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Monoacylglycerol O-acyltransferase 1 (Mogat1), a lipogenic enzyme that converts monoacylglycerol to diacylglycerol, is highly expressed in adipocytes and may regulate lipolysis by re-esterifying fatty acids released during times when lipolytic rates are low. However, the role of Mogat1 in regulating adipocyte fat storage during differentiation and diet-induced obesity is relatively understudied.Here, adipocyte-specific Mogat1 knockout mice were generated and subjected to a high-fat diet to determine the effects of Mogat1 deficiency on diet-induced obesity. Mogat1 floxed mice were also used to develop preadipocyte cell lines wherein Mogat1 could be conditionally knocked out to study adipocyte differentiation in vitro.In preadipocytes, it was found that Mogat1 knockout at the onset of preadipocyte differentiation prevented the accumulation of glycerolipids and reduced the differentiation capacity of preadipocytes. However, the loss of adipocyte Mogat1 did not affect weight gain or fat mass induced by a high-fat diet in mice. Furthermore, loss of Mogat1 in adipocytes did not affect plasma lipid or glucose concentrations or insulin tolerance.These data suggest Mogat1 may play a role in adipocyte differentiation in vitro but not adipose tissue expansion in response to nutrient overload in mice.

Pyruvate metabolism, a central nexus of carbon homeostasis, is an evolutionarily conserved process and aberrant pyruvate metabolism is associated with and contributes to numerous human metabolic disorders including diabetes, cancer, and heart disease. As a product of glycolysis, pyruvate is primarily generated in the cytosol before being transported into the mitochondrion for further metabolism. Pyruvate entry into the mitochondrial matrix is a critical step for efficient generation of reducing equivalents and ATP and for the biosynthesis of glucose, fatty acids, and amino acids from pyruvate. However, for many years, the identity of the carrier protein(s) that transported pyruvate into the mitochondrial matrix remained a mystery. In 2012, the molecular-genetic identification of the mitochondrial pyruvate carrier (MPC), a heterodimeric complex composed of protein subunits MPC1 and MPC2, enabled studies that shed light on the many metabolic and physiological processes regulated by pyruvate metabolism. A better understanding of the mechanisms regulating pyruvate transport and the processes affected by pyruvate metabolism may enable novel therapeutics to modulate mitochondrial pyruvate flux to treat a variety of disorders. Herein, we review our current knowledge of the MPC, discuss recent advances in the understanding of mitochondrial pyruvate metabolism in various tissue and cell types, and address some of the outstanding questions relevant to this field.

Dysfunctional adipose tissue is believed to promote the development of hepatic steatosis and systemic insulin resistance, but many of the mechanisms involved are still unclear. Lipin 1 catalyzes the conversion of phosphatidic acid to diacylglycerol (DAG), the penultimate step of triglyceride synthesis, which is essential for lipid storage. Herein we found that adipose tissueLPIN1expression is decreased in people with obesity compared to lean subjects and lowLPIN1expression correlated with multi-tissue insulin resistance and increased rates of hepatic de novo lipogenesis. Comprehensive metabolic and multi-omic phenotyping demonstrated that adipocyte-specificLpin1-/-mice had a metabolically-unhealthy phenotype, including liver and skeletal muscle insulin resistance, hepatic steatosis, increased hepatic de novo lipogenesis, and transcriptomic signatures of nonalcoholic steatohepatitis that was exacerbated by high-fat diets. We conclude that adipocyte lipin 1-mediated lipid storage is vital for preserving adipose tissue and systemic metabolic health and its loss predisposes mice to nonalcoholic steatohepatitis.

