Inhibition of microRNA-182 reduces hypoxia/re-oxygenation-induced HL-1 cardiomyocyte apoptosis by targeting the nuclear respiratory factor-1/mitochondrial transcription factor A (NRF-1/mtTFA) pathway

Apoptosis is a component of cardiac remodeling following myocardial ischemia and infarction. MicroRNAs (miRNAs) are involved in regulating a range of cellular biological processes, including apoptosis, by regulating gene expression at the post-transcriptional level. Nuclear respiratory factor-1 (NRF-1) is a transcriptional activator of nuclear genes that encode a range of mitochondrial proteins that sensitize cells to apoptosis. The HL-1 mouse cardiomyocyte hypoxia/re-oxygenation (H/R) in vitro model included a 24 h period of hypoxia followed by a 48 h period of re-oxygenation. qRT-PCR was used to study gene expression at the mRNA level. MicroRNA target identification was done using bioinformatics data with analysis using a luciferase dual reporter assay. SiRNA-NRF-1 was used to inhibit NRF1 gene expression. Functional studies included the assessment of cardiomyocyte apoptosis and lactate dehydrogenase (LDH) levels. The protein levels of mitochondrial transcription factor A (mtTFA) and NRF-1 were measured using western blots. The results of this study showed that increasing the duration of HL-1 cardiomyocyte H/R resulted in increased expression of miRNA-182 and a reduction in the expression of NRF-1. Inhibition of miRNA-182 reduced HL-1 cardiomyocyte apoptosis and cell death, increased NRF-1 gene expression and protein levels. These effects were reduced with co-transfection of HL-1 cardiomyocytes using siRNA-NRF-1. A role for miRNA-182 in H/Rinduced cardiomyocyte apoptosis involves regulation of the NRF-1/mtTFA pathway. Future studies may determine whether miRNA-182 knocked down can reduce reperfusion injury associated with myocardial ischemia and infarction.