Effects of palmitate on the PERK- eIF2α-ATF4 pathway. (A) Western blotting analysis. HepG2 cells were treated with 2% BSA (as a negative control) and 700 μM Palmitate (PA). After 3, 6, 24 hrs, the cell extracts were collected and subjected to immunoblot analysis for total PERK (both phosphorylated and non-phosphorylated proteins), p-PERK (phosphorylated proteins), eIF2α, p-eIF2α, and ATF4. Actin served as a loading control. (B) Quantification. From Figure 1A, the phosphorylation levels of PERK and eIF2α were quantified and normalized to the total protein levels of PERK and eIF2α, respectively. The protein expression levels of ATF4 were also quantified. The protein fold changes were calculated at each time point using the following equation: (Protein expression level)palmitate/(protein expression level)BSA. Data represent the mean and standard deviation of three independent experiments: *p < 0.05 and **p < 0.01 vs. the control at each time point.