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The mitochondrial calcium uniporter (MCU) facilitates mitochondrial calcium influx, and several enzymes that catalyze key steps of the tricarboxylic acid (TCA) cycle require calcium for their activation. As such, it is generally accepted that rates of mitochondrial oxidation correlate with mitochondrial calcium activity. Decreased rates of hepatic mitochondrial oxidation are associated with metabolic dysfunction and ectopic lipid accumulation as seen with type 2 diabetes and nonalcoholic fatty liver disease. Despite this, a role for MCU and mitochondrial calcium in regulating hepatic mitochondrial oxidation is still unclear. To address this question, we generated a liver-specific MCU knockout (MCU KO) mouse model which displayed significantly reduced mitochondrial calcium ([Ca2+]mt, p Disclosure T. E. Lamoia: n/a. B. T. Hubbard: None. M. Guerra: None. R. Goodman: None. M. H. Nathanson: None. V. Mootha: Advisory Panel; 5AM Ventures, LLC, Janssen Research & Development, LLC. G. I. Shulman: Advisory Panel; 89bio, Inc., AstraZeneca, Equator Therapeutics, Inc., Janssen Research & Development, LLC, Merck & Co., Inc., Consultant; DiCerna Pharmaceuticals, Inc. , Novo Nordisk, Other Relationship; Generian Pharmaceuticals, iMetabolic Biopharma Corporation, Maze Therapeutics, The Liver Company, Stock/Shareholder; Levels Health, Inc. . Funding F31 DK126362/DK/NIDDK NIH HHST32 GM007324/GM/NIGMS NIH HHS

Metformin is a first-line therapy for the treatment of type 2 diabetes, due to its robust glucose-lowering effects, well-established safety profile, and relatively low cost. While metformin has been shown to have pleotropic effects on glucose metabolism, there is a general consensus that the major glucose-lowering effect in patients with type 2 diabetes is mostly mediated through inhibition of hepatic gluconeogenesis. However, despite decades of research, the mechanism by which metformin inhibits this process is still highly debated. A key reason for these discrepant effects is likely due to the inconsistency in dosage of metformin across studies. Widely studied mechanisms of action, such as complex I inhibition leading to AMPK activation, have only been observed in the context of supra-pharmacological (>1 mM) metformin concentrations, which do not occur in the clinical setting. Thus, these mechanisms have been challenged in recent years and new mechanisms have been proposed. Based on the observation that metformin alters cellular redox balance, a redox-dependent mechanism of action has been described by several groups. Recent studies have shown that clinically relevant (50-100 μM) concentrations of metformin inhibit hepatic gluconeogenesis in a substrate-selective manner both in vitro and in vivo, supporting a redox-dependent mechanism of metformin action. Here, we review the current literature regarding metformin’s cellular and molecular mechanisms of action., Graphical Abstract Graphical Abstract

Significance Metformin is the most commonly prescribed drug for the treatment of type 2 diabetes mellitus, yet the mechanism by which it lowers plasma glucose concentrations has remained elusive. Most studies to date have attributed metformin’s glucose-lowering effects to inhibition of complex I activity. Contrary to this hypothesis, we show that inhibition of complex I activity in vitro and in vivo does not reduce plasma glucose concentrations or inhibit hepatic gluconeogenesis. We go on to show that metformin, and the related guanides/biguanides, phenformin and galegine, inhibit complex IV activity at clinically relevant concentrations, which, in turn, results in inhibition of glycerol-3-phosphate dehydrogenase activity, increased cytosolic redox, and selective inhibition of glycerol-derived hepatic gluconeogenesis both in vitro and in vivo.

