Twenty-four hours after plating, mouse hepatocytes were infected with a combination of adenovirus as indicated (GFP, SCD1, ChREBP, shCTRL, and/or shSCD1) and described in the Methods section. Cells were then incubated for 24 hours with BSA or 0.48 mM albumin-bound PALM. An insulin (1 nM) time course was performed for 2 and 5 minutes (n = 6 independent cultures). (A) Western blot analysis of insulin-mediated Akt phosphorylation on Ser473, Thr308, total Akt, ChREBP, and SCD1. Representative Western blots are shown. β-Actin was used as a loading control. (B) Quantification of the ratio of Thr308 compared with total Akt protein content is shown. *P < 0.05 compared with GFP BSA. (C) qRT-PCR analysis of chop mRNA under the indicated culture conditions. *P < 0.05 GFP PALM compared with GFP BSA.