SRPIN340, a selective serine-arginine protein kinase 1/2 (SRPK1/2) inhibitor, has been shown to have antiviral and anti-angiogenesis effects. However, its role in the heart is unknown. The present study explored the role of SRPIN340 in myocardial protection and the related mechanisms. During challenge with H2O2, cardiomyocytes (CMs) pretreated with SRPIN340 showed strikingly more injury tolerance, which was manifested as reduced lactate dehydrogenase (LDH) release and lower apoptotic index. Further research showed that SRPIN340 activated AKT under basal conditions, and AKT inhibition abolished the protective effects of SRPIN340 treatment during H2O2 stress. The protective effect of SRPIN340 was also demonstrated in perfused rat hearts subjected to ischemia/reperfusion (I/R). Collectively, our results reveal the beneficial effects of SRPIN340 against H2O2-induced oxidative damage in CMs and 1/R-induced injury in a Langendorff heart model, supporting a potential application of SRPIN340 in the clinically relevant context of reperfusion. The effectiveness of SRPIN340 may be attributed to AKT signal activation. (C) 2019 Elsevier Inc. All rights reserved.