Protective Effect Of Ginsenoside Rg1 On Myocardial Ischemia-Reperfusion Injury And The Regulatory Mechanism Of MiR-34a

Objective: Serious myocardial ischemia/reperfusion(I/R)injury could cause heart failure.How to reduce myocardial ischemia-reperfusion injury is currently an urgent need to be solved.GinsenosideRgl has many pharmacological functions.However,whether it can reduce myocardial ischemia/ reperfusion injury has not yet been clarified.MicroRNA(miRNA)is crucial in the pathology and physiology process of cell.Inhibition of miR-34 a may improve cardiac function in patients with heart failure.Whether miR-34 a inhibition can reduce myocardial ischemia-reperfusion injury is still unclear.This study will investigate whether ginsenosideRgl has the protective effect on myocardial ischemia-reperfusion injury and explore its underlying mechanism,as well as explore whether ginsenosideRgl can mediate change of miR-34 a and the regulatory mechanisms of miR-34 a during myocardial ischemia-reperfusion injury.Methods: This study is executed in vitro and in vivo.In vitro: hypoxia/reoxygenation(H/R)model of H9c2 cells was established and divided into different treatment groups.The activity of cardiomyocytes was detected by CCK8.The proliferation of H9c2 cells was detected by Brd U.The morphology of H9c2 cells was observed.The activity of caspase-3,the activity of antioxidant enzyme and the changes of reactive oxygen species(ROS)and mitochondrial membrane potential(MMP)were determined.Western blot was used to detect the protein expression.Cardiomyocyte apoptosis andROS levels were detected by flow cytometry.In vivo experiments: we firstly established myocardial ischemia-reperfusion(I/R)model in adult male Sprague Dawley(SD)rats and were given different treatments.Left ventricular ejection fraction was assessed by echocardiography and heart was stained with Evans blue and triphenyl tetrazolium chloride(TTC)used for calculating relative myocardial infarction area.Cardiac blood samples were collected to measure the level of serum myocardial enzymes,and the contents of malondialdehyde(MDA)in the tissues were detected by kit.Cell apoptosis was detected by TUNEL assay and protein expression was detected by western blot.Results:(1)H9c2 cardiomyocytes exposed to H/R stimulation showed obvious injury.Ginsenoside Rg1 could promote Nrf2 nuclear transcription and HO-1 protein expression to resist oxidative stress,which decreased cell injury.Nrf2-siRNA and HO-1 inhibitors prevented cardiomyocytes protection of Rg1.Ginsenoside Rg1 can also reduce cell injury by inhibiting JNK pathway.(2)In the cardiomyocytes exposed to hypoxia/reoxygenation model,the level of miR-34 a was elevated.Ginsenoside Rg1 treatment decreased the level of miR-34 a.MiR-34 a mimic(overexpression)can prevent the protective effect of ginsenoside Rg1 on cardiomyocytes.(3)MiR-34 a level was elevated in ischemia-reperfusion myocardium of rats.The adenovirus injected into myocardium that inhibits the expression of miR-34 a,can alleviate myocardial ischemia-reperfusion injury.MiR-34 a inhibitor decreased H/R-induced cell injury in H9c2 cells.In contrast,miR-34 a mimic exacerbated cell injury.Dual luciferase reporter verified Bcl-2 and SIRT1 were the target genes of miR-34 a.Bcl-2-siRNA and SIRT1-siRNA prevented the protective effect of miR-34 a inhibitor on cardiomyocytes.Conclusion:(1)Ginsenoside Rg1 can relieve hypoxia/reoxygenation induced cardiomyocyte injury by resisting oxidative stress.(2)Ginsenoside Rg1 resisted hypoxia/reoxygenation induced cardiomyocyte injury by reducing miR-34 a level.(3)Inhibition of miR-34 a alleviated cardiac ischemia/reperfusion injury.Bcl-2 and SIRT1 were the target genes of miR-34 a.