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Metformin inhibits gluconeogenesis via a redox-dependent mechanism in vivo

An Author Correction to this article was published on 07 February 2019

This article has been updated

Abstract

Metformin, the universal first-line treatment for type 2 diabetes, exerts its therapeutic glucose-lowering effects by inhibiting hepatic gluconeogenesis. However, the primary molecular mechanism of this biguanide remains unclear, though it has been suggested to act, at least partially, by mitochondrial complex I inhibition. Here we show that clinically relevant concentrations of plasma metformin achieved by acute intravenous, acute intraportal or chronic oral administration in awake normal and diabetic rats inhibit gluconeogenesis from lactate and glycerol but not from pyruvate and alanine, implicating an increased cytosolic redox state in mediating metformin’s antihyperglycemic effect. All of these effects occurred independently of complex I inhibition, evidenced by unaltered hepatic energy charge and citrate synthase flux. Normalizing the cytosolic redox state by infusion of methylene blue or substrates that contribute to gluconeogenesis independently of the cytosolic redox state abrogated metformin-mediated inhibition of gluconeogenesis in vivo. Additionally, in mice expressing constitutively active acetyl-CoA carboxylase, metformin acutely decreased hepatic glucose production and increased the hepatic cytosolic redox state without altering hepatic triglyceride content or gluconeogenic enzyme expression. These studies demonstrate that metformin, at clinically relevant plasma concentrations, inhibits hepatic gluconeogenesis in a redox-dependent manner independently of reductions in citrate synthase flux, hepatic nucleotide concentrations, acetyl-CoA carboxylase activity, or gluconeogenic enzyme protein expression.

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Fig. 1: Acute treatment with 50 mg kg–1 metformin administered i.v. inhibits contributions from lactate but not alanine to hepatic gluconeogenesis in normal SD rats.
Fig. 2: Acute treatment with 50 mg kg–1 metformin administered i.v. inhibits contributions from lactate but not alanine to hepatic gluconeogenesis in ZDF rats.
Fig. 3: Acute treatment with 50 mg kg–1 metformin administered i.v. decreases fasting plasma glucose and inhibits EGP in the ACC DKI mouse model by modulating redox.
Fig. 4: Infusion of pyruvate and DHA and infusion of MB both abrogate the effects of acute i.v. treatment with 50 mg kg–1 metformin on plasma glucose and EGP in normal SD rats.
Fig. 5: Portal vein infusion of 50 mg kg–1 metformin acutely inhibits hepatic gluconeogenesis and selectively alters lactate, β-OHB and glycerol turnover and clearance rates without altering alanine turnover and alanine clearance rates or impacting mitochondrial VCS in a HFD–STZ SD rat model of T2D.
Fig. 6: Chronic oral metformin treatment (3.5 mg mL–1 in the drinking water for 14 d) specifically inhibits contributions from lactate but not alanine to hepatic gluconeogenesis without impacting mitochondrial VCS in a flux HFD–STZ SD rat model of T2D.

Change history

  • 07 February 2019

    In the version of this article originally published, the VPC and VCS flux data shown in Fig. 6e,f were inadvertently duplicated from Fig. 5j,k. The correct data are now shown in Fig. 6e,f. In these corrected data, VPC flux in response to chronic oral metformin treatment was still significantly decreased (Fig. 6e), and there was still no impact of metformin on VCS flux (Fig. 6f). Therefore, the text describing these data remains the same and this correction does not change the conclusion of this study.

References

  1. 1.

    Owen, M. R., Doran, E. & Halestrap, A. P. Evidence that metformin exerts its anti-diabetic effects through inhibition of complex 1 of the mitochondrial respiratory chain. Biochem. J. 348, 607–614 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. 2.

    El-Mir, M.-Y. et al. Dimethylbiguanide inhibits cell respiration via an indirect effect targeted on the respiratory chain complex I. J. Biol. Chem. 275, 223–228 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. 3.

    He, L. et al. Metformin and insulin suppress hepatic gluconeogenesis through phosphorylation of CREB binding protein. Cell 137, 635–646 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. 4.

    Cao, J. et al. Low concentrations of metformin suppress glucose production in hepatocytes through AMP-activated protein kinase (AMPK). J. Biol. Chem. 289, 20435–20446 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. 5.

