J Vet Sci.  2014 Jun;15(2):179-185. 10.4142/jvs.2014.15.2.179.

Effects of resveratrol on the insulin signaling pathway of obese mice

• Hong HJ
• Kang W
• Kim DG
• Lee DH
• Lee Y
• Han CH

• KMID: 1784637
• DOI: http://doi.org/10.4142/jvs.2014.15.2.179

Abstract

The present study was conducted to investigate the effects of resveratrol on the insulin signaling pathway in the liver of obese mice. To accomplish this, we administered resveratrol to high fat diet-induced obese mice and examined the levels of protein phosphorylation in the liver using an antibody array. The phosphorylation levels of 10 proteins were decreased in the high fat diet and resveratrol (HFR) fed group relative to the levels in the high fat diet (HF) fed group. In contrast, the phosphorylation levels of more than 20 proteins were increased in the HFR group when compared with the levels of proteins in the HF group. Specifically, the phosphorylation levels of Akt (The308, Tyr326, Ser473) were restored to normal by resveratrol when compared with the levels in the HF group. In addition, the phosphorylation levels of IRS-1 (Ser636/Ser639), PI-3K p85-subunit alpha/gamma(Tyr467/Tyr199), PDK1 (Ser241), GSK-3alpha (S21) and GSK-3 (Ser9), which are involved in the insulin signaling pathway, were decreased in the HF group, whereas the levels were restored to normal in the HFR group. Overall, the results show that resveratrol restores the phosphorylation levels of proteins involved in the insulin signaling pathway, which were decreased by a high fat diet.

Keyword

Akt phosphorylation; antibody array; insulin signaling pathway; obese mice; resveratrol

MeSH Terms

Animals
Anti-Inflammatory Agents/*pharmacology
Fluorescent Antibody Technique
Insulin/*physiology
Liver/*metabolism
Male
Mice
Mice, Inbred C57BL
Mice, Obese
Phosphorylation
Proteins/metabolism
Signal Transduction/*drug effects
Stilbenes/*pharmacology
Anti-Inflammatory Agents
Insulin
Proteins
Stilbenes

Figure

• 

  Fig. 1 Scanned image of antibody array slides. Proteins were extracted from livers of mice fed a normal diet (control), high fat diet (HF), and high fat diet with resveratrol (HFR). Proteins extracted from each group (n = 12) were combined, and phosphorylation levels were analyzed using an antibody array. Each slide consists of an array of antibodies with six replicates per antibody, and multiple positive and negative controls to maximize data quality and reproducibility. Signals from three different array chips were normalized based on the GAPDH signal after background subtraction.

• 

  Fig. 2 Restoration effects of resveratrol on protein phosphorylation in the insulin signaling pathway of obese mice. (A) The phosphorylation levels of IRS-1 (Ser636, Ser639). (B) The levels of PI3-kinase p85-subunit α/γ (Tyr467/Tyr199) and PDK1 (Ser241) among the control, HF, and HFR groups were compared. Proteins extracted from each group (n = 12) were combined, and phosphorylation levels were analyzed using the antibody array.

• 

  Fig. 3 Restoration effects of resveratrol on the phosphorylation of Akt and GSK3 of obese mice. (A) The phosphorylation levels of Akt (Thr308, Tyr326, Ser473). (B) The levels of GSK-3α (S21) and GSK-3 (Ser9) among the control, HF, and HFR groups were compared. Proteins extracted from each group (n = 12) were combined, and phosphorylation levels were analyzed using the antibody array.

• 

  Fig. 4 Effects of resveratrol on protein phosphorylation in the insulin signaling pathway and the mTOR pathway of obese mice.

Reference

1. Alberti KG, Zimmet P, Shaw J. The metabolic syndrome-a new worldwide definition. Lancet. 2005; 366:1059–1062.

2. Baur JA, Pearson KJ, Price NL, Jamieson HA, Lerin C, Kalra A, Prabhu VV, Allard JS, Lopez-Lluch G, Lewis K, Pistell PJ, Poosala S, Becker KG, Boss O, Gwinn D, Wang M, Ramaswamy S, Fishbein KW, Spencer RG, Lakatta EG, Le Couteur D, Shaw RJ, Navas P, Puigserver P, Ingram DK, de Cabo R, Sinclair DA. Resveratrol improves health and survival of mice on a high-calorie diet. Nature. 2006; 444:337–342.

3. Bhat KPL, Kosmeder JW 2nd, Pezzuto JM. Biological effects of resveratrol. Antioxid Redox Signal. 2001; 3:1041–1064.

4. Casamayor A, Morrice NA, Alessi DR. Phosphorylation of Ser-241 is essential for the activity of 3-phosphoinositide-dependent protein kinase-1: Identification of five sites of phosphorylation in vivo. Biochem J. 1999; 342:287–292.

