Fig. 6.

Effects of AICAR, Dorsomorphin and EX-527 on p66Shc mRNA expression and MDA levels under HG ambience in HK-2 cells treated with probucol. A: Real-time PCR showing p66Shc mRNA expression in HK-2 cells under HG conditions treated with or without probucol and AICAR, Dorsomorphin or EX-527. B: MDA content in the medium of HK-2 cells. Values are the mean ± SE (n = 4), *P < 0.01 vs. LG; #P < 0.01 vs. HG; $P < 0.01 vs. HG + Probucol; @ P < 0.01 vs. HG + Probucol + Ex-527. C: Cellular immunofluorescence with anti-p-AMPK, anti-Sirt1 and anti-p66Shc antibodies. High levels of p-AMPK were observed under LG conditions and were accompanied by Sirt1 expression. HG reduced this fluorescence. This effect was reversed by probucol treatment and Dorsomorphin and EX-527 treatment but was increased by AICAR treatment. Contrasting results were observed regarding p66Shc and DHE staining (lower panel). D–F: Representative Western blots (D) and densitometric analysis of Sirt1 (E) and p66Shc protein expression (F) in HK-2 cells exposed to HG conditions with or without treatment with various reagents. Quantitative analyses of the Western blotting results are included, and the data were normalized to β-actin and are presented as the mean ± SEM (n = 4). *P < 0.01 versus LG; #P < 0.01 vs. HG; $P < 0.01 vs. HG + Probucol; @ P < 0.01 vs. HG + Probucol + Ex-527.