    Fullerton, M. D. et al. Single phosphorylation sites in Acc1 and Acc2 regulate lipid homeostasis and the insulin-sensitizing effects of metformin. Nat. Med. 19, 1649–1654 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. 6.

    Cool, B. et al. Identification and characterization of a small molecule AMPK activator that treats key components of type 2 diabetes and the metabolic syndrome. Cell Metab. 3, 403–416 (2006).

    Article  CAS  Google Scholar 

  7. 7.

    Guigas, B. & Viollet, B. Targeting AMPK: from ancient drugs to new small-molecule activators. EXS 107, 327–350 (2016).

    CAS  PubMed  Google Scholar 

  8. 8.

    Rena, G., Hardie, D. G. & Pearson, E. R. The mechanisms of action of metformin. Diabetologia 60, 1577–1585 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. 9.

    Shaw, R. J. et al. The kinase LKB1 mediates glucose homeostasis in liver and therapeutic effects of metformin. Science 310, 1642–1646 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. 10.

    Smith, B. K. et al. Treatment of nonalcoholic fatty liver disease: role of AMPK. Am. J. Physiol. Endocrinol. Metab. 311, E730–E740 (2016).

    Article  Google Scholar 

  11. 11.

    Foretz, M. et al. Metformin inhibits hepatic gluconeogenesis in mice independently of the LKB1/AMPK pathway via a decrease in hepatic energy state. J. Clin. Invest. 120, 2355–2369 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. 12.

    Miller, R. A. et al. Biguanides suppress hepatic glucagon signalling by decreasing production of cyclic AMP. Nature 494, 256–260 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Johanns, M. et al. AMPK antagonizes hepatic glucagon-stimulated cyclic AMP signalling via phosphorylation-induced activation of cyclic nucleotide phosphodiesterase 4B. Nat. Commun. 7, 10856 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. 14.

    Hawley, S. A., Gadalla, A. E., Olsen, G. S. & Hardie, D. G. The antidiabetic drug metformin activates the AMP-activated protein kinase cascade via an adenine nucleotide-independent mechanism. Diabetes 51, 2420–2425 (2002).

    Article  CAS  Google Scholar 

  15. 15.

    Madiraju, A. K. et al. Metformin suppresses gluconeogenesis by inhibiting mitochondrial glycerophosphate dehydrogenase. Nature 510, 542–546 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Cederbaum, A. I., Lieber, C. S., Beattie, D. S. & Rubin, E. Characterization of shuttle mechanisms for the transport of reducing equivalents into mitochondria. Arch. Biochem. Biophys. 158, 763–781 (1973).

    Article  CAS  Google Scholar 

  17. 17.

    Kobayashi, K. et al. The gene mutated in adult-onset type II citrullinaemia encodes a putative mitochondrial carrier protein. Nat. Genet. 22, 159–163 (1999).

    Article  CAS  Google Scholar 

  18. 18.

    LaNoue, K. F. & Williamson, J. R. Interrelationships between malate–aspartate shuttle and citric acid cycle in rat heart mitochondria. Metabolism 20, 119–140 (1971).

    Article  CAS  Google Scholar 

  19. 19.

    Jomain-Baum, M. & Hanson, R. W. Regulation of hepatic gluconeogenesis in the guinea pig by fatty acids and ammonia. J. Biol. Chem. 250, 8978–8985 (1975).

    CAS  PubMed  Google Scholar 

  20. 20.

    Sugano, T. et al. Intracellular redox state and stimulation of gluconeogenesis by glucagon and norepinephrine in the perfused rat liver. J. Biochem. 87, 153–166 (1980).

    Article  CAS  Google Scholar 

  21. 21.

    Williamson, J. R., Scholz, R. & Browning, E. T. Control mechanisms of gluconeogenesis and ketogenesis. II Interactions between fatty acid oxidation and the citric acid cycle in perfused rat liver.J. Biol. Chem 244, 4617–4627 (1969).

    CAS  PubMed  Google Scholar 

  22. 22.

    Williamson, J. R., Scholz, R., Browning, E. T., Thurman, R. G. & Fukami, M. H. Metabolic effects of ethanol in perfused rat liver. J. Biol. Chem. 244, 5044–5054 (1969).

    CAS  PubMed  Google Scholar 

  23. 23.

    Perry, R. J. et al. Hepatic acetyl CoA links adipose tissue inflammation to hepatic insulin resistance and type 2 diabetes. Cell 160, 745–758 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. 24.

