Calorie restriction-like effects of 30 days of resveratrol supplementation on energy metabolism and metabolic profile in obese humans

Abstract

Resveratrol is a natural compound that affects energy metabolism and mitochondrial function and serves as a calorie restriction mimetic, at least in animal models of obesity. Here, we treated 11 healthy, obese men with placebo and 150 mg/day resveratrol (resVida) in a randomized double-blind crossover study for 30 days. Resveratrol significantly reduced sleeping and resting metabolic rate. In muscle, resveratrol activated AMPK, increased SIRT1 and PGC-1α protein levels, increased citrate synthase activity without change in mitochondrial content, and improved muscle mitochondrial respiration on a fatty acid-derived substrate. Furthermore, resveratrol elevated intramyocellular lipid levels and decreased intrahepatic lipid content, circulating glucose, triglycerides, alanine-aminotransferase, and inflammation markers. Systolic blood pressure dropped and HOMA index improved after resveratrol. In the postprandial state, adipose tissue lipolysis and plasma fatty acid and glycerol decreased. In conclusion, we demonstrate that 30 days of resveratrol supplementation induces metabolic changes in obese humans, mimicking the effects of calorie restriction.

Figure 1. Resveratrol decreases postprandial energy expenditure and lowers adipose tissue lipolysis

Data are obtained on whole body (A-D), by microdialysis (E-F) and indirect calorimetry (G-I). Plasma metabolites during the postprandial microdialysis test after 30 days of placebo or resveratrol supplementation: (A) glucose response (B) insulin response (C) NEFA response (D) free glycerol response. Interstitial glycerol responses in (E) adipose tissue and (F) skeletal muscle during the postprandial microdialysis test. Indirect calorimetry results during the postprandial microdialysis test: (G) energy expenditure (H) respiratory quotient (I) fat oxidation. Values are given as means ± SEM (n=10).

Figure 2. Resveratrol increases oxidative phosphorylation gene expression in man and mice

(A) One-way hierarchical clustering of genes significantly changed in a microarray of vastus lateralis of subjects receiving resveratrol (RSV) or placebo. In muscle, 469 genes were differentially expressed between placebo and RSV (n=10, p<0.05). (B) Gene set enrichment analysis (GSEA) of muscle microarray revealed that gene sets related to mitochondrial oxidative phosphorylation were significantly increased (left panel) and cytokine signaling decreased (right panel) following resveratrol supplementation in humans. (C) Previously published microarray data of mice treated with 400 mpkd RSV (Lagouge et al., 2006) were reanalyzed by GSEA, showing, like the human data, a marked enrichment of oxidative phosphorylation genes. See also table S3.

Figure 3. Resveratrol increases AMPK activity, increases mitochondrial efficiency and respiration on fatty acid substrates

After 30 days of resveratrol or placebo, a muscle biopsy was obtained from the vastus lateralis muscle. Of total protein extracts from the vastus lateralis muscle, 50 μg of protein was used to check the expression levels of several proteins by Western blotting. (A) The level of phosphorylation of residue Thr¹⁷² of the AMPKa subunit (top panel), indicative of AMPK activity, by Western blotting in a total of 9 subjects per group (representative subset is shown). Total level of the AMPKa protein is shown in the bottom panel. The ratio resveratrol/placebo is shown on the right of panel A. Relative quantification of the ratio of pAMPK/AMPK is shown in figure S1A. (B) The protein content of the individual complexes of the electron transport chain are quantified by Western blotting in vastus lateralis muscle of 11 subjects per group. An antibody cocktail that detects all five complexes is used, and the data are represented as mean ± SEM for the individual complexes as well as for the average expression of all the complexes. A representative Western blot is included in figure S1B. (C) SIRT1 and PGC-1α protein levels in vastus lateralis muscle by Western blotting (n=8 for SIRT1 and n=11 for PGC-1α, a representative subset is shown). The relative expression of both proteins is corrected for the loading control, tubulin for SIRT1 and actin for PGC-1α. The ratio of resveratrol/placebo is shown on the right of panel C. The relative quantification of the Western blots is shown in figure S1C. (D) Citrate Synthase activity was significantly increased in vastus lateralis muscle of resveratrol (RSV)-treated subjects (n=11), * p<0.05. (E) In vivo mitochondrial function expressed as PCr half-time (n=11). Evaluation of ex vivo mitochondrial function in vastus lateralis muscle: (F) ADP-stimulated respiration (state 3) upon a lipid substrate. (G) State 3 respiration fueled by Complex I-linked substrates. (H) State 3 respiration upon parallel electron input into Complex I and II, * p<0.05. (I) Maximally uncoupled respiration upon FCCP, * p<0.05. (J) Mitochondrial respiration uncoupled from ATP synthesis (state 4o). Values are given as means ± SEM (n=10). M, malate; O, octanoyl-carnitine; G, glutamate; S, succinate.

Figure 4. Resveratrol decreases intrahepatic lipid content but increases intramyocellular lipid content

Ectopic lipid deposition after 30 days of placebo or resveratrol supplementation. (A) Typical lipid regions of hepatic ¹H-MR spectra from a subject receiving placebo (left) and resveratrol (right) at 3T, scaled to the water resonance (reference). The CH₂ peak is used for quantification, the area under the curve is proportional to the lipid content in the liver. Quantification revealed that intrahepatic lipid content was reduced by resveratrol (n=9), * p<0.05. (B) Images of representative cross section of the vastus lateralis muscle from a subject receiving placebo (top) and resveratrol (bottom). Sections are stained for intramyocellular lipids with Oil Red O staining (in red), muscle laminin (in blue) and type 1 muscle fibers (in green) (400x magnification). IMCL content significantly increased by resveratrol treatment in muscle fibers, which was mostly accounted for by an increased lipid storage in type I muscle fibers (n=10), * p<0.05. Values are given as means ± SEM.