Extracellular Visfatin Activates Gluconeogenesis in HepG2 Cells Through the Classical PKA/CREB-Dependent Pathway

Abstract

Adipokines reportedly affect hepatic gluconeogenesis, and the adipokine visfatin is known to be related to insulin resistance and type 2 diabetes. However, whether visfatin contributes to hepatic gluconeogenesis remains unclear. Visfatin, also known as nicotinamide phosphoribosyltransferase (NAMPT), modulates sirtuin1 (SIRT1) through the regulation of nicotinamide adenine dinucleotide (NAD). Therefore, we investigated the effect of extracellular visfatin on glucose production in HepG2 cells, and evaluated whether extracellular visfatin affects hepatic gluconeogenesis via an NAD⁺-SIRT1-dependent pathway. Treatment with visfatin significantly increased glucose production and the mRNA expression and protein levels of phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) in HepG2 cells in a time- and concentration-dependent manner. Knockdown of SIRT1 had no remarkable effect on the induction of gluconeogenesis by visfatin. Subsequently, we evaluated if extracellular visfatin stimulates the production of gluconeogenic enzymes through the classical protein kinase A (PKA)/cyclic AMP-responsive element (CRE)-binding protein (CREB)-dependent process. The phosphorylation of CREB and PKA increased significantly in HepG2 cells treated with visfatin. Additionally, knockdown of CREB and PKA inhibited visfatin-induced gluconeogenesis in HepG2 cells. In summary, extracellular visfatin modulates glucose production in HepG2 cells through the PKA/CREB pathway, rather than via SIRT1 signaling.