5. Conus NM, Hannan KM, Cristiano BE, Hemmings BA, Pearsom RB. Direct identification of tyrosine 474 as a regulatory phosphorylation site for the Akt protein kinase. J Biol Chem. 2002; 277:38021–38028.

6. Duffy SJ, Vita JA. Effects of phenolics on vascular endothelial function. Curr Opin Lipidol. 2003; 14:21–27.

7. Fauconneau B, Waffo-Teguo P, Huguet F, Barrier L, Decendit A, Merillon JM. Comparative study of radical scavenger and antioxidant properties of phenolic compounds from Vitis vinifera cell cultures using in vitro tests. Life Sci. 1997; 61:2103–2110.

8. Frankel EN, Waterhouse AL, Kinsella JE. Inhibition of human LDL oxidation by resveratrol. Lancet. 1993; 341:1103–1104.

9. Hansen TK, Thiel S, Wouters PJ, Christiansen JS, Van den Berghe G. Intensive insulin therapy exerts antiinflammatory effects in critically ill patients and counteracts the adverse effect of low mannose-binding lectin levels. J Clin Endocrinol Metab. 2003; 88:1082–1088.

10. Hsieh T, Juan G, Darzynkiewicz Z, Wu JM. Resveratrol increases nitric oxide synthase, induces accumulation of p53 and p21^WAF1/CIP1, and suppresses cultured bovine pulmonary artery endothelial cell proliferation by perturbing progression through S and G₂. Cancer Res. 1999; 59:2596–2601.

11. Kang W, Hong HJ, Guan J, Kim DG, Yang EJ, Koh G, Park D, Han CH, Lee YJ, Lee DH. Resveratrol improves insulin signaling in a tissue-specific manner under insulin-resistant conditions only: in vitro and in vivo experiments in rodents. Metabolism. 2012; 61:424–433.

12. Lagouge M, Argmann C, Gerhart-Hines Z, Meziane H, Lerin C, Daussin F, Messadeq N, Milne J, Lambert P, Elliott P, Geny B, Laakso M, Puigserver P, Auwerx J. Resveratrol improves mitochondrial function and protects against metabolic disease by activating SIRT1 and PGC-1α. Cell. 2006; 127:1109–1122.

13. Li X, Lu Y, Jin W, Liang K, Mills GB, Fan Z. Autophosphorylation of Akt at threonine 72 and serine 246. A potential mechanism of regulation of Akt kinase activity. J Biol Chem. 2006; 281:13837–13843.

14. McManus EJ, Sakamoto K, Armit LJ, Ronaldson L, Shpiro N, Marquez R, Alessi DR. Role that phosphorylation of GSK3 plays in insulin and Wnt signalling defined by knockin analysis. EMBO J. 2005; 24:1571–1583.

15. Pace-Asciak CR, Rounova O, Hahn SE, Diamandis EP, Goldberg DM. Wines and grape juices as modulators of platelet aggregation in healthy human subjects. Clin Chim Acta. 1996; 246:163–182.

16. Postic C, Girard J. Contribution of de novo fatty acid synthesis to hepatic steatosis and insulin resistance: lessons from genetically engineered mice. J Clin Invest. 2008; 118:829–838.

17. Renaud S, de Lorgeril M. Wine, alcohol, platelets, and the French paradox for coronary heart disease. Lancet. 1992; 339:1523–1526.

18. Sipula IJ, Brown NF, Perdomo G. Rapamycin-mediated inhibition of mammalian target of rapamycin in skeletal muscle cells reduces glucose utilization and increases fatty acid oxidation. Metabolism. 2006; 55:1637–1644.

19. Soleas GJ, Diamandis EP, Goldberg DM. The world of resveratrol. Adv Exp Med Biol. 2001; 492:159–182.

20. Thorburn AW, Gumbiner B, Bulacan F, Wallace P, Henry RR. Intracellular glucose oxidation and glycogen synthase activity are reduced in non-insulin-dependent (Type II) diabetes independent of impaired glucose uptake. J Clin Invest. 1990; 85:522–529.

21. Valenzano DR, Terzibasi E, Genade T, Cattaneo A, Domenici L, Cellerino A. Resveratrol prolongs lifespan and retards the onset of age-related markers in a short-lived vertebrate. Curr Biol. 2006; 16:296–300.

22. White MF. IRS proteins and the common path to diabetes. Am J Physiol Endocrinol Metab. 2002; 283:E413–E422.