    Perry, R. J. et al. Leptin reverses diabetes by suppression of the hypothalamic–pituitary–adrenal axis. Nat. Med. 20, 759–763 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. 25.

    Sistare, F. D. & Haynes, R. C.Jr. The interaction between the cytosolic pyridine nucleotide redox potential and gluconeogenesis from lactate/pyruvate in isolated rat hepatocytes. Implications for investigations of hormone action.J. Biol. Chem. 260, 12748–12753 (1985).

    CAS  PubMed  Google Scholar 

  26. 26.

    Goodman, L. S. Goodman & Gilman's The Pharmacological Basis of Therapeutics, 12th edn (eds Brunton, L. L., Chabner, B. & Knollmann, B. C.) 1906–2000 (McGraw-Hill, New York, 2011).

  27. 27.

    Duca, F. A. et al. Metformin activates a duodenal Ampk-dependent pathway to lower hepatic glucose production in rats. Nat. Med. 21, 506–511 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. 28.

    Guigas, B. et al. 5-Aminoimidazole-4-carboxamide-1-β-d-ribofuranoside and metformin inhibit hepatic glucose phosphorylation by an AMP-activated protein kinase-independent effect on glucokinase translocation. Diabetes 55, 865–874 (2006).

    Article  CAS  Google Scholar 

  29. 29.

    Zhou, G. et al. Role of AMP-activated protein kinase in mechanism of metformin action. J. Clin. Invest. 108, 1167–1174 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. 30.

    Hundal, R. S. et al. Mechanism by which metformin reduces glucose production in type 2 diabetes. Diabetes 49, 2063–2069 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. 31.

    Inzucchi, S. E. et al. Efficacy and metabolic effects of metformin and troglitazone in type II diabetes mellitus. N. Engl. J. Med. 338, 867–872 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. 32.

    Chu, C. A. et al. The acute effect of metformin on glucose production in the conscious dog is primarily attributable to inhibition of glycogenolysis. Metabolism 49, 1619–1626 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. 33.

    Mithieux, G., Guignot, L., Bordet, J. C. & Wiernsperger, N. Intrahepatic mechanisms underlying the effect of metformin in decreasing basal glucose production in rats fed a high-fat diet. Diabetes 51, 139–143 (2002).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. 34.

    Lavine, J. E. et al. Effect of vitamin E or metformin for treatment of nonalcoholic fatty liver disease in children and adolescents: the TONIC randomized controlled trial. J. Am. Med. Assoc. 305, 1659–1668 (2011).

    Article  CAS  Google Scholar 

  35. 35.

    Rakoski, M. O., Singal, A. G., Rogers, M. A. & Conjeevaram, H. Meta-analysis: insulin sensitizers for the treatment of non-alcoholic steatohepatitis. Aliment. Pharmacol. Ther. 32, 1211–1221 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. 36.

    Shields, W. W., Thompson, K. E., Grice, G. A., Harrison, S. A. & Coyle, W. J. The effect of metformin and standard therapy versus standard therapy alone in nondiabetic patients with insulin resistance and nonalcoholic steatohepatitis (NASH): a pilot trial. Ther. Adv. Gastroenterol. 2, 157–163 (2009).

    Article  Google Scholar 

  37. 37.

    Gormsen, L. C. et al. Metformin does not affect postabsorptive hepatic free fatty acid uptake, oxidation or resecretion in humans: a 3-month placebo-controlled clinical trial in patients with type 2 diabetes and healthy controls. Diabetes Obes. Metab. https://doi.org/10.1111/dom.13244 (2018).

  38. 38.

    Ryle, P. R., Chakraborty, J. & Thomson, A. D. The effect of methylene blue on the hepatocellular redox state and liver lipid content during chronic ethanol feeding in the rat. Biochem. J. 232, 877–882 (1985).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Baur, J. A. & Birnbaum, M. J. Control of gluconeogenesis by metformin: does redox trump energy charge? Cell Metab. 20, 197–199 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    He, L. & Wondisford, F. E. Metformin action: concentrations matter. Cell Metab. 21, 159–162 (2015).

    Article  CAS  Google Scholar 

  41. 41.

