AIMS: Apoptosis plays a critical role in cardiomyocyte loss during ischaemic heart injury. A detailed understanding of the mechanism involved has a substantial impact on the optimization and development of treatment strategies. Here, we report that the expression of SIRT4, a mitochondrial sirtuin, is markedly down-regulated in hypoxia-induced apoptosis of H9c2 cardiomyoblast cells. METHODS AND RESULTS: SIRT4 interference significantly alters H9c2 cell viability, apoptotic cell number and caspase-3/7 activity. Furthermore, SIRT4 expression can affect the ratio of pro-caspase 9/caspase 9 or pro-caspase 3/caspase 3, an affect Bax translocation, which in turn alters the development of H9c2 cell apoptosis. CONCLUSION: These results suggest that SIRT4 is a key player in hypoxia-induced cardiomyocyte apoptosis, and that strategies based on its enhancement might be of benefit in the treatment of ischaemic heart disease.