Effect of C3G gene on apoptosis and proliferation of H9C2 cardiomyocytes / 解放军医学杂志

ABSTRACT

ObjectiveTo study the effect of C3G gene on apoptosis and proliferation of H9C2 cardiomyocytes and its mechanism. MethodsThe RNA lentivirus was constructed, and pCXN2-Flag plasmid (empty plasmid) and pCXN2-Flag-hC3G plasmid (over-expressed human C3G mRNA) were purchased. H9C2 cardiomyocytes were respectively infected and transfected with blank reagent, negative lentivirus, C3G siRNA lentivirus, C3G siRNA lentivirus+pCXN2-Flag plasmid and C3G siRNA lentivirus+pCXN2-Flag-hC3G plasmid with stochastic method. Thus the experiments were randomly divided into five groups, namely blank group, negative control group, C3G silence group, C3G silence+empty plasmid group, and C3G silence+C3G overexpression group. Seventy two hours after plasmid transfection, the expression of C3G mRNA was detected by PT-RCR, cell apoptosis was determined by flow cytometry, cell proliferation rate was determined by MTT, and the protein levels of C3G, p-ERK1/2, Bax and Flag were determined by Western blotting. Results As screened by puromycin, it was found that more than 85% of the cells in negative control group and C3G silence group were labeled with green fluorescent protein, showing that more than 85% of the cells were infected with lentivirus. Compared with blank group and negative control group, the expressed Bax protein and cell apoptosis rate were increased significantly (P<0.01, P<0.05), and the expression levels of C3G mRNA, C3G protein and p-ERK1/2 protein and the cell proliferation rate were remarkably decreased in C3G silence group and C3G silence+empty plasmid group (P<0.01, P<0.05); Compared with C3G silence group and C3G silence+empty plasmid group, the expression level of Bax protein and the cell apoptosis rate were decreased obviously (P<0.01, P<0.05), while the expression levels of C3G mRNA, C3G protein and p-ERK1/2 protein and the cell proliferation rate were remarkably increased in C3G silence+C3G overexpression group (P<0.01, P<0.05). Conclusion C3G gene silence can induce apoptosis and inhibit proliferation in H9C2 cardiomyocytes, and overexpression of C3G gene can reverse the effects of C3G gene silence affecting in H9C2 cardiomyocytes, characterized by a reduction of apoptosis rate and promotion of proliferation, and they may be related to p-ERK1/2 protein and pro-apoptotic molecule Bax.