Non-alcoholic fatty liver disease (NAFLD) has emerged as the most prevalent liver disease in the world, yet there are still no approved pharmacological therapies to prevent or treat this condition. NAFLD encompasses a spectrum of severity, ranging from simple steatosis to non-alcoholic steatohepatitis (NASH). Although NASH is linked to an increased risk of hepatocellular carcinoma and cirrhosis and has now become the leading cause of liver failure-related transplantation, the majority of patients with NASH will ultimately die as a result of complications of type 2 diabetes mellitus (T2DM) and cardiometabolic diseases. Importantly, NAFLD is closely linked to obesity and tightly interrelated with insulin resistance and T2DM. Thus, targeting these interconnected conditions and taking a holistic attitude to the treatment of metabolic disease could prove to be a very beneficial approach. This Review will explore the latest relevant literature and discuss the ongoing therapeutic options for NAFLD focused on targeting intermediary metabolism, insulin resistance and T2DM to remedy the global health burden of these diseases.

Alzheimer’s (AD) and Parkinson’s Diseases (PD) are common neurodegenerative disorders growing in incidence and prevalence and for which there are no disease-modifying treatments. While there are considerable complexities in the presentations of these diseases, the histological pictures of these pathologies, as well as several rare genetic predispositions for each, point to the involvement of maladaptive protein processing and inflammation. Importantly, the common presentations of AD and PD are connected to aging and to dysmetabolism, including common co-diagnosis of metabolic syndrome or diabetes. Examination of anti-diabetic therapies in preclinical models and in some observational clinical studies have suggested effectiveness of the first generation insulin sensitizer pioglitazone in both AD and PD. Recently, the mitochondrial pyruvate carrier (MPC) was shown to be a previously unrecognized target of pioglitazone. New insulin sensitizers are in development that can be dosed to full engagement of this previously unappreciated mitochondrial target. Here we review molecular mechanisms that connect modification of pyruvate metabolism with known liabilities of AD and PD. The mechanisms involve modification of autophagy, inflammation, and cell differentiation in various cell types including neurons, glia, macrophages, and endothelium. These observations have implications for the understanding of the general pathology of neurodegeneration and suggest general therapeutic approaches to disease modification.

Macrophages reprogram their metabolism to promote appropriate responses. Proresolving macrophages primarily use fatty acid oxidation as an energy source. Metabolites generated during the catabolism of fatty acids aid in the resolution of inflammation and tissue repair, but the regulatory mechanisms that control lipid metabolism in macrophages are not fully elucidated. Lipin-1, a phosphatidic acid phosphatase that has transcriptional coregulator activity, regulates lipid metabolism in a variety of cells. In this current study, we show that lipin-1 is required for increased oxidative phosphorylation in IL-4 stimulated mouse (Mus musculus) macrophages. We also show that the transcriptional coregulatory function of lipin-1 is required for β-oxidation in response to palmitate (free fatty acid) and apoptotic cell (human) stimulation. Mouse bone marrow–derived macrophages lacking lipin-1 have a reduction in critical TCA cycle metabolites following IL-4 stimulation, suggesting a break in the TCA cycle that is supportive of lipid synthesis rather than lipid catabolism. Together, our data demonstrate that lipin-1 regulates cellular metabolism in macrophages in response to proresolving stimuli and highlights the importance of aligning macrophage metabolism with macrophage phenotype.

The pervasion of three daily meals and snacks is a relatively new introduction to our shared experience, and is coincident with an epidemic rise in obesity and cardiometabolic disorders of overnutrition. The past two decades have yielded convincing evidence regarding the adaptive, protective effects of calorie restriction (CR) and intermittent fasting (IF) against cardiometabolic, neurodegenerative, proteostatic and inflammatory diseases. Yet, durable adherence to intensive lifestyle changes is rarely attainable. New evidence now demonstrates that restricting carbohydrate entry into the hepatocyte by itself mimics several key signaling responses and physiological outcomes of IF and CR. This discovery raises the intriguing proposition that targeting hepatocyte carbohydrate transport to mimic fasting and caloric restriction can abate cardiometabolic and perhaps other fasting-treatable diseases. Here, we review the metabolic and signaling fates of a hepatocyte carbohydrate, identify evidence to target the key mediators within these pathways, and provide rationale and data to highlight carbohydrate transport as a broad, proximal intervention to block the deleterious sequelae of hepatic glucose and fructose metabolism.

