Figure 7. Resveratrol activates AMPK in a SIRT1 dependent manner through deacetylation of LKB1.

(A) Representative immunoblot for p-AMPK (Thr172) and total AMPK in C2C12 cells infected with SIRT1 or non-targeting shRNA and treated with 10, 25 or 50 μM resveratrol for 24h.

(B) ATP content in C2C12 cells treated with 25 or 50 μM resveratrol for 24h (n=4) (*p<0.05 versus DMSO).

(C) Mitochondrial membrane potential in C2C12 cells treated with 25 or 50 μM resveratrol for 24h (n=4) (*p<0.05 versus DMSO).

(D) ATP content in C2C12 cells treated with 25 or 50 μM resveratrol for 1, 4, 6 and 12h (n=4) (*p<0.05 versus DMSO).

(E) Representative immunoblot for for p-AMPK (Thr172), and total AMPK in C2C12 cells treated with 25 or 50 μM resveratrol for 1, 4, 6 and 12h.

(F) NAD⁺ content in C2C12 cells treated with 25 or 50 μM resveratrol for 1, 4, 6 and 12h (n=4) (*p<0.05 versus 50 μM DMSO, #p<0.05 versus 25 μM DMSO).

(G) Representative immunoblot for for p-AMPK (Thr172), and total AMPK in primary myoblasts isolated from wild type and SIRT1 knockout mice and treated with 500 μM AICAR for 24h.

(H) Mitochondrial DNA content analyzed by quantitative PCR in primary myoblasts isolated from wild type and SIRT1 knockout mice and treated with 500 μM AICAR for 24h. Relative expression values were normalized to control. (n=3) experiments (*p<0.05 versus empty DMSO).

(I) ATP content in primary myoblasts isolated from wild type and SIRT1 knockout mice and treated with 500 μM AICAR for 24h (n=3) (*p<0.05 versus DMSO).

(J) C2C12 cells infected with SIRT1 or non-targeting shRNA, and expressing Flag-LKB1 were treated with resveratrol 25 μM for 24h and LKB1 acetylation was tested in Flag immunoprecipitates. Total LKB1 was evaluated in total extracts as input.

(K) Moderate doses of resveratrol activate AMPK and stimulate mitochondrial biogenesis in a SIRT1-dependent manner that results in improvement of mitochondrial function.

Values are expressed as mean ± SEM.