The mammalian succinate dehydrogenase (SDH) complex has recently been shown as capable of operating bidirectionally. Here, we develop a method (Q-Flux) capable of measuring absolute rates of both forward (V

The transmembrane protein, cluster of differentiation 36 (CD36), facilitates tissue fatty acid uptake and is highly expressed in a variety of different cell types, including endothelial cells (ECs). Loss of EC Cd36 reduces parenchymal long-chain fatty acid uptake and improves whole-body glucose tolerance and insulin sensitivity; however, the mechanisms by which this occurs remain unclear. Here, we report that EC-specific deletion of Cd36 in chow-fed male mice (EC-Cd36-/-) leads to significant reductions in whole-body fat utilization during fasting, subsequently increasing plasma non-esterified fatty acid levels by 30% (P Conclusion: Taken together, these results demonstrate a key role for EC Cd36 in regulating parenchymal fuel selection and hepatic glucose production and provide insight into the mechanisms by which EC Cd36 controls systemic glucose and lipid metabolism. Disclosure L. Goedeke: None. N. Son: None. T. E. Lamoia: None. A. Nasiri: Employee; Spouse/Partner; Medtronic. M. Kahn: None. X. Zhang: None. G. Cline: None. I. J. Goldberg: Advisory Panel; Self; Akcea Therapeutics, Arrowhead Pharmaceuticals, Inc., Consultant; Self; Esperion. G. I. Shulman: Consultant; Self; 89bio, Inc., BridgeBio, Ionis Pharmaceuticals, Maze Therapeutics, Novo Nordisk, Other Relationship; Self; AstraZeneca, Esperion Therapeutics, Inc, Generian Pharmaceuticals, Inc., Gilead Sciences, Inc., iMetabolic Biopharma Corporation, Janssen Research & Development, LLC, Merck & Co., Inc., The Liver Company. Funding National Institutes of Health (HL150234, R01DK113984, R01DK045735)

Background Non-caloric artificial sweeteners (NCAS) are widely used as a substitute for dietary sugars to control body weight or glycemia. Paradoxically, some interventional studies in humans and rodents have shown unfavorable changes in glucose homeostasis in response to NCAS consumption. The causative mechanisms are largely unknown, but adverse changes in gut microbiota have been proposed to mediate these effects. These findings have raised concerns about NCAS safety and called into question their broad use, but further physiological and dietary considerations must be first addressed before these results are generalized. We also reasoned that, since NCAS are bona fide ligands for sweet taste receptors (STRs) expressed in the intestine, some metabolic effects associated with NCAS use could be attributed to a common mechanism involving the host. Results We conducted a double-blind, placebo-controlled, parallel arm study exploring the effects of pure saccharin compound on gut microbiota and glucose tolerance in healthy men and women. Participants were randomized to placebo, saccharin, lactisole (STR inhibitor), or saccharin with lactisole administered in capsules twice daily to achieve the maximum acceptable daily intake for 2 weeks. In parallel, we performed a 10-week study administering pure saccharin at a high dose in the drinking water of chow-fed mice with genetic ablation of STRs (T1R2-KO) and wild-type (WT) littermate controls. In humans and mice, none of the interventions affected glucose or hormonal responses to an oral glucose tolerance test (OGTT) or glucose absorption in mice. Similarly, pure saccharin supplementation did not alter microbial diversity or composition at any taxonomic level in humans and mice alike. No treatment effects were also noted in readouts of microbial activity such as fecal metabolites or short-chain fatty acids (SCFA). However, compared to WT, T1R2-KO mice were protected from age-dependent increases in fecal SCFA and the development of glucose intolerance. Conclusions Short-term saccharin consumption at maximum acceptable levels is not sufficient to alter gut microbiota or induce glucose intolerance in apparently healthy humans and mice. Trial registration Trial registration number NCT03032640, registered on January 26, 2017.