ObjectiveMonoacylglycerol O-acyltransferase 1 (Mogat1), a lipogenic enzyme that converts monoacylglycerol to diacylglycerol, is highly expressed in adipocytes and may regulate lipolysis by re-esterifying fatty acids released during times when lipolytic rates are low. However, the role of Mogat1 in regulating adipocyte fat storage during differentiation and diet-induced obesity is relatively understudied.MethodsHere we generated adipocyte-specific Mogat1 knockout mice and subjected them to a high-fat diet to determine the effects of Mogat1 deficiency on diet-induced obesity. We also used Mogat1 floxed mice to develop preadipocyte cell lines wherein Mogat1 could be conditionally knocked out to study adipocyte differentiation in vitro.ResultsIn preadipocytes, we found that Mogat1 knockout at the onset of preadipocyte differentiation prevented the accumulation of glycerolipids and reduced the differentiation capacity of preadipocytes. However, the loss of adipocyte Mogat1 did not affect weight gain or fat mass induced by high-fat diet in mice. Furthermore, loss of Mogat1 in adipocytes did not affect plasma lipid or glucose concentrations or insulin tolerance.ConclusionsThese data suggest Mogat1 may play a role in adipocyte differentiation in vitro but not adipose tissue expansion in response to nutrient overload in mice.STUDY IMPORTANCEWhat is already known?Adipose tissue expansion through adipocyte precursor cell differentiation is critical for proper lipid storage during nutrient overload.Monoacylglycerol O-acyltransferase 1 (Mogat1), a lipogenic enzyme, is highly induced during adipocyte differentiation of human and mouse precursor cells and is reduced in patients with obesity and metabolic dysfunction.What does this study add?Mogat1 deletion during early adipocyte differentiation reduces differentiation capacity, adipogenic gene expression and lowers glycerolipid content of differentiated adipocytes.Adipocyte Mogat1 expression is dispensable for adiposity and metabolic outcomes high-fat fed mice and suggests compensation from other glycerolipid synthesis enzymes.How might these results change the direction of research?Understanding the molecular mechanisms of glycerolipid metabolism healthy adipose tissue expansion.

Hepatic gluconeogenesis from amino acids contributes significantly to diabetic hyperglycemia, but the molecular mechanisms involved are incompletely understood. Alanine transaminases (ALT1 and ALT2) catalyze the interconversion of alanine and pyruvate, which is required for gluconeogenesis from alanine. We find that ALT2 is overexpressed in the liver of diet-induced obese and db/db mice and that the expression of the gene encoding ALT2 (GPT2) is downregulated following bariatric surgery in people with obesity. The increased hepatic expression of Gpt2 in db/db liver is mediated by activating transcription factor 4, an endoplasmic reticulum stress-activated transcription factor. Hepatocyte-specific knockout of Gpt2 attenuates incorporation of

SUMMARYHepatic gluconeogenesis from amino acids contributes significantly to diabetic hyperglycemia, but the molecular mechanisms involved are incompletely understood. Alanine transaminases (ALT1 and ALT2) catalyze the interconversion of alanine and pyruvate, which is required for gluconeogenesis from alanine. We found that ALT2 was overexpressed in liver of diet-induced obese and db/db mice and that the expression of the gene encoding ALT2 (GPT2) was downregulated following bariatric surgery in people with obesity. The increased hepatic expression of Gpt2 in db/db liver was mediated by activating transcription factor 4; an endoplasmic reticulum stress-activated transcription factor. Hepatocyte-specific knockout of Gpt2 attenuated incorporation of 13C-alanine into newly synthesized glucose by hepatocytes. In vivo Gpt2 knockdown or knockout in liver had no effect on glucose concentrations in lean mice, but Gpt2 suppression alleviated hyperglycemia in db/db mice. These data suggest that ALT2 plays a significant role in hepatic gluconeogenesis from amino acids in diabetes.

