Study On The Protective Effect Of Inhibiting MEG3 On Chronic Heart Failure

BackgroundChronic heart failure（CHF）is the terminal stage of cardiovascular disease with the highest recurrence rate,disability rate and mortality rate among chronic diseases.Its pathogenesis is not fully understood,and clinical prevention and treatment are not effective,which seriously endangers human health.The occurrence and development of CHF may involve changes in many genes,proteins and signal transduction pathways,which may hinder the clinical prevention and treatment of CHF.In recent years,long-non coding RNA（Lnc RNA）has become a research hotspot in the prevention and treatment of many diseases.Some studies have shown that inhibition of MEG3（Maternally expressed gene 3,Lnc RNA）may have a protective effect on myocardial fibrosis and cardiac systolic and diastolic dysfunction,but the research on the role and mechanism of CHF is very inadequate,and further research is needed.Euparin is a monomer compound with antioxidant effect.Our previous study found that it can improve the heart function of CHF mice and reduce the expression level of MEG3 in the myocardial tissue of mice,which has the value of further research.This study can provide experimental and theoretical basis for discovering new mechanisms of CHF and potential new drugs for prevention and treatment of CHF.ObjectivesThe protective effect and mechanism of MEG3 inhibition and Euparin intervention on CHF were discussed by constructing mouse model and myocardial cell oxidative damage model.Methods1.The mouse model of CHF was established by using Isoprenaline,and the oxidative damage model was constructed by stimulating H9C2 cardiomyocytes in rat with Hydrogen peroxide（H₂O₂）.2.Echocardiography was used to detect the cardiac function indexes of mice,and cardiac weight/body weight was used to detect the changes of cardiac mass index.3.The pathological changes of cardiac myocyte hypertrophy and fibrosis were detected by HE staining and Masson staining.4.Detection of reactive oxygen species（ROS）levels in H9C2 cells by superoxide anion fluorescent probe kit.5.CCK8 kit was used to detect the survival rate of H9C2 cell injury induced by H₂O₂,and flow cytometry was used to detect the apoptosis rate of H9C2 cells induced by H₂O₂.6.Detection of MEG3/mi RNA-129-5p gene levels in mouse left ventricular tissue and H9C2 cells by real-time fluorescent quantitative PCR（q PCR）technology.7.The expression of NPPA、NPPB、ATG14、Akt、p-Akt、GSK3β、p-GSK3β、Bcl2/Bax、m TOR、p-m TOR、Beclin-1、LC3Ⅱ/LC3Ⅰ、P62 protein in left ventricular tissue and H9C2cardiomyocytes was detected by Western blot technique.Results1.Both MEG3 inhibition and Euparin can reduce the heart mass index of CHF mice,and improve the heart function,cardiomyocyte hypertrophy and myocardial fibrosis of the mice.2.MEG3 inhibition and Euparin can increase the survival rate of H9C2 cells stimulated by H₂O₂,and reduce the apoptosis rate and oxidative stress level of cells.3.MEG3 expression increased and target gene expression decreased in CHF animal and oxidative damage cell models.MEG3 inhibition and Euparin therapy can reverse this result.4.MEG3 inhibition and Euparin treatment can reverse the increase in the expression of the apoptosis-related molecular marker Bax protein and the decrease in the expression of p-GSK3βand Bcl2 in the left ventricle of CHF mice and oxidatively damaged H9C2 cells.5.MEG3 inhibition and Euparin treatment reduced the expression of Beclin-1 and LC3Ⅱ/LC3Ⅰin the left ventricle and oxidatively damaged H9C2 cells of CHF mice,and increased the expression levels of p-m TOR and p62 autophagy-related proteins.ConclusionMEG3 inhibition and Euparin can improve the cardiac function of CHF in mice,reduce the hypertrophy and fibrosis of cardiomyocytes,reduce the autophagy,apoptosis and oxidative stress state of cardiomyocytes,thereby having a protective effect on the ventricular remodeling of CHF.Their mechanism of action mainly involves their regulation of mi RNA-129-5p/ATG14/Akt signaling pathway and its downstream related autophagy and apoptosis signaling molecules.