The mitochondrial calcium uniporter (MCU) is a key regulator of mitochondrial calcium, however, the role of MCU in mediating hepatic mitochondrial metabolism and gluconeogenesis has received limited attention. In order to address these questions, we examined rates of mitochondrial oxidation in both liver slices and in awake liver-specific MCU knockout (MCU KO) mice. We found that deletion of MCU significantly increases glucagon-induced intrahepatic lipolysis (p In conclusion, these results demonstrate a critical role for MCU in mediating hepatic mitochondrial oxidation and hepatic gluconeogenesis. Disclosure T.E. LaMoia: None. J. Lee: None. M. Guerra: None. R. Goodman: None. G.I. Shulman: Advisory Panel; Self; AstraZeneca, Janssen Research & Development, LLC, Merck & Co., Inc. Advisory Panel; Spouse/Partner; Merck & Co., Inc. Consultant; Self; Novo Nordisk A/S. Consultant; Spouse/Partner; Novo Nordisk A/S. Other Relationship; Self; Gilead Sciences, Inc., iMetabolic Biopharma Corporation. Other Relationship; Spouse/Partner; iMetabolic Biopharma Corporation. Other Relationship; Self; Maze Therapeutics. Funding National Institutes of Health (T32GM0007324)

Background : Non-caloric artificial sweeteners (NCAS) are widely used as a substitute for dietary sugars to control body weight or glycemia. Paradoxically, saccharin and other NCAS have been reported to induce glucose intolerance in mice fed a high-fat diet and in a subset of humans by directly inducing unfavorable changes in gut microbiota. These findings have raised concerns about NCAS and called into question their broad use. Whether these results can be generalized to healthy populations consuming conventional diets is unknown. It is also unclear how different NCAS, that do not share a common chemical structure, can produce identical direct effects on gut microbiota. A common feature of all NCAS is their strong affinity for sweet taste receptors (STRs) which are expressed in the intestine. However, their role in mediating NCAS-induced effects has not been addressed. Results : We conducted a double-blind, placebo-controlled, parallel arm study exploring the effects of saccharin on gut microbiota and glucose tolerance in healthy men and women. Participants were randomized to placebo, saccharin, lactisole (STR inhibitor), or saccharin with lactisole administered in capsules twice daily to achieve the maximum acceptable daily intake for two weeks. In parallel, we performed a ten-week study administering high-dose saccharin in the drinking water of chow-fed mice with genetic ablation of STRs (T1R2-KO) and wild-type (WT) littermate controls. In humans and mice alike, none of the interventions affected glucose or hormonal responses to a glucose tolerance test, nor ex vivo glucose absorption in mice. Similarly, saccharin supplementation did not alter microbial diversity or abundance at any taxonomic level in humans or mice. No treatment effects were also noted in readouts of microbial activity such as fecal metabolites or short chain fatty acids (SCFA). However, compared to WT, T1R2-KO mice were protected from age-dependent increases in fecal SCFA and the development of glucose intolerance.

Objective Beyond the taste buds, sweet taste receptors (STRs; T1R2/T1R3) are also expressed on enteroendocrine cells, where they regulate gut peptide secretion but their regulatory function within the intestine is largely unknown. Methods Using T1R2-knock out (KO) mice we evaluated the role of STRs in the regulation of glucose absorption in vivo and in intact intestinal preparations ex vivo. Results STR signaling enhances the rate of intestinal glucose absorption specifically in response to the ingestion of a glucose-rich meal. These effects were mediated specifically by the regulation of GLUT2 transporter trafficking to the apical membrane of enterocytes. GLUT2 translocation and glucose transport was dependent and specific to glucagon-like peptide 2 (GLP-2) secretion and subsequent intestinal neuronal activation. Finally, high-sucrose feeding in wild-type mice induced rapid downregulation of STRs in the gut, leading to reduced glucose absorption. Conclusions Our studies demonstrate that STRs have evolved to modulate glucose absorption via the regulation of its transport and to prevent the development of exacerbated hyperglycemia due to the ingestion of high levels of sugars., Highlights • The intestinal T1R2 receptor enhances glucose absorption in vivo and ex vivo. • Pharmacological inhibition of STRs reduces glucose flux in human intestinal preparations. • T1R2 regulates glucose absorption dependent on GLUT2 activity in enterocytes. • GLP-2 mediates the effects of T1R2 signaling through activation of enteric neurons. • High sucrose diet rapidly downregulates STRs leading to reduced glucose absorption.