Fig. 3. FFA activate PERK signaling. INS-1E cells were cultured in the presence or absence of oleate (O, gray diamonds), palmitate (P, black squares) or oleate plus palmitate (OP, white circles) at 11 mM glucose. Western blots using phospho-specific PERK (P-PERK), phospho-specific eIF2 (P-eIF2) and anti-ATF3 antibodies after 6-12 hours exposure to FFA are shown. Total eIF2 or β-actin was used as control for protein loading. One representative experiment for three to six similar experiments is shown, as well as mean optical density measurements of the western blots following 6-, 12- and 24-hour FFA exposure; n=2-6. ATF3 and CHOP mRNA expression following a 6- to 48-hour FFA exposure was analyzed by real-time PCR, normalized for the expression level of the housekeeping gene GAPDH and expressed as fold induction of the control. The results represent means ± s.e.m. of 6-11 independent experiments. ^*P<0.05, ^**P<0.01 vs control.