We studied the role of hypoxia-inducible factor 1-alpha (HIF-1 alpha) 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 48 h and were submitted to 5 h of hypoxia followed by 2, 6, or 12 h of reoxygenation. Small interfering RNA was used to target the HIF-1a gene. Cardiac myocyte apoptosis induced by H/R was assessed by Annexin V-FITC apoptosis assay. HIF-1 alpha, 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-1 alpha and proapoptotic Bnip3 mRNA and protein expression. HIF-1 alpha activity inhibited by siRNA significantly decreased (P <0.01) the rate of apoptotic cardiomyocytes induced by 5 h of hypoxia followed by 6 h of reoxygenation compared with cardiomyocytes without siRNA treatment. Additionally, the expression of Bnip3 and caspase-3 was also markedly reduced. We conclude that HIF-1 alpha 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-1 alpha and the mechanism might involve Bnip3 and caspase-3. HIF-1 alpha may be a possible therapeutic target to limit myocardial injury after myocardial infarction. (C) 2011 Elsevier Inc. All rights reserved.