Objective To explore the protective effects of erythropoietin (EPO) on cardiomyocytes against hypoxia/reoxygenation (HR) injury. Method Isolated and cultured rat eardiomyocytes were subjected to 2-hour hypoxia followed by 1 hour reoxygenation to establish model of HR injury. Cardiomyocytes were randomly divided into 4 groups: sham group, HR group, HR + EPO-treated group (EPO 10 U/ml), and HR + EPO + UO126-treated group (U0126 10?mol/L). The concentration of lactate dehydrogenase (LDH) in culture medium was detected by automatic biochemical analyzer; viability of eardiomyocytes was measured by MTT assay; apoptosis ratio was determined by TUNEL technology and Annexin-V-FITC with flow cytometer (FCM); level of extracellular signal- regulated kinase 1/2 (ERK_(1/2)) and phospho-ERK_(1/2) were measured by Western-blot analysis. Results EPO significantly decreased the leakage of LDH, enhanced activity, reduced apoptosis ratio, and increased level of phospho-ERK_(1/2).However, the effects were blocked by U0126, an inhibitor of MAPK. Conclusions EPO has the protective effects on cardiomyocytes against HR injury possibly via the mechanism of activation of ERK_(1/2) and inhibition of apoptosis.