Cytokines seem to play an important role in the metabolic disturbances that are commonly associated with sepsis. In this study, we analyzed the effect of tumor necrosis factor, interleukin-1 and interleukin-6, as well as that of tumor necrosis factor in combination with interleukin-1 or interleukin-6, both on free fatty acids and on phospholipid synthesis by isolated rat hepatocytes. All three cytokines and combinations caused inhibited D-[U-14C]glucose incorporation into phosphatidylcholine (tumor necrosis factor = 6.39 ± 1.13 pmol/μg protein vs. control = 12.90 ± 0.98 pmol/μg protein, n = 7; p < 0.001). However, when [U-14C]palmitate was used as radioactive precursor, tumor necrosis factor, either alone or in the presence of the other cytokines, stimulated phosphatidylcholine synthesis. D-[U-14C]glucose incorporation into free fatty acids and triacylglycerol was also significantly stimulated, whereas phosphatidylinositol labeling was found inhibited by the assayed cytokines. Our results demonstrate an effect of sepsis-related cytokines, more evident for tumor necrosis factor, on hepatocyte lipid synthesis either from glucose or palmitate. Also, the findings support the hypothesis that cytokine-induced changes in hepatocyte lipid synthesis can contribute to the impairment in lipidic metabolism seen in patients with sepsis. (Hepatology 1994;20:924–931).