Adipose-derived stem cells (ASCs) induce therapeutic angiogenesis due to pro-angiogenic cytokines secretion. Superparamagnetic iron oxide (SPIO) nanoparticles are critical for magnetic resonance (MR) tracking of implanted cells. Hypoxia is a powerful stimulus for angiogenic activity of ASCs. In this study, we investigated whether therapeutic potency could be enhanced by implantation of hypoxia-preconditioned SPIO-labeled ASCs (ASCs) into the infarcted myocardium. ASCs and ASCs were cultured under 2% O (hypoxia) or 95% air (normoxia). Cells were intramyocardially injected into the infarcted myocardium after 48-h culture. We found that hypoxia culture increased the mRNA expression of hypoxia-inducible factor-1 alpha (HIF-1α) and vascular endothelial growth factor (VEGF) in ASCs and ASCs. The VEGF protein in the conditioned medium was significantly higher in hypoxic ASCs and ASCs than in normoxic ASCs and ASCs. The capillary density and left ventricular contractile function in the infarcted myocardium were significantly higher 4 weeks after implantation with hypoxic ASCs and ASCs than with normoxic ASCs and ASCs. Improvement in the capillary density and left ventricle function didn't differ between hypoxic ASCs-transplanted rats and hypoxic ASCs-transplanted rats. Hypoxic culture enhanced the angiogenic efficiency of ASCs. It was concluded that implantation of hypoxic ASCs or ASCs promotes therapeutic angiogenesis and cardiac function recovery in the infarcted myocardium. SPIO labeling does not impact the beneficial effect of hypoxic ASCs.