Fig. 6. Ceramides facilitate lipid uptake by mechanisms involving activation of PKCζ and CD36.

A) Relative abundance of CD36 mRNA was assessed by qPCR from livers of WT, Alb-AC, and Art-AC mice following 8 weeks of HFD-dox. B) Hepatic expression of CD36 was assessed by Western blotting, and representative results are shown in duplicate. C) Hepatic expression of PKCζ were assessed by western blotting and representative results are shown in duplicate. D) PKCζ activity was assessed from livers of WT, Alb-AC, and Art-AC mice following 8 weeks of HFD-dox. E-F) Following transfection with GFP, dnPKCζ, or caPKCζ H4IIe hepatocytes were treated with C2-ceramide (100 M, in 0.2% BSA) or BSA for 1 hour prior to assessment of ³H-palmitate uptake. Myristoylated PKCζ pseudosubstrate inhibitor was included for 90 minutes prior to ceramide treatments in non-transfected cells (n=4-8 from separate experiments). F) PKCζ activity was assessed from cell lysates harvested from cells represented in panel D. ‡ corresponds to comparison of C2-Ceramide treated cells with or without the PKCζ inhibitor. # corresponds to comparison of BSA treated cells that was transfected with either GFP or caPKCζ. G) Representative immunofluorescence of CD36 in H4IIe cells transfected with GFP, dnPKCζ, or caPKCζ. Each group is treated with either DMSO or C2-Ceramide. H) Circulating glucose levels measured during an insulin tolerance test (ITT) (0.75 U/kg) of Alb-AC or Art-AC mice and littermate controls after daily injections of ACPD. * corresponds to comparison of WT to Alb or Art-AC. ‡ corresponds to comparison of WT and WT + ACPD. # corresponds to comparison of WT and Alb or Art-AC + ACPD. I) Biochemical quantification of hepatic lipid accumulation in Alb-AC or Art-AC mice and littermate controls after daily injections of ACPD. *,‡,#: P<0.05, **P<0.01 by Student’s t test.