Abstract: Objective To explore the effect and the mechanism of isoflurane on human cardiac myocytes (HCM) injury induced by anoxia/reoxygenation (AR). Methods HCM cells were divided into control group (con), anoxia/reoxygenation group (AR) and isoflurane (0.5%, 1%, 1.5% and 2%) treatment group (n=6). Cell viability , LDH activity, apoptosis and the expression level of Anoxia inducible factor-1α (HIF-1α) were detected using CCK-8 assay, LDH activity assay kit, Annexin V-FITC/PI staining, PCR and western blot, respectively. Results Compared with the con group, cell viability decreased, LDH activity and apoptosis cells increased in AR group. Isoflurane can significantly relieve the decrease of cell viability, the increase of LDH activity and apoptosis cells, and the down-regulation of the mRNA and protein expression level of HIF-1α induced by AR (P<0.05). Compared with AR group, the mRNA and protein expression level of HIF-1α in siRNA transfected group is significantly decreased (P<0.05). 2% isoflurane significantly relieve the increase of cell viability, and the decrease of LDH activity and apoptotic cells induced by HIF-siRNA in AR injuried HCM cells(P<0.05).Conclusion Isoflurane can protect HCM cells from AR injury partly through up-regulate the expression of HIF-1α.