    Puhakainen, I., Koivisto, V. A. & Yki-Järvinen, H. Lipolysis and gluconeogenesis from glycerol are increased in patients with noninsulin-dependent diabetes mellitus. J. Clin. Endocrinol. Metab. 75, 789–794 (1992).

    CAS  PubMed  Google Scholar 

  42. 42.

    Nurjhan, N., Consoli, A. & Gerich, J. Increased lipolysis and its consequences on gluconeogenesis in non-insulin-dependent diabetes mellitus. J. Clin. Invest. 89, 169–175 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. 43.

    Wu, H. et al. Metformin alters the gut microbiome of individuals with treatment-naive type 2 diabetes, contributing to the therapeutic effects of the drug. Nat. Med. 23, 850–858 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. 44.

    Schlienger, J. L., Frick, A., Marbach, J., Freund, H. & Imler, M. Effects of biguanides on the intermediate metabolism of glucose in normal and portal-strictured rats. Diabete Metab. 5, 5–9 (1979).

    CAS  PubMed  Google Scholar 

  45. 45.

    Gormsen, L. C. et al. In vivo imaging of human 11C-metformin in peripheral organs: dosimetry, biodistribution, and kinetic analyses. J. Nucl. Med. 57, 1920–1926 (2016).

    Article  CAS  Google Scholar 

  46. 46.

    Brønden, A. A. A., et al Single-dose metformin enhances bile acid–induced GLP-1 secretion in patients with type 2 diabetes. J. Clin. Endocrinol. Metab. (in the press) (2017).

  47. 47.

    Timmins, P., Donahue, S., Meeker, J. & Marathe, P. Steady-state pharmacokinetics of a novel extended-release metformin formulation. Clin. Pharmacokinet. 44, 721–729 (2005).

    Article  CAS  Google Scholar 

  48. 48.

    Larsen, S. et al. Metformin-treated patients with type 2 diabetes have normal mitochondrial complex I respiration. Diabetologia 55, 443–449 (2012).

    Article  CAS  Google Scholar 

  49. 49.

    Said, A. & Akhter, A. Meta-analysis of randomized controlled trials of pharmacologic agents in non-alcoholic steatohepatitis. Ann. Hepatol. 16, 538–547 (2017).

    Article  CAS  Google Scholar 

  50. 50.

    Sawangjit, R. et al. Comparative efficacy of interventions on nonalcoholic fatty liver disease (NAFLD): a PRISMA-compliant systematic review and network meta-analysis. Med. (Baltim.) 95, e4529 (2016).

    Article  CAS  Google Scholar 

  51. 51.

    Kim, C. W. et al. Acetyl CoA carboxylase inhibition reduces hepatic steatosis but elevates plasma triglycerides in mice and humans: a bedside to bench investigation. Cell Metab. 26, 576 (2017).

    Article  CAS  Google Scholar 

  52. 52.

    Konopka, A. R. et al. Hyperglucagonemia mitigates the effect of metformin on glucose production in prediabetes. Cell Rep. 15, 1394–1400 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. 53.

    Lewis, A. J.et al. Assessment of metformin-induced changes in cardiac and hepatic redox state using hyperpolarized [1-13C] pyruvate. Diabetes 65, 3544–3551 (2016).

    Article  CAS  Google Scholar 

  54. 54.

    Qi, H. et al. Acute renal metabolic effect of metformin assessed with hyperpolarised MRI in rats. Diabetologia 61, 445–454 (2018).

    Article  CAS  Google Scholar 

  55. 55.

    Nattrass, M., Todd, P. G., Hinks, L., Lloyd, B. & Alberti, K. G. Comparative effects of phenformin, metformin and glibenclamide on metabolic rhythms in maturity-onset diabetics. Diabetologia 13, 145–152 (1977).

    Article  CAS  Google Scholar 

  56. 56.

    Hussey, E. K. et al. Safety, pharmacokinetics and pharmacodynamics of remogliflozin etabonate, a novel SGLT2 inhibitor, and metformin when co-administered in subjects with type 2 diabetes mellitus. BMC Pharmacol. Toxicol. 14, 25 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. 57.

    Comte, B., Vidal, H., Laville, M. & Riou, J. P. Influence of thyroid hormones on gluconeogenesis from glycerol in rat hepatocytes: a dose-response study. Metabolism 39, 259–263 (1990).

    Article  CAS  Google Scholar 

  58. 58.