SummaryHematopoietic homeostasis is maintained by stem and progenitor cells in part by extrinsic feedback cues triggered by mature cell loss. We demonstrate a different mechanism by which hematopoietic progenitors intrinsically anticipate and prevent the loss of mature progeny through metabolic switches. We examined hematopoiesis in mice conditionally deficient in long-chain fatty acid oxidation (carnitine palmitoyltransferase 2,Cpt2), glutaminolysis (glutaminase,Gls), or mitochondrial pyruvate import (mitochondrial pyruvate carrier 2,Mpc2). While genetic ablation ofCpt2orGlsminimally impacted most blood lineages, deletion ofMpc2led to a sharp decline in mature myeloid cells. However, MPC2-deficient myeloid cells rapidly recovered due to a transient increase in myeloid progenitor proliferation. Competitive bone marrow chimera and stable isotope tracing experiments demonstrated that this proliferative burst was intrinsic to MPC2-deficient progenitors and accompanied by a metabolic switch to glutaminolysis. Thus, hematopoietic progenitors intrinsically adjust to metabolic perturbations independently of feedback from downstream mature cells to maintain homeostasis.

Insulin signaling is known to regulate essential proteostasis mechanisms.The analyses here examined effects of insulin signaling in the PiZ mouse model of α1-antitrypsin deficiency in which hepatocellular accumulation and proteotoxicity of the misfolded α1-antitrypsin Z variant (ATZ) causes liver fibrosis and cancer.We first studied the effects of breeding PiZ mice to liver-insulin-receptor knockout (LIRKO) mice (with hepatocyte-specific insulin-receptor gene disruption). The results showed decreased hepatic ATZ accumulation and liver fibrosis in PiZ x LIRKO vs PiZ mice, with reversal of those effects when we bred PiZ x LIRKO mice onto a FOXO1-deficient background. Increased intracellular degradation of ATZ mediated by autophagy was identified as the likely mechanism for diminished hepatic proteotoxicity in PiZ x LIRKO mice and the converse was responsible for enhanced toxicity in PiZ x LIRKO x FOXO1-KO animals. Transcriptomic studies showed major effects on oxidative phosphorylation and autophagy genes, and significant induction of peroxisome proliferator-activated-receptor-γ-coactivator-1α (PGC1α) expression in PiZ-LIRKO mice. Because PGC1α plays a key role in oxidative phosphorylation, we further investigated its effects on ATZ proteostasis in our ATZ-expressing mammalian cell model. The results showed PGC1α overexpression or activation enhances autophagic ATZ degradation.These data implicate suppression of autophagic ATZ degradation by down-regulation of PGC1α as one mechanism by which insulin signaling exacerbates hepatic proteotoxicity in PiZ mice, and identify PGC1α as a novel target for development of new human α1-antitrypsin deficiency liver disease therapies.

The peptide hormone adropin regulates energy metabolism in skeletal muscle and plays important roles in the regulation of metabolic homeostasis. Besides muscle, the liver has an essential role in regulating glucose homeostasis. Previous studies have reported that treatment of diet-induced obese (DIO) male mice with adropin(34–76) (the putative secreted domain) reduces fasting blood glucose independently of body weight changes, suggesting that adropin suppresses glucose production in the liver. Here, we explored the molecular mechanisms underlying adropin's effects on hepatic glucose metabolism in DIO mice. Male DIO B6 mice maintained on a high-fat diet received five intraperitoneal injections of adropin(34–76) (450 nmol/kg/injection) over a 48-h period. We found that adropin(34–76) enhances major intracellular signaling activities in the liver that are involved in insulin-mediated regulation of glucose homeostasis. Moreover, treatment with adropin(34–76) alleviated endoplasmic reticulum stress responses and reduced activity of c-Jun N-terminal kinase in the liver, explaining the enhanced activities of hepatic insulin signaling pathways observed with adropin(34–76) treatment. Furthermore, adropin(34–76) suppressed cAMP activated protein kinase A (PKA) activities, resulting in reduced phosphorylation of inositol trisphosphate receptor, which mediates endoplasmic reticulum calcium efflux, and of cAMP-responsive element–binding protein, a key transcription factor in hepatic regulation of glucose metabolism. Adropin(34–76) directly affected liver metabolism, decreasing glucose production and reducing PKA-mediated phosphorylation in primary mouse hepatocytes in vitro. Our findings indicate that major hepatic signaling pathways contribute to the improved glycemic control achieved with adropin(34–76) treatment in situations of obesity.

