The Effect Of MiR-138-5p Mediating Endoglin On Diabetic Myocardial Fibrosis

Objective: To investigate the effect of miR-138-5p targeting endothelial glycoprotein(Endoglin),on myocardial fibrosis in diabetic mice and explore its possible mechanism.Methods: Ten of the 40 male C57BL/6 mice were randomly selected as the normal control group,the other mice were treated with low-dose intraperitoneal injection of STZ(40 mg/kg/d)for 5 consecutive days to establish a model of type 2 diabetes mice.HE staining to observe the morphology and pathological changes of apical tissue in mice;Masson staining to observe the accumulation of collagen fibers in myocardial tissue of mice;qRT-PCR to detect the expression of Collagen ?,Collagen ?,Endoglin and miR-138-5p in myocardial tissue of mice;Western blot to determine the protein expression of Collagen ?,Collagen ? and Endoglin in myocardial tissue of mice.In vitro experiments,primary isolate and culture C57BL/6 mouse rat myocardial fibroblasts(mCFs),and establish a model of myocardial fibrosis by high glucose(30mmol/L)induction of mCFs for 24 h to detect the gene expression of miR-138-5p,Collagen ?,Collagen ? and Endoglin in mouse cardiac fibroblasts and the protein expression of Collagen ?,Collagen ? and Endoglin;Use TargetScan software to predict and analyze the binding miRNA of Endoglin genes and double luciferase reporter gene detection technology to verify the target association between miR-138-5p and Endoglin 3'UTR;Transfection of miR-138-5p mimics and inhibitor in mCFs and induction with high glucose and establish blank and negative control group,and detecte the gene and protein expression of Collagen ? and Endoglin in cells by qRT-PCR and Western blot method.Results:The results of HE staining and Masson staining in mouse heart tissue sections showed that compared with the normal control group,the myocardial tissue arrangement disorder and collagen deposition appeared in the DM group;The results of qRT-PCR and Western blot showed that the mRNA and protein expression levels of Collagen ?,Collagen ? and Endoglin in the heart tissues of DM mice were up-regulated compared with the normal control group,while the miR-138-5p expression was down-regulated(P<0.05).In vitro,after high glucose induced mCFs for 24 h,the mRNA and protein expression of Collagen ? and Endoglin were higher than that of normal control group,the expression of Collagen ? was higher than that of normal group while there was no statistical difference,but the expression of mir-138-5p mRNA was opposite to that of Endoglin group,and the expression of mRNA was down-regulated compared with that of CON group.Using TargetScan software to predict the binding miRNA of Endoglin genes,it was found that the miR-138-5p contained a binding domain of CACCAGCA sequence in human and mouse,which was highly conserved,indicating that Endoglin mRNA could be a target for miR-138-5p;The results of double luciferase reporter gene detection revealed targeted binding between miR-138-5p and the 3'UTR of Endoglin(P<0.05).The expression of Collagen ? and Endoglin decreased in the miR-138-5p mimics group and the opposite in the miR-138-5p inhibitor group after transfected with miR-138-5p mimics and inhibitor in mCFs and induction with high glucose(P<0.05).Conclusion: miR-138-5p may influence mCFs activation and proliferation by targeting down-regulated Endoglin,thereby regulating the course of diabetic myocardial fibrosis.