MicroRNAs (miRNAs) are a class of endogenous non-coding RNAs involved in regulating various biological processes at the post-transcription level. Accumulating evidence suggests that hypoxia caused by acute myocardial infarction induces cardiomyocyte damage including apoptosis. Previous studies regarding the miRNAome in H9c2 cells under hypoxia have shown that hypoxia modulates miRNA expression in H9c2 cells, including miR-532-5p. We therefore investigated whether miR-532-5p has a potential function in the cardiomyocyte response to hypoxia. In the present study, we found that miR-532-5p, which was down-regulated in hypoxia-exposed H9c2 cells and the myocardium of acute myocardial infarction rats, alleviated hypoxia-induced H9c2 cell apoptosis. Additionally, we identified PDCD4 as the direct target of miR-532-5p, which partly elucidates the anti-apoptotic mechanism of miR-532-5p. In summary, this study revealed that miR-532-5p has cardioprotective effects against hypoxia-induced apoptosis in H9c2 cells.