Research Paper Volume 11, Issue 23 pp 11520—11540

Figure 2. BMP4 inhibits triglyceride accumulation through regulating the genes involved in lipid metabolism and members of mTORC1 signaling pathway in hepatocytes. (A) Primary mouse hepatocytes were infected with Ad-B4 or Ad-GFP for 7 days, and subjected to ORO staining. (B) Primary mouse hepatocytes were infected with Ad-B4 or Ad-GFP for 36h and 72h. Total RNA was isolated and subjected to TqPCR analysis of the expression of the genes involved in triglyceride synthesis and storage (a and b) and triglyceride breakdown (c and d). Relative expression was calculated by dividing the relative expression values (i.e., gene/Gapdh) in “**” p < 0.001, “*” p < 0.05, Ad-B4 group vs. Ad-GFP group. (C) Oleic acid (0.05mM)-induced hepatocytes were treated with 1nM PF-04691502 or DMSO for 7 days, and subjected to ORO staining. (D) Primary mouse hepatocytes were infected with Ad-B4 or Ad-GFP for 36h and 72h. Total RNA was isolated and subjected to TqPCR analysis of the expression of the members of mTORC1 signaling pathway (a and b). Relative expression was calculated by dividing the relative expression values (i.e., gene/Gapdh) in “**” p < 0.01, “*” p < 0.05, Ad-B4 group vs. Ad-GFP group. (E) Primary mouse hepatocytes were infected with Ad-B4 or Ad-GFP for 72h, and total cell lysate was subjected to Western blotting analysis of the expression of the members of mTORC1 signaling pathway. Each assay condition was done in triplicate, and representative images are shown or indicated by arrows.