Previous studies have demonstrated that a high-fat diet enriched with polyunsaturated fatty acids (ω-3 PUFA) has a protective effect on the liver. Lipid emulsion increases hepatocyte uncoupling protein 2 (UCP2) production that seems to have a protective effect by inhibiting mitochondrial ROS production. We hypothesized that a high-fat diet enriched with ω-3 PUFA could modify mitochondrial functions and therefore protect the liver from ischemia/reperfusion injury.Aim: To evaluate the effect of a high-fat diet enriched with ω-3 PUFA (cold fish oil) on hepatic mitochondrial functions and hepatic disturbances associate with hepatic ischemia/reperfusion (I/R) injury.Methods: Wistar male rats were divided in 2 groups: Group I (n=2):rats with fatty liver as induced by high-fat diet enriched with ω-3 PUFAs given for 4 weeks and Group II (n=20) that received standard diet. Ten animals of each group were killed and their livers were collected for determination of oxidation and phosphorylation of liver mitochondria and liver histological analysis. Activities of AST and ALT were also determined. Ten animals of each group were submitted to a partial liver ischemia performed by clamping the pedicle from medium and left anterior lateral liver segments during an hour. After 2 hours of reperfusion, liver mitochondrial oxidation and phosphorylation were analyzed. Serum levels of TNF-α, IL-6, IL-10, and AST, ALT activities were also determined. Results: Mild liver steatosis was observed in the group I in comparison to group II. The group I showed a significant liver mitochondrial uncoupling: respiration state 4 (23.6±1.97 vs 15.5±1.35 nmol O2 mgprot/min) and RCR (3.09±0.22 vs 3.73±0.14) in relation to the group II (p<0.05). AST and ALT activities were not changed. After I/R, there was an increase in state 3 (79.1±5.42 vs 30.8±2.97) and state 4 (26.5±2.45 vs 15.5±1.01) respiration, and in the respiration control rate (3.06±0.11 vs 1.98±0.15) in the group I compared to group II(p<0.05). The group I showed also lower activities of AST: 1087±775 vs 6607±806 and ALT: 1333±892 vs 7132±1102 and the histological changes observed were significantly less intense (p<0.05). A significant reduction was observed in the levels of TNF-α, IL-6, and IL-10 in the group I as compared to group II (p<0.05).Conclusion: High-fat diet enriched with ω-3 PUFA protected the liver from ischemia/reperfusion injury probably by a mechanism associated to changes mitochondrial function.FAPESP, CNPq