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. 2020 Nov 13;8(4):531–544. doi: 10.1016/j.gendis.2020.11.004

Figure 5.

Figure 5

The signaling mechanism of BMP4-regulated glucose metabolism in hepatocytes in vivo. Recombinant adenovirus Ad-B4 or Ad-GFP was injected intrahepatically into 4-week old C57BL/6 mice (male, n = 10/time point/group). The mice were sacrificed after 4 weeks and 12 weeks, respectively. Total RNA was isolated from the liver of Ad-B4 group and Ad-GFP group. TqPCR analysis was carried out to detect the expression of the members of mTORC2 signaling pathway. 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 (A) Total tissue proteins were isolated from the liver samples of Ad-B4 group and Ad-GFP group after 4 weeks and 12 weeks, and Western blotting was carried out to detect the expression or phosphorylation level of proteins related to glucose levels regulated by mTORC2 signaling pathway, including AKT1+2+3, p-AKT, GSK-3β, p-GSK-3β, FOXO1, p-FOXO1, and β-ACTIN was used as a loading control (B) The liver sections were subjected to IHC staining to detect the expression or phosphorylation level of genes related to glucose metabolism regulated by mTORC2 signaling pathway, including AKT1+2+3, p-AKT, GSK-3β, p-GSK-3β, FOXO1, and p-FOXO1. Staining results were recorded by light microscopic examination (x400) (C) Each assay condition was done in triplicate. Representative images are shown.