The Role Of MiR-367 In Promoting The Proliferation Of Mouse Cardiomyocytes

Objective:To compare the differences of the members of miR-302/367 family in promoting the proliferation of HL-1(mouse atrial cardiomyocyte).Further,apply the screened miRNAs with the strongest HL-1 proliferation effect to the mouse myocardial infarction(MI)model to evaluate its proliferation promoting ability and the recovery of cardiac function in vivo.Methods:In this study,the experiments were mainly divided into two parts:in vitro and in vivo.In in vitro experiments,the mimic and negative control(NC)of each member of the miR-302/367 family(miR-302a/b/c/d and miR-367)were respectively transfected into HL-1 by using the transfection reagent Lipofectamine RNAiMAX(RNAiMAX),and then the RNA of each group was extracted for real-time quantitative PCR(qPCR)to detect the expression of miRNA in HL-1 cells.Then on the 1st,3rd and 5th days after transfection,the changes of cell proliferation ability after miRNA overexpression were detected by CCK-8 method.Finally,EdU experiment was used to evaluate whether there was any difference in the ability of each miRNA to promote HL1 cell proliferation.The miRNA having the strongest proliferation effect was screened and used for in vivo experiments.The myocardial infarction model was constructed by permanent ligation of the anterior descending coronary artery in mouse,and and then the miRNA mimic with the strongest proliferative ability was screened out by intramyocardial injection in the border zone of myocardial infarction.Before the operation and on the 3rd,7th and 14th days after the operation,the heart function of the mouse was measured by small animal ultrasound.After the 14th day,myocardial infarction tissue was taken out for paraffin embedding and sectioning and histological analysis.The Ki-67 and Masson staining experiment to analyze the proliferation of mouse cardiomyocytes and the area of myocardial fibrosis after myocardial infarction,further verifying the effect of miRNA on promoting myocardial proliferation and improving heart function in vivo.Results:In vitro experiments,it was proved that RNAiMAX has a good transfection effect and the overexpression of miR-302a/b/c/d and miR-367 can promote the proliferation of HL-1 cells,and there was no significant difference in the proliferation ability of each member.The results of in vivo studies show that the mouse model of myocardial infarction can be successfully constructed by ligating the anterior descending coronary artery in mice.And on the 14th day after myocardial infarction in mouse,it was found by cardiac ultrasound that the heart function of mouse treated with miR-367 mimic was to some extent improved.In paraffin sections,it was observed that the proportion of Ki-67+cardiomyocytes in the miR-367 mimic group was highest than that in other experimental groups,and the area of fibrosis after myocardial infarction was smallest.Conclusion:In conclusion,the overexpression of miR-302/367 family in HL-1 cells can promote the proliferation of these cells and there was no statistical difference in the ability of miR-302a/b/c/d and miR-367 to promote cardiomyocyte proliferation.Intramyocardial injection of miR-367 mimic can promote cardiomyocyte proliferation and reduce myocardial fibrosis area in mice after myocardial infarction,thereby improving heart function.