Role of microRNA-133b-5p in apoptosis induced by hypoxia/reoxygenation in rat cardiomyocytes

Objective To evaluate the role of microRNA-133b-5p (miR-133b-5p) in apoptosis hypoxia/reoxygenation (H/R) induced by in rat cardiomyocytes. Methods Rat myocardial cell line H9c2 was cultured in DMEM/F12 culture medium supplemented with 10% fetal bovine serum. The cells were seeded in 96-well or 6-well plates and randomly divided into 4 groups (n= 64 wells each): control group (group C); group H/R; miR-133b-5p mimic + H/R group (group M+ H/R); miR-133b-5p negative control + H/R group (group NC+ H/R). The cells were exposed to 95% N2-5% CO2 for 5 h at 37 ℃ followed by 1 h reoxygenation in DMEM/F12 culture medium supplemented with 10% fetal bovine serum in all the groups except group C. The cells were cultured in normal culture atmosphere in group C. In M+ H/R and NC+ H/R groups, the cells were transfected with miR-133b-5p mimic (final concentration 30 nmol/L) and miR-133b-5p negative control (final concentration 30 nmol/L), respectively, for 24 h before H/R. Total RNA was extracted from cells to detect the expression of miR-133b-5p using quantitative real-time PCR. The cell viability (by CCK-8) and lactic dehydrogenase (LDH) activity in the culture medium were detected. Cell apoptosis was assessed by Annexin V/PI flow cytometry. Apoptotic rate was calculated. Result Compared with group C, the cell viability was significantly decreased, and the LDH activity and apoptotic rate were increased in H/R, M+ H/R and NC+ H/R groups, the expression of miR-133b-5p was down-regulated in H/R and NC+ H/R groups, and the expression of miR-133b-5p was up-regulated in group M+ H/R. Compared with group H/R, the cell viability was significantly increased, the LDH activity and apoptotic rate were decreased, and the expression of miR-133b-5p was up-regulated in group M+ H/R, and no significant change was found in the parameters mentioned above in group NC+ H/R. Conclusion H/R in rat cardiomyocytes can induce cell apoptosis possibility through down-regulating the expression of miR-133b-5p Key words: MicroRNAs; Myocytes, cardiac; Apoptosis; Myocardial reperfusion injury