During prolonged fasting, the liver plays a central role in maintaining systemic energy homeostasis by producing glucose and ketones in processes fueled by oxidation of fatty acids liberated from adipose tissue. In mice, this is accompanied by transient hepatic accumulation of glycerolipids. We found that the hepatic expression of monoacylglycerol acyltransferase 1 (Mogat1), an enzyme with monoacylglycerol acyltransferase (MGAT) activity that produces diacyl­glycerol from monoacylglycerol, was significantly increased in the liver of fasted mice compared with mice given ad libitum access to food. Basal and fasting-induced expression of Mogat1 was markedly diminished in the liver of mice lacking the transcription factor PPARα. Suppressing Mogat1 expression in liver and adipose tissue with antisense oligonucleotides (ASOs) reduced hepatic MGAT activity and triglyceride content compared with fasted controls. Surprisingly, the expression of many other PPARα target genes and PPARα activity was also decreased in mice given Mogat1 ASOs. When mice treated with control or Mogat1 ASOs were gavaged with the PPARα ligand, WY-14643, and then fasted for 18 h, WY-14643 administration reversed the effects of Mogat1 ASOs on PPARα target gene expression and liver triglyceride content. In conclusion, Mogat1 is a fasting-induced PPARα target gene that may feed forward to regulate liver PPARα activity during food deprivation.

A hepatic comorbidity of metabolic syndrome, known as nonalcoholic fatty liver disease (NAFLD), is increasing in prevalence in conjunction with the pandemics of obesity and diabetes. The spectrum of NAFLD ranges from simple hepatic fat accumulation to a more severe disease termed nonalcoholic steatohepatitis (NASH), involving inflammation, hepatocyte death, and fibrosis. Importantly, NASH is linked to a much higher risk of cirrhosis, liver failure, and hepatocellular carcinoma, as well as an increased risk for nonhepatic malignancies and cardiovascular disease. Interest in the understanding of the disease processes and search for treatments for the spectrum of NAFLD-NASH has increased exponentially, but there are no approved pharmacologic therapies. In this review, we discuss the existing literature supporting insulin-sensitizing thiazolidinedione compounds as potential drug candidates for the treatment of NASH. In addition, we put these results into new context by summarizing recent studies suggesting these compounds alter mitochondrial metabolism by binding and inhibiting the mitochondrial pyruvate carrier. Keywords: Nonalcoholic Steatohepatitis, Mitochondria, Pyruvate, Thiazolidinedione

The mitochondrial pyruvate carrier (MPC) has emerged as a therapeutic target for treating insulin resistance, type 2 diabetes, and nonalcoholic steatohepatitis (NASH). However, the molecular mechanisms by which MPC inhibition leads to metabolic improvements are incompletely understood. We evaluated whether MPC inhibitors might correct impairments in branched chain amino acid (BCAA) catabolism, which are predictive of developing diabetes and NASH. BCAA concentrations were assessed in plasma collected in EMMINENCE (NCT02784444), a randomized, placebo-controlled trial of the MPC inhibitor MSDC-0602K in people with NASH. MSDC-0602K treatment, which led to marked improvements in insulin sensitivity and diabetes endpoints, decreased plasma concentrations of BCAAs compared to baseline while placebo had no effect. The rate-limiting enzyme in BCAA catabolism is the mitochondrial branched chain ketoacid dehydrogenase (BCKDH). BCKDH activity is suppressed by phosphorylation mediated by a kinase (BDK) and activated by the phosphatase (PPM1K). In Huh7 or HepG2 cells, MPC inhibitors markedly reduced BCKDH phosphorylation, suggesting increased BCKDH activity. Chemical inhibition or siRNA-mediated silencing of BDK had no effect on suppression of BCKDH phosphorylation by MPC inhibitors. In contrast, PPM1K knockdown prevented the effects of MPC inhibition on phospho-BCKDH. Lastly, we quantified BCKDH phosphorylation in liver of wild-type and hepatocyte-specific MPC2 knockout (LS-Mpc2-/-) mice after 20 weeks of high fat diet. LS-Mpc2-/- mice exhibited reduced phosphorylation of BCKDH compared to wild-type mice. Lean LS-Mpc2-/- mice also exhibited reduced plasma BCAA concentrations after an overnight fast. These data demonstrate novel cross talk between mitochondrial pyruvate and BCAA metabolism and suggest that MPC inhibition leads to lower plasma BCAA concentrations and BCKDH phosphorylation via the phosphatase PPM1K. Disclosure D. Ferguson: None. N. K. H. Yiew: None. K. S. Mccommis: None. J. R. Colca: Board Member; Self; Metabolic Solutions Development Company, LLC, Employee; Self; Cirius Therapeutics. B. N. Finck: Advisory Panel; Self; Cirius Therapeutics, Stock/Shareholder; Self; Cirius Therapeutics. Funding National Institutes of Health (R01DK104735, T32DK007120)

