S6K signal, but not PTEN, is altered by exposure to palmitic acid (PA). A: Mouse embryonic fibroblasts (mEF), beta-cells, and cultured islets treated with 0.4 mmol/L PA or dimethyl sulfoxide (DMSO; Veh) were lysed and analyzed for PTEN expression. Each lane represents an independent cell culture experiment. Experiment was repeated multiple times. B: Gene enrichment analysis of mTOR signal genes in islets from mice fed HFD for 4 months versus those fed normal chow (NC). C: mEF cells exposed to 0.4 mmol/L PA or vehicle with 6 mmol/L glucose were analyzed for phospho-AKT (p-AKT), total AKT (tAKT), and phospho-S6K (P-S6K). Each lane represents an independent cell culture experiment. Right two panels, densitometry quantification of the ratio of P-AKT versus tAKT, and S6K cyclins versus actin. D and E: Three beta-cell lines were treated with 0.4 mmol/L PA with 6 mmol/L glucose (D), and isolated islets from HFD (4 months) versus normal chow (NC) fed mice (E) were analyzed for P-AKT, tAKT, phospho-insulin receptor substrate 1 (P-IRS1), phospho-mTOR (P-mTOR), P-S6K, and total S6K (tS6K). Each lane represents an independent cell culture experiment or mouse. Bottom panels, densitometry quantification of the ratio of P-AKT versus tAKT. Experiments were repeated multiple times. n = 3 (B and C); n = 5 (D and E). ∗P < 0.05 between PA- and vehicle-treated samples.