Effect of hypoxia-inducible factor 1-alpha on hypoxia/reoxygenation-induced apoptosis in primary neonatal rat cardiomyocytes.

We studied the role of hypoxia-inducible factor 1-alpha (HIF-1α) in hypoxia/reoxygenation (H/R)-induced apoptosis in primary neonatal rat cardiomyocytes and its possible molecular mechanisms. Isolated neonatal and adult rat cardiac myocytes were cultured for 48h and were submitted to 5h of hypoxia followed by 2, 6, or 12h of reoxygenation. Small interfering RNA was used to target the HIF-1α gene. Cardiac myocyte apoptosis induced by H/R was assessed by Annexin V-FITC apoptosis assay. HIF-1α, Bnip3 and caspase-3 levels were determined by real-time reverse transcription polymerase chain reaction and western blot for mRNA and protein, respectively. H/R resulted in severe injury in cultured rat cardiomyocytes and it upregulated HIF-1α and proapoptotic Bnip3 mRNA and protein expression. HIF-1α activity inhibited by siRNA significantly decreased (P<0.01) the rate of apoptotic cardiomyocytes induced by 5h of hypoxia followed by 6h of reoxygenation compared with cardiomyocytes without siRNA treatment. Additionally, the expression of Bnip3 and caspase-3 was also markedly reduced. We conclude that HIF-1α is a key regulator of apoptosis of cardiomyocytes induced by H/R. H/R enhances primary neonatal rat cardiomyocyte apoptosis through the activation of HIF-1α and the mechanism might involve Bnip3 and caspase-3. HIF-1α may be a possible therapeutic target to limit myocardial injury after myocardial infarction.