Introduction: Ischemic preconditioning (IPC) is an adaptive response of the heart to stress resulting in cardioprotection in following Ischiemia/reperfusion(I/R) injury. I/R injury induces hydrolysis of membrane phospholipids to arachidonic acids (AA) and AA is metabolised by cyclooxygenases (COXs), lipoxygenases (LOXs), and cytochrome P450 (CYPs) enzymes. It���s been well known that (4.PGE2 and PGI2 by) COX-2 mediates of cardioprotective effects through PI3K-Akt signaling pathways in IPC. Recent studies suggest that overexpression of CYP2C induce COX-2 expression and epoxyeicosatrienoic acids (EETs) mediate cardioprotection during I/R injury, however, the relation between CYPs and COX-2 in IPC is poorly understood. Therefore, we examine the role of CYP2J3, which is considered as major epoxygenase in heart, COX-2 and its metabolites in IPC. Meothods: IPC was achieved by 2 cycles of each 5 minutes ischemia and reperfusion before sustained 45 minutes I/R injury (1. in Langendorff mode) (n=5, each group). CYPs and COX-2 inhibitors were added to (2.the perfusion buffer) 20 min before IPC. LDH and infarct size was measured at 30 min and at 45 min after reperfusion. CYP2J3 and COX-2 expression were determined by westernblot and RT-PCR. AA metabolites were measured by liquid-chromatography mass spectrometry (LC-MS). Results: CYP2J3 expression was down-regulated in I/R injury, whereas its activity were recovered by IPC (3. which associated with an increased COX-2 expression). CYP2J3 expression was suppressed by EET antagonist EEZE and CYP epoxygenase inhibitor MS-PPOH, however, COX-2 expression was not affected by EEZE but reduced by MS-PPOH. The LDH and infarct size were reduced corresponding to COX-2 expression. Conclusion: I/R injury reduced CYP2J3 expression and enhanced by IPC. COX-2 expression was inhibited by CYP epoxygenase inhibitor MS-PPOH and COX-2 inhibitors. These data indicated that the enhanced activity of CYP epoxygenase is prerequisite for COX-2 mediated cardioprotection in IPC.