Lipin 1 is a bifunctional protein that is a transcriptional regulator and has phosphatidic acid (PA) phosphohydrolase activity, which dephosphorylates PA to generate diacylglycerol. Human lipin 1 mutations lead to episodic rhabdomyolysis, and some affected patients exhibit cardiac abnormalities, including exercise-induced cardiac dysfunction and cardiac triglyceride accumulation. Furthermore, lipin 1 expression is deactivated in failing heart, but the effects of lipin 1 deactivation in myocardium are incompletely understood. We generated mice with cardiac-specific lipin 1 KO (cs-Lpin1-/-) to examine the intrinsic effects of lipin 1 in the myocardium. Cs-Lpin1-/- mice had normal systolic cardiac function but mild cardiac hypertrophy. Compared with littermate control mice, PA content was higher in cs-Lpin1-/- hearts, which also had an unexpected increase in diacylglycerol and triglyceride content. Cs-Lpin1-/- mice exhibited diminished cardiac cardiolipin content and impaired mitochondrial respiration rates when provided with pyruvate or succinate as metabolic substrates. After transverse aortic constriction-induced pressure overload, loss of lipin 1 did not exacerbate cardiac hypertrophy or dysfunction. However, loss of lipin 1 dampened the cardiac ionotropic response to dobutamine and exercise endurance in association with reduced protein kinase A signaling. These data suggest that loss of lipin 1 impairs cardiac functional reserve, likely due to effects on glycerolipid homeostasis, mitochondrial function, and protein kinase A signaling.

We previously demonstrated that endogenous phosphatidic acid (PA) promotes liver regeneration after acetaminophen (APAP) hepatotoxicity. Here, we hypothesized that exogenous PA is also beneficial. To test that, we treated mice with a toxic APAP dose at 0 h, followed by PA or vehicle (Veh) post-treatment. We then collected blood and liver at 6, 24, and 52 h. Post-treatment with PA 2 h after APAP protected against liver injury at 6 h, and the combination of PA and N-acetyl-l-cysteine (NAC) reduced injury more than NAC alone. Interestingly, PA did not affect canonical mechanisms of APAP toxicity. Instead, transcriptomics revealed that PA activated interleukin-6 (IL-6) signaling in the liver. Consistent with that, serum IL-6 and hepatic signal transducer and activator of transcription 3 (Stat3) phosphorylation increased in PA-treated mice. Furthermore, PA failed to protect against APAP in IL-6-deficient animals. Interestingly, IL-6 expression increased 18-fold in adipose tissue after PA, indicating that adipose is a source of PA-induced circulating IL-6. Surprisingly, however, exogenous PA did not alter regeneration, despite the importance of endogenous PA in liver repair, possibly due to its short half-life. These data demonstrate that exogenous PA is also beneficial in APAP toxicity and reinforce the protective effects of IL-6 in this model., Graphical abstract Exogenous phosphatidic acid (PA) increases interleukin-6 (IL-6) production in peripheral adipose tissue. The IL-6 then enters circulation and signals to the liver to reduce early acetaminophen (APAP)-induced liver injury.Image 1