    Gregory, R. B., Phillips, J. W. & Berry, M. N. Reducing-equivalent transfer to the mitochondria during gluconeogenesis and ureogenesis in hepatocytes from rats of different thyroid status. Biochim. Biophys. Acta 1137, 34–38 (1992).

    Article  CAS  Google Scholar 

  59. 59.

    Kneer, N. & Lardy, H. Thyroid hormone and dehydroepiandrosterone permit gluconeogenic hormone responses in hepatocytes. Arch. Biochem. Biophys. 375, 145–153 (2000).

    Article  CAS  Google Scholar 

  60. 60.

    Drahota, Z. et al. Biguanides inhibit complex I, II and IV of rat liver mitochondria and modify their functional properties. Physiol. Res. 63, 1–11 (2014).

    CAS  PubMed  Google Scholar 

  61. 61.

    Reed, M. J. et al. Effect of masoprocol on carbohydrate and lipid metabolism in a rat model of type II diabetes. Diabetologia 42, 102–106 (1999).

    Article  CAS  Google Scholar 

  62. 62.

    Perry, R. J. et al. Non-invasive assessment of hepatic mitochondrial metabolism by positional isotopomer NMR tracer analysis (PINTA). Nat. Commun. 8, 798 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. 63.

    Perry, R. J. et al. Reversal of hypertriglyceridemia, fatty liver disease, and insulin resistance by a liver-targeted mitochondrial uncoupler. Cell Metab. 18, 740–748 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. 64.

    Alves, T. C. et al. Regulation of hepatic fat and glucose oxidation in rats with lipid-induced hepatic insulin resistance. Hepatology 53, 1175–1181 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  65. 65.

    Perry, R. J. et al. Mechanism for leptin’s acute insulin-independent effect to reverse diabetic ketoacidosis. J. Clin. Invest. 127, 657–669 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  66. 66.

    Ayala, J. E. et al. Standard operating procedures for describing and performing metabolic tests of glucose homeostasis in mice. Dis. Model. Mech. 3, 525–534 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  67. 67.

    Passonneau, J. V. & Lauderdale, V. R. A comparison of three methods of glycogen measurement in tissues. Anal. Biochem. 60, 405–412 (1974).

    Article  CAS  Google Scholar 

  68. 68.

    Bligh, E. G. & Dyer, W. J. A rapid method of total lipid extraction and purification. Can. J. Biochem. Physiol. 37, 911–917 (1959).

    Article  CAS  Google Scholar 

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Acknowledgements

The authors would like to thank J. Dong, M. Kahn, S. Dufour, J. Stack, Y.Kosover, A. Nasiri, X. Ma, W. Zhu, and K. Harry for their technical support and V. Samuel, S. Caprio, and D. Kibbey for helpful discussions. This publication was supported by grants from the US Department of Health and Human Services: R01 DK113984, P30 DK45735, P30 DK034989, K99 CA215315 and R01 DK114793. Its contents are solely the responsibility of the authors and do not necessarily represent the official view of NCRR or the NIH. G.R.S. is supported by grants and fellowships from a Canada Research Chair in Metabolism and Obesity, the J. Bruce Duncan Endowed Chair in Metabolic Diseases at McMaster University and Diabetes Canada. B.E.K. is supported by grants and a Fellowship (BEK) from the National Health and Medical Research Council (1068813 and 1085460) and the Victorian Government Operational Infrastructure Support Scheme.

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A.K.M., Y.Q., R.J.P., J.-P.G.C., D.F.V., K.F.P., and G.I.S. designed the experimental protocols. A.K.M., Y.Q., Y.R., X.-M.Z., D.Z., G.W.C., G.M.B., G.C., J.-P.G.C., and K.F.P. performed the studies. A.K.M., Y.Q., R.J.P., X-M. Z., G.W.C., G.C., and J.-P.G.C. analyzed the data. B.E.K. and G.R.S. supplied reagents.A.K.M., Y.Q., R.J.P, G.W.C., D.F.V., K.F.P., and G.I.S. wrote the manuscript with contributions from all of the other authors.

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Madiraju, A.K., Qiu, Y., Perry, R.J. et al. Metformin inhibits gluconeogenesis via a redox-dependent mechanism in vivo. Nat Med 24, 1384–1394 (2018). https://doi.org/10.1038/s41591-018-0125-4

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