Figure 10.

The metformin-mediated Hh activation regulates autophagy through a mechanism involving BNIP3. (a) Cell lysates of HUVECs were used to detect the expression of BNIP3 and ATG proteins by immunoblotting. HUVECs were transduced with Ad-sh-GLI1 and Ad-Scrambled shRNA, respectively. After transduction, HUVECs were cultured either in NG or HG medium in the presence or absence of MET (50 μM) for 72 h, MAN was used as the osmotic control for HG. (b) The quantitative analysis of each immunoblot relative to GAPDH protein level in (a). Data are expressed as fold change relative to Ad-Scrambled shRNA-transduced HUVECs cultured in NG, values displayed are means ± SEM of 6 independent experiments. # P < 0.05 vs. Ad-Scrambled shRNA-transduced HUVECs cultured in NG or MAN; * P < 0.05 vs. Ad-Scrambled shRNA-transduced HUVECs cultured in HG; % P < 0.05 vs. Ad-Scrambled shRNA-transduced HUVECs cultured in HG coincubated with MET; ns, non-significant. (c) HUVECs were treated as in (a), and the cell lysates were subjected to immunoprecipitation with BCL2 antibody, followed by immunoblotting with the indicated antibodies. Cell lysates were also subjected to immunoprecipitation with IgG as negative control. (d) HUVECs were treated as in (a), and the cell lysates were subjected to immunoprecipitation with either BCL2 or BECN1 antibody, followed by immunoblotting with the indicated antibodies. Cell lysates were also subjected to immunoprecipitation with IgG as negative control.