Objective Monoacylglycerol acyltransferase (MGAT) enzymes catalyze the synthesis of diacylglycerol from monoacylglycerol. Previous work has suggested the importance of MGAT activity in the development of obesity-related hepatic insulin resistance. Indeed, antisense oligonucleotide (ASO)-mediated knockdown of Mogat1 mRNA, which encodes MGAT1, reduced hepatic MGAT activity and improved glucose tolerance and insulin resistance in high-fat diet (HFD)-fed mice. However, recent work has suggested that some ASOs may have off-target effects on body weight and metabolic parameters via activation of the interferon alpha/beta receptor 1 (IFNAR-1) pathway. Methods Mice with whole-body Mogat1 knockout or a floxed allele for Mogat1 to allow for liver-specific Mogat1-knockout (by either a liver-specific transgenic or adeno-associated virus-driven Cre recombinase) were generated. These mice were placed on an HFD, and glucose metabolism and insulin sensitivity were assessed after 16 weeks on diet. In some experiments, mice were treated with control scramble or Mogat1 ASOs in the presence or absence of IFNAR-1 neutralizing antibody. Results Genetic deletion of hepatic Mogat1, either acutely or chronically, did not improve hepatic steatosis, glucose tolerance, or insulin sensitivity in HFD-fed mice. Furthermore, constitutive Mogat1 knockout in all tissues actually exacerbated HFD-induced obesity, insulin sensitivity, and glucose intolerance on an HFD. Despite markedly reduced Mogat1 expression, liver MGAT activity was unaffected in all knockout mouse models. Mogat1 overexpression in hepatocytes increased liver MGAT activity and TAG content in low-fat-fed mice but did not cause insulin resistance. Multiple Mogat1 ASO sequences improved glucose tolerance in both wild-type and Mogat1 null mice, suggesting an off-target effect. Hepatic IFNAR-1 signaling was activated by multiple Mogat1 ASOs, but its blockade did not prevent the effects of either Mogat1 ASO on glucose homeostasis. Conclusion These results indicate that genetic loss of Mogat1 does not affect hepatic MGAT activity or metabolic homeostasis on HFD and show that multiple Mogat1 ASOs improve glucose metabolism through effects independent of targeting Mogat1 or activation of IFNAR-1 signaling., Graphical abstract Image 1, Highlights • Mogat1 liver-specific KO or KD does not improve metabolism in HFD-fed mice. • Whole-body Mogat1 deletion impairs insulin tolerance in HFD-fed mice. • Mogat1 ASOs improve whole-body metabolism independently of gene knockdown. • Blockade of the INFAR1 response does not prevent off-target effects of Mogat1 ASOs.

Nicotinamide adenine dinucleotide (NAD+) is an essential coenzyme that regulates cellular energy metabolism in many cell types. The major purpose of the present study was to test the hypothesis that NAD+ in white adipose tissue (WAT) is a regulator of whole-body metabolic flexibility in response to changes in insulin sensitivity and with respect to substrate availability and use during feeding and fasting conditions. To this end, we first evaluated the relationship between WAT NAD+ concentration and metabolic flexibility in mice and humans. We found that WAT NAD+ concentration was increased in mice after calorie restriction and exercise, 2 enhancers of metabolic flexibility. Bariatric surgery-induced 20% weight loss increased plasma adiponectin concentration, skeletal muscle insulin sensitivity, and WAT NAD+ concentration in people with obesity. We next analyzed adipocyte-specific nicotinamide phosphoribosyltransferase (Nampt) knockout (ANKO) mice, which have markedly decreased NAD+ concentrations in WAT. ANKO mice oxidized more glucose during the light period and after fasting than control mice. In contrast, the normal postprandial stimulation of glucose oxidation and suppression of fat oxidation were impaired in ANKO mice. Data obtained from RNA-sequencing of WAT suggest that loss of NAMPT increases inflammation, and impairs insulin sensitivity, glucose oxidation, lipolysis, branched-chain amino acid catabolism, and mitochondrial function in WAT, which are features of metabolic inflexibility. These results demonstrate a novel function of WAT NAMPT-mediated NAD+ biosynthesis in regulating whole-body metabolic flexibility, and provide new insights into the role of adipose tissue NAD+ biology in metabolic health.

