MiR-9-5p调控瞬时受体电位M7对心肌缺血/再灌注大鼠的保护作用  

Protective effect of miR-9-5p regulating transient receptor potential melastatin 7 on myocardial ischemia-reperfusion in rats

作　　者：刘向前[1] 周仪梦 谢向荣[2] 倪进忠[3] 杨浩 LIU Xiang-qian;ZHOU Yi-meng;XIE Xiang-rong;NI Jin-zhong;YANG Hao(Department of Cardiology,the First People's Hospital of Chuzhou,Anhui Chuzhou 239000,China;Department of Cardiology,Yijishan Hospital of Wannan Medical College,Anhui Wuhu 241001,China;Department of Anatomy,Wannan Medical College,Anhui Wuhu 241002,China)

机构地区：[1]滁州市第一人民医院心血管内科,安徽滁州239000 [2]皖南医学院弋矶山医院心血管内科,安徽芜湖241001 [3]皖南医学院解剖学教研室,安徽芜湖241002

出　　处：《解剖学报》2022年第2期246-253,共8页Acta Anatomica Sinica

摘　　要：目的探讨miR-9-5p调控瞬时受体电位M7(TRPM7)对心肌缺血/再灌注(MIR)大鼠的作用。方法将32只SD大鼠分为假手术组、模型组、miR-9-5p过表达组及空载体对照组,通过左冠状动脉结扎法建立MIR模型,假手术组不结扎,miR-9-5p过表达组和空载体对照组于模型建立前24 h分别尾静脉注入miR-9-5p agomir和agomir NC。通过HE染色观察各组大鼠心肌损伤情况;Real-time PCR检测各组大鼠心肌组织中miR-9-5p表达水平;酶联免疫吸附测定法检测各组大鼠血清中白细胞介素(IL)-6、肿瘤坏死因子α(TNF-α)、IL-1β、肌酸激酶同工酶MB(CK-MB)、心肌肌钙蛋白Ⅰ(cTnI)、乳酸脱氢酶(LDH)水平以及心肌组织中丙二醛(MDA)、超氧化物歧化酶(SOD)含量;TUNEL法检测心肌细胞凋亡情况;双荧光素酶实验验证miR-9-5p与TRPM7的关系;免疫印迹法检测各组大鼠TRPM7、Bcl-2、Bax、磷酸化核因子κB 65(p-NF-κB p65)、toll样受体4(TLR4)蛋白表达。结果MiR-9-5p在大鼠心肌组织中低表达(P<0.05);miR-9-5p过表达能够降低CK-MB、cTnI、LDH表达水平,改善大鼠心肌损伤程度;与模型组相比,miR-9-5p过表达组心肌细胞凋亡率、Bax蛋白表达及MDA、IL-6、TNF-α及IL-1β含量均下降,而Bcl-2蛋白表达和SOD含量上升(P<0.05);双荧光素酶实验结果显示,TRPM7为miR-9-5p的靶基因,且miR-9-5p过表达组TRPM7、p-NF-κB p65、TLR4蛋白表达均较模型组降低(P<0.05)。结论MiR-9-5p通过调控TRPM7抑制心肌缺血再灌注导致的心肌细胞凋亡、氧化应激及炎症反应,并抑制TLR4/NF-κB通路。Objective To investigate the effect of microRNA-9-5p(miR-9-5p)regulating transient receptor potential melastatin 7(TRPM7)on myocardial ischemia-reperfusion(MIR)in rats.Methods Thirty-two SD rats were divided into sham operation group,model group,miR-9-5p overexpression group and empty vector control group.The MIR model was established by ligation of left coronary artery.The sham operation group was not ligated.miR-9-5p agomir and agomir NC were injected into tail vein 24 hours before model establishment in miR-9-5p overexpression group and empty vector control group.The myocardial injury was observed by HE staining.The expression of miR-9-5p was detected by Real-time PCR.The serum levels of interleukin(IL)-6,tumor necrosis factor alpha(TNF-α),IL-1β,creatine kinase isoenzyme MB(CK-MB),cardiac troponinⅠ(cTnI),lactate dehydrogenase(LDH)and the contents of malondialdehyde(MDA)and superoxide dismutase(SOD)in myocardium were measured were measured by ELISA.Cardiomyocyte apoptosis was detected by TUNEL.Double luciferase assay verified the relationship between miR-9-5p and TRPM7.The protein expressions of TRPM7,Bcl-2,Bcl-2 associated X(Bax),phosphorylated nuclear factor kappa-B 65(p-NF-κB p65)and toll like receptor 4(TLR4)were detected by Western blotting.Results The expression of miR-9-5p was low in myocardial tissue of rats(P<0.05).Overexpression of miR-9-5p could reduce the expression levels of CK-MB,cTnI and LDH,and improve the degree of myocardial injury.Compared with the model group,the apoptosis rate,Bax protein expression,MDA,IL-6,TNF-αand IL-1βcontents in myocardial cells of miR-9-5p overexpression group decreased,while Bcl-2 protein expression and SOD content increased(P<0.05).The result of dual luciferase assay showed that TRPM7 was the target gene of miR-9-5p,and the protein expressions of TRPM7,p-NF-κB p65 and TLR4 in miR-9-5p overexpression group were lower than those in model group(P<0.05).Conclusion MiR-9-5p can inhibit myocardial cell apoptosis,oxidative stress and inflammation induced by myocard

关 键 词：微小RNA-9-5p 瞬时受体电位M7 心肌缺血/再灌注 Toll样受体4/核因子κB信号通路 实时定量聚合酶链反应 酶联免疫吸附测定 免疫印迹法 大鼠 

分 类 号：R542.2[医药卫生—心血管疾病]