Metformin is the most commonly prescribed medication for type 2 diabetes, owing to its glucose-lowering effects, which are mediated through the suppression of hepatic glucose production (reviewed in refs. 1,2,3). However, in addition to its effects on the liver, metformin reduces appetite and in preclinical models exerts beneficial effects on ageing and a number of diverse diseases (for example, cognitive disorders, cancer, cardiovascular disease) through mechanisms that are not fully understood1,2,3. Given the high concentration of metformin in the liver and its many beneficial effects beyond glycemic control, we reasoned that metformin may increase the secretion of a hepatocyte-derived endocrine factor that communicates with the central nervous system4. Here we show, using unbiased transcriptomics of mouse hepatocytes and analysis of proteins in human serum, that metformin induces expression and secretion of growth differentiating factor 15 (GDF15). In primary mouse hepatocytes, metformin stimulates the secretion of GDF15 by increasing the expression of activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP; also known as DDIT3). In wild-type mice fed a high-fat diet, oral administration of metformin increases serum GDF15 and reduces food intake, body mass, fasting insulin and glucose intolerance; these effects are eliminated in GDF15 null mice. An increase in serum GDF15 is also associated with weight loss in patients with type 2 diabetes who take metformin. Although further studies will be required to determine the tissue source(s) of GDF15 produced in response to metformin in vivo, our data indicate that the therapeutic benefits of metformin on appetite, body mass and serum insulin depend on GDF15.
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The authors would like to thank A. Božović and V. Kulasingam for measuring serum metformin levels. E.A.D. was a recipient of an Ontario Graduate Scholarship (Queen Elizabeth II Graduate Scholarship in Science and Technology) and a Douglas C. Russell Memorial Scholarship. G.R.S. is a Canada Research Chair and the J. Bruce Duncan Chair in Metabolic Diseases. This study was supported by research grants from the Canadian Institutes of Health Research (201709FDN-CEBA-116200 to G.R.S.) and Diabetes Canada (DI-5-17-5302-GS). We thank Sanofi for providing heterozygous breeding pairs of GDF15-null mice.
S.H., G.P., H.C.G. and G.R.S. hold a patent entitled ‘Growth differentiation factor 15 as biomarker for metformin’ (WO/2017/108941). S.H. and M.K. are employees of Sanofi. All other authors have no competing interests.
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Extended Data Fig. 1 Metformin, phenformin and buformin increase GDF15 release independent of complex 1 inhibition or AMPK.
a, Volcano plot showing differentially regulated genes after 24 h of metformin treatment (metformin versus control, n = 4 per group). b,c, GDF15 release is stimulated in a dose-dependent manner by two biguanides that are structurally similar to metformin: (b) phenformin (0 μM n = 4, 10 μM n = 3, 30 μM n = 3 and 50 μM n = 4) and (c) buformin (0 μM n = 3, 10 μM n = 2, 30 μM n = 2 and 50 μM n = 3). d, The complex I inhibitor rotenone (0 μM, 0.1 μM, 1 μM, 5 μM n = 3) does not increase GDF15 release. e, Metformin increases GDF15 release in primary hepatocytes from WT (control n = 7, metformin n = 7), AMPK β1KO (control n = 4, metformin n = 3) and ACC DKI (control n = 3, metformin n = 3) mice. Data are presented as mean ± s.e.m. For b–d, * indicates P < 0.05 and *** indicates P < 0.001 for one-way ANOVA with Sidak multiple comparison test. For e, * indicates P < 0.05 for two-way ANOVA with Sidak multiple comparison test.
a, Serum metformin 1 h after acute saline (n = 4) or metformin gavage (n = 7). *P < 0.05, for unpaired two-sided t-test. b, Serum GLP-1 10 min after acute metformin gavage in wild-type (n = 8) and GDF15-KO (n = 6) mice fed a 45% HFD. c–f, Wild-type (control n = 9, metformin n = 9) and GDF15-KO (control n = 6, metformin n = 6) mice were fed a 45% HFD, placed in metabolic cages, and allowed to acclimatize for approximately 24 h before a single oral gavage of metformin (250 mg kg–1) or appropriate volume of saline 2 h before the onset of the dark period. RER, beam breaks and energy expenditure were measured for 24 h after gavage, and data are presented as mean ± s.e.m. *P < 0.05 between control and metformin, two-way ANOVA with Sidak multiple comparison test.
Extended Data Fig. 3 Chronic metformin treatment does not alter lean mass, RER or energy expenditure.
Wild-type (control n = 8, metformin n = 9) and GDF15-KO (control n = 8, metformin n = 8) mice were fed a 45% HFD for 4 weeks prior to being switched to control (tap water) or metformin water (3 g l−1) for 10 weeks. a,b, Body composition was assessed at week 4 of treatment (wild-type control n = 8, wild-type metformin n = 9, GDF15-KO control n = 8 and GDF15-KO metformin n = 8). c–i, Wild-type (control n = 7, metformin n = 8) and GDF15-KO (control n = 8, metformin n = 8) mice were placed in metabolic cages and allowed to acclimatize for approximately 24 h. Food intake, activity, RER and energy expenditure were measured over 48 h. Energy expenditure is shown (e) uncorrected, (f,g) corrected for body mass, and (h,i) corrected for lean mass, and data are presented as mean ± s.e.m.
Extended Data Fig. 4 Metformin in drinking water elicits clinically relevant serum metformin levels.
Wild-type and GDF15-KO mice were fed a 45% HFD for 4 weeks prior to being switched to control (tap water) or metformin water (3 g l−1) for 10 weeks. Wild-type (control n = 6, metformin n = 8) and GDF15-KO (control n = 6, metformin n = 7) mice were placed in metabolic cages and allowed to acclimatize for approximately 24 h. a, Water intake was measured over 48 h. b, Metformin dose was calculated based on water intake and body mass. c, Serum metformin was measured after mice were killed at the onset of the light period. d, Food intake of 45% HFD was monitored every 3–4 days in mice fed ad libitum (n = 9), mice fed ad libitum treated with metformin (n = 10), and mice pair-fed with metformin-treated animals (n = 10). e, Serum GDF15 was assessed at 3 weeks. Data are presented as mean ± s.e.m. *P < 0.05, **P < 0.005, ***P < 0.001 between control and metformin, two-way ANOVA with Sidak multiple comparison test (a–c), one-way ANOVA with Sidak multiple comparison test (d,e).
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Day, E.A., Ford, R.J., Smith, B.K. et al. Metformin-induced increases in GDF15 are important for suppressing appetite and promoting weight loss. Nat Metab 1, 1202–1208 (2019). https://doi.org/10.1038/s42255-019-0146-4
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