Overexpression merely of p53 decreases SIRT1 abundance and attenuates the effects of metformin on AMPK activation and cellular triglycerides. Overexpression of p53 decreased SIRT1 protein abundance and attenuated metformin-induced AMPK and ACC phosphorylation (Fig. 5, A and B). In addition, treatment with nutlin-3, which increased the abundance of p53 (Fig. 4C), reduced the transcription of SIRT1 as evidenced by reduced SIRT gene expression at both 12 and 16 h (Fig. 5C). Since both AMPK and SIRT1 have lipid-lowering effects in vitro and in vivo (8, 12, 22, 35, 60, 62), we hypothesized that p53 overexpression would attenuate the lipid-lowering effect of metformin. As shown in Fig. 6, the ability of metformin to inhibit triglyceride accumulation was blunted in HepG2 cells in which p53 was overexpressed.

Fig. 5. Overexpression of p53 decreases SIRT1 abundance and diminishes AMPK activation by metformin. Cells infected with adenovirus Ad-��-gal (control) or Ad-p53 for 24 h in 25 mM glucose, followed by 24 h incubation in 25 mM glucose with or without 2 … Fig. 6. Increasing p53 abundance attenuates the effect of metformin on cellular triglycerides. HepG2 cells infected with adenovirus Ad-��-gal (control) or Ad-p53 for 24 h in 25 mM glucose media, followed by 24 h incubation in 25 mM glucose with or without … Metformin effects are partially restored by the SIRT1 activator SRT2183 in cells overexpressing p53. We next tested whether SIRT1 activation in cells overexpressing p53 would restore the effects of metformin on AMPK activity and triglyceride accumulation.

Coincubation with the SIRT1 activator SRT2183 restored some, but not all, of the effects of metformin in these cells, as evidenced by their decreased triglyceride content and increased AMPK phosphorylation (Fig. 7). Although no effect on ACC phosphorylation was observed (Fig. 7A), these results support the notion that p53 overexpression attenuates the effects of metformin in part by decreasing SIRT1 activity. Fig. 7. The effects of metformin are partially restored by the SIRT1 activator SRT2183. A: representative Western blot demonstrating partial restoration of p-AMPK with SRT2183, with no apparent effect on Ac-H3 or Ac-p53. B: triglyceride content in cellular lysates … DISCUSSION In this study, we investigated the interactions between AMPK/SIRT1 signaling and p53 protein abundance in HepG2 cells under conditions of nutrient excess.

The results demonstrate that activation of AMPK and SIRT1 in response to metformin treatment decreases p53 abundance. They also reveal that metformin diminishes oxidative stress and enhances p53 deacetylation, likely contributing to the MDM2-mediated increase in p53 degradation. Conversely, we showed that overexpression of p53 diminishes SIRT1 abundance and prevents metformin-induced activation Anacetrapib of AMPK and reductions in triglyceride content.