Key words

visfatin - gluconeogenesis - insulin resistance

^* These authors contributed equally to this work

• References

• 1 Samal B, Sun Y, Stearns G, Xie C, Suggs S, McNiece I. Cloning and characterization of the cDNA encoding a novel human pre-B-cell colony-enhancing factor. Mol Cell Biol 1994; 14: 1431-1437
• 2 Revollo JR, Körner A, Mills KF, Satoh A, Wang T, Garten A, Dasgupta B, Sasaki Y, Wolberger C, Townsend RR, Milbrandt J, Kiess W, Imai S. Nampt/PBEF/visfatin regulates insulin secretion in beta cells as a systemic NAD biosynthetic enzyme. Cell Metab 2007; 6: 363-375
• 3 Dahl TB, Holm S, Aukrust P, Halvorsen B. Visfatin/NAMPT: A Multifaceted Molecule with Diverse Roles in Physiology and Pathophysiology. Annu Rev Nutr 2012; 32: 229-243
• 4 Rasouli N, Kern PA. Adipocytokines and the metabolic complications of obesity. J Clin Endocrinol Metab 2008; 93 (Suppl. 01) S64-S73
• 5 Aller R, de Luis DA, Izaola O, Sagrado MG, Conde R, Velasco MC, Alvarez T, Pacheco D, González JM. Influence of visfatin on histopathological changes of non-alcoholic fatty liver disease. Dig Dis Sci 2009; 54: 1772-1777
• 6 Garten A, Petzold S, Barnikol-Oettler A, Körner A, Thasler WE, Kratzsch J, Kiess W, Gebhardt R. Nicotinamide phosphoribosyltransferase (NAMPT/PBEF/visfatin) is constitutively released from human hepatocytes. Biochem Biophys Res Commun 2010; 391: 376-381
• 7 Lee WJ, Wu CS, Lin H, Lee IT, Wu CM, Tseng JJ, Chou MM, Sheu WH. Visfatin-induced expression of inflammatory mediators in human endothelial cells through the NF-kappaB pathway. Int J Obes (Lond) 2009; 33: 465-472
• 8 Dahl TB, Yndestad A, Skjelland M, Øie E, Dahl A, Michelsen A, Damås JK, Tunheim SH, Ueland T, Smith C, Bendz B, Tonstad S, Gullestad L, Frøland SS, Krohg-Sørensen K, Russell D, Aukrust P, Halvorsen B. Increased expression of visfatin in macrophages of human unstable carotid and coronary atherosclerosis: possible role in inflammation and plaque destabilization. Circulation 2007; 115: 972-980
• 9 Laudes M, Oberhauser F, Schulte DM, Freude S, Bilkovski R, Mauer J, Rappl G, Abken H, Hahn M, Schulz O, Krone W. Visfatin/PBEF/Nampt and resistin expressions in circulating blood monocytes are differentially related to obesity and type 2 diabetes in humans. Horm Metab Res 2010; 42: 268-273
• 10 Zhong M, Tan HW, Gong HP, Wang SF, Zhang Y, Zhang W. Increased serum visfatin in patients with metabolic syndrome and carotid atherosclerosis. Clin Endocrinol (Oxf) 2008; 69: 878-884
• 11 Retnakaran R, Youn BS, Liu Y, Hanley AJ, Lee NS, Park JW, Song ES, Vu V, Kim W, Tungtrongchitr R, Havel PJ, Swarbrick MM, Shaw C, Sweeney G. Correlation of circulating full-length visfatin (PBEF/NAMPT) with metabolic parameters in subjects with and without diabetes: a cross-sectional study. Clin Endocrinol (Oxf.) 2008; 69: 885-893
• 12 Rabe K, Lehrke M, Parhofer KG, Broedl UC. Adipokines and insulin resistance. Mol Med 2008; 14: 741-751
• 13 Zhou H, Song X, Briggs M, Violand B, Salsgiver W, Gulve EA, Luo Y. Adiponectin represses gluconeogenesis independent of insulin in hepatocytes. Biochem Biophys Res Commun 2005; 338: 793-799
• 14 Miller RA, Chu Q, Le Lay J, Scherer PE, Ahima RS, Kaestner KH, Foretz M, Viollet B, Birnbaum MJ. Adiponectin suppresses gluconeogenic gene expression in mouse hepatocytes independent of LKB1-AMPK signaling. J Clin Invest 2011; 121: 2518-2528
• 15 Anderwald C, Müller G, Koca G, Fürnsinn C, Waldhäusl W, Roden M. Short-term leptin-dependent inhibition of hepatic gluconeogenesis is mediated by insulin receptor substrate-2. Mol Endocrinol 2002; 16: 1612-1628
• 16 Chang YH, Chang DM, Lin KC, Shin SJ, Lee YJ. Visfatin in overweight/obesity, type 2 diabetes mellitus, insulin resistance, metabolic syndrome and cardiovascular diseases: a meta-analysis and systemic review. Diabetes Metab Res Rev 2011; 27: 515-527
• 17 Rodgers JT, Lerin C, Haas W, Gygi SP, Spiegelman BM, Puigserver P. Nutrient control of glucose homeostasis through a complex of PGC-1alpha and SIRT1. Nature 2005; 434: 113-118
• 18 Zhang T, Berrocal JG, Frizzell KM, Gamble MJ, DuMond ME, Krishnakumar R, Yang T, Sauve AA, Kraus WL. Enzymes in the NAD+salvage pathway regulate SIRT1 activity at target gene promoters. J Biol Chem 2009; 284: 20408-20417
• 19 Herzig S, Long F, Jhala US, Hedrick S, Quinn R, Bauer A, Rudolph D, Schutz G, Yoon C, Puigserver P, Spiegelman B, Montminy M. CREB regulates hepatic gluconeogenesis via the co-activator PGC-1. Nature 2001; 413: 179-183
• 20 Ozcan L, Wong CC, Li G, Xu T, Pajvani U, Park SK, Wronska A, Chen BX, Marks AR, Fukamizu A, Backs J, Singer HA, Yates 3rd JR, Accili D, Tabas I. Calcium Signaling through CaMKII Regulates Hepatic Glucose Production in Fasting and Obesity. Cell Metab 2012; 15: 739-751
• 21 Cao W, Collins QF, Becker TC, Robidoux J, Lupo Jr EG, Xiong Y, Daniel KW, Floering L, Collins S. p38 Mitogen-activated protein kinase plays a stimulatory role in hepatic gluconeogenesis. J Biol Chem 2005; 280: 42731-42737
• 22 Asada S, Daitoku H, Matsuzaki H, Saito T, Sudo T, Mukai H, Iwashita S, Kako K, Kishi T, Kasuya Y, Fukamizu A. Mitogen-activated protein kinases, Erk and p38, phosphorylate and regulate Foxo1. Cell Signal 2007; 19: 519-527
• 23 Mlinar B, Marc J, Janez A, Pfeifer M. Molecular mechanisms of insulin resistance and associated diseases. Clin Chim Acta 2007; 375: 20-35
• 24 Esteghamati A, Alamdari A, Zandieh A, Elahi S, Khalilzadeh O, Nakhjavani M, Meysamie A. Serum visfatin is associated with type 2 diabetes mellitus independent of insulin resistance and obesity. Diabetes Res Clin Pract 2011; 91: 154-158
• 25 Jiang G, Zhang BB. Glucagon and regulation of 3 glucose metabolism. Am J Physiol Endocrinol Metab 2003; 284: E671-E678