF-κB信号通路的关系。方法通过肿瘤坏死因子-α(TNF-α)处理H9C2细胞建立炎症诱导的H9C2心肌细胞凋亡体外模型。将不同处理的H9C2细胞分为对照组、TNF-α组、TNF-α+miR-nc组、TNF-α+miR-103a组、TNF-α+si-nc组、TNF-α+si-TAK1组、miR-nc+AAV-nc组和miR-103a+AAV-TAK1组。对照组不进行任何处理,其他各组(除TNF-α组外)细胞均进行转染,并经20μg/mL TNF-α处理24 h,使用流式细胞仪检测细胞凋亡和CCK-8试剂盒检测细胞活性。通过caspase 3、8和9试剂盒测定凋亡相关蛋白caspase 3、8和9蛋白表达。qRT-PCR检测miR-103a表达,免疫印迹检测TAK1、NF-κB和p-NF-κB蛋白表达。结果与对照组比较,TNF-α组H9C2细胞中miR-103a表达显著降低,TAK1蛋白表达显著升高。过表达miR-103a或敲低TAK1均可显著下调TNF-α诱导的H9C2细胞凋亡(P<0.05),并显著下调caspase 3、8和9蛋白表达(P<0.05)。双荧光素酶基因报告系统检测结果显示:miR-103a靶向抑制H9C2细胞中TAK1表达。过表达miR-103a或下调TAK1均显著降低NF-κB的磷酸化(P<0.05),而过表达TAK1显著降低miR-103a过表达对TNF-α诱导的H9C2细胞凋亡的抑制作用和对NF-κB的磷酸化的抑制作用。结论TNF-α处理H9C2心肌细胞下调miR-103a和上调TAK1蛋白表达,miR-103a可通过靶向抑制TAK1而抑制NF-κB信号通路的激活,最终抑制TNF-α诱导的H9C2心肌细胞凋亡。Objective To study the influence of miR-103a expression on cardiomyocyte apoptosis induced by inflammation,and to discuss the relationship among which molecular mechanism,transforming growth factor-β-activated kinase(TAK1)and nuclear factor-κB(NF-κB)signaling pathway.Methods The in vitro model of inflammation-induced H9C2 cardiomyocyte apoptosis was established through treating H9C2 cells with tumor necrosis factor-α(TNF-α).All H9C2 cells with treated with different treatments were divided into control group,TNF-αgroup,TNF-α+miR-nc group,TNF-α+miR-103a group,TNF-α+si-nc group,TNF-α+si-TAK1 group,miR-nc+AAV-nc group and miR-103a+AAV-TAK1 group.The control group was not given any treatment.The cells in other groups(except of TNF-αgroup)were transfected and treated with TNF-α(20μg/mL)for 24 h.The protein expressions of apoptosis-related proteins,including caspase-3,caspase-8 and caspase-9 were detected by using kits of caspase-3,caspase-8 and caspase-9.The expression of miR-103a was detected by using qRT-PCR.The protein expressions of TAK1,NF-κB and p-NF-κB were detected by using Western blotting assay.Results Compared with control group,the expression of miR-103a in H9C2 cells decreased significantly,and expression of TAK1 protein increased significantly in TNF-αgroup.The overexpression of miR-103a or knockdown of TAK1 down-regulated significantly H9C2 apoptosis induced by TNF-α(P<0.05),and down�regulated significantly the protein expressions of caspase-3,caspase-8 and caspase-9(P<0.05).The testing results of dual luciferase gene reporter system showed that miR-103a inhibited targeted TAK1 expression in H9C2 cells.The overexpression of miR-103a or knockdown of TAK1 reduced significantly NF-κB phosphorylation(P<0.05).The overexpression of TAK1 reduced significantly the inhibitory effects of miR-103a overexpression on TNF-α-induced H9C2 apoptosis and NF-κB phosphorylation.Conclusion The treatment of H9C2 cardiomyocytes with TNF-αdown-regulates miR-103a expression and up-regulates TAK1 protein e

关 键 词：miR-103a 肿瘤坏死因子-Α 凋亡 NF-ΚB通路 

分 类 号：R322.11[医药卫生—人体解剖和组织胚胎学]