Epoxyeicosatrienoic acids (EET), the primary arachidonic acid metabolites of cytochrome P450 2J (CYP2J) epoxygenases, possess potent vasodilatory, anti-inflammatory, antiapoptotic, and mitogenic effects. To date, little is known about the role of CYP2J2 and EETs in tumor necrosis factor (TNF)-alpha-induced cardiac injury. We utilized cell culture and in vivo models to examine the effects of exogenously applied EETs or CYP2J2 overexpression on TNF-alpha-induced cardiac apoptosis and cardiac dysfunction. In neonatal rat cardiomyocytes, TNF-alpha-induced apoptosis was markedly attenuated by EETs or CYP2J2 overexpression, leading to significantly improved cell survival. Further studies showed that TNF-alpha decreased expression of the antiapoptotic proteins Bcl-2 and Bcl-xL, decreased I kappa B alpha and PPAR gamma, and also inhibited PI3K-dependent Akt and EGFR signaling. Both EETs and CYP2J2 overexpression reversed the effects of TNF-alpha on these pathways. Furthermore, overexpression of CYP2J2 in rats prevented the decline in cardiac function that is normally observed in TNF-alpha challenged animals. These results demonstrate that EETs or CYP2J2 overexpression can prevent TNF-alpha-induced cardiac cell injury and cardiac dysfunction by inhibiting apoptosis, reducing inflammation, and enhancing PPAR gamma expression. Targeting the CYP2J2 epoxygenase pathway may represent a novel approach to mitigate cardiac injury in diseases such as heart failure, where increased TNF-alpha levels are known to occur.-Zhao, G., J. Wang, X. Xu, Y. Jing, L. Tu, X. Li, C. Chen, K. Cianflone, P. Wang, R. T. Dackor, D. C. Zeldin, and D. W. Wang. Epoxyeicosatrienoic acids protect rat hearts against tumor necrosis factor-alpha-induced injury. J. Lipid Res. 2012. 53: 456-466.