Nicorandil, an adenosine triphosphate (ATP)-sensitive potassium (K(ATP)) channel opener, has been shown to exert a significant protective effect against ischemic heart injury. In the present study, we investigated the anti-apoptotic effects of nicorandil on rat mesenchymal stem cells (MSCs) subjected to hypoxia and serum deprivation (H/SD), as well as the potential underlying mechanisms. Apoptosis was induced in the MSCs by exposure to H/SD, and the apoptotic rates and reactive oxygen species (ROS) levels were determined by flow cytometry. The mitochondrial inner membrane potential was measured using the membrane-permeable dye, JC-1. Western blot analysis was used to measure the levels of Akt, Bcl-2, Bax, cytochrome c and cleaved caspase-3. The cell proliferative ability was assessed using the cell counting kit-8 (CCK-8) and 5-ethynyl-2'-deoxyuridine (EdU) assay. The results revealed that H/SD-induced apoptosis was significantly attenuated by treatment with nicorandil in a concentration-dependent manner. Moreover, nicorandil markedly reduced the levels of ROS which were induced by exposure to H/SD, and increased the stability of mitochondrial membrane potential and the Bcl-2/Bax ratio, while it concomitantly decreased the H/SD-induced cleavage of caspase-3 and the release of cytochrome c. Treatment with the phosphoinositide 3-kinase (PI3K) inhibitor, LY294002, abolished the beneficial effects of nicorandil on the MSCs. In conclusion, the findings of the present study indicate that nicorandil exerts protective effects against MSC apoptosis induced by H/SD and that these effects are mediated through the PI3K/Akt, mitochondrial and ROS signaling pathways.