The myocardium is metabolically flexible; however, impaired flexibility is associated with cardiac dysfunction in conditions including diabetes and heart failure. The mitochondrial pyruvate carrier (MPC) complex, composed of MPC1 and MPC2, is required for pyruvate import into the mitochondria. Here we show that MPC1 and MPC2 expression is downregulated in failing human and mouse hearts. Mice with cardiac-specific deletion of Mpc2 (CS-MPC2−/−) exhibited normal cardiac size and function at 6 weeks old, but progressively developed cardiac dilation and contractile dysfunction, which was completely reversed by a high-fat, low-carbohydrate ketogenic diet. Diets with higher fat content, but enough carbohydrate to limit ketosis, also improved heart failure, while direct ketone body provisioning provided only minor improvements in cardiac remodelling in CS-MPC2−/− mice. An acute fast also improved cardiac remodelling. Together, our results reveal a critical role for mitochondrial pyruvate use in cardiac function, and highlight the potential of dietary interventions to enhance cardiac fat metabolism to prevent or reverse cardiac dysfunction and remodelling in the setting of MPC deficiency. Impaired myocardial metabolic flexibility is associated with cardiac dysfunction in diabetes and heart failure. McCommis et al. reveal a critical role for mitochondrial pyruvate use in cardiac function, and use dietary interventions to enhance cardiac fat metabolism in dilated cardiomyopathy.

Phosphatidic acid (PA) is a glycerophospholipid intermediate in the triglyceride synthesis pathway that has incredibly important structural functions as a component of cell membranes and dynamic effects on intracellular and intercellular signaling pathways. Although there are many pathways to synthesize and degrade PA, a family of PA phosphohydrolases (lipin family proteins) that generate diacylglycerol constitute the primary pathway for PA incorporation into triglycerides. Previously, it was believed that the pool of PA used to synthesize triglyceride was distinct, compartmentalized, and did not widely intersect with signaling pathways. However, we now know that modulating the activity of lipin 1 has profound effects on signaling in a variety of cell types. Indeed, in most tissues except adipose tissue, lipin-mediated PA phosphohydrolase activity is far from limiting for normal rates of triglyceride synthesis, but rather impacts critical signaling cascades that control cellular homeostasis. In this review, we will discuss how lipin-mediated control of PA concentrations regulates metabolism and signaling in mammalian organisms.

ObjectiveMonoacylglycerol acyltransferase (MGAT) enzymes catalyze the synthesis of diacylglycerol from monoacylglycerol. Previous work has suggested the importance of MGAT activity in the development of obesity-related hepatic insulin resistance. Indeed, antisense oligonucleotide (ASO)-mediated knockdown of the gene encoding MGAT1, Mogat1, reduced hepatic MGAT activity and improved glucose tolerance and insulin resistance in high fat diet (HFD) fed mice. However, recent work has suggested that some ASOs may have off-target effects on body weight and metabolic parameters via activation of the interferon alpha/beta receptor 1 (IFNAR-1) pathway.MethodsMice with whole-body Mogat1 knockout or a floxed allele for Mogat1 to allow for liver-specific Mogat1-knockout (by either a liver-specific transgenic or adeno-associated virus-driven Cre recombinase) were generated. These mice were placed on a high fat diet and glucose metabolism and insulin sensitivity was assessed after 16 weeks on diet. In some experiments, mice were treated with control or Mogat1 or control ASOs in the presence or absence of IFNAR-1 neutralizing antibody.ResultsGenetic deletion of hepatic Mogat1, either acutely or chronically, did not improve hepatic steatosis, glucose tolerance, or insulin sensitivity in HFD-fed mice. Furthermore, constitutive Mogat1 knockout in all tissues actually exacerbated HFD-induced weight gain, insulin resistance, and glucose intolerance on a HFD. Despite markedly reduced Mogat1 expression, liver MGAT activity was unaffected in all knockout mouse models. Mogat1 overexpression hepatocytes increased liver MGAT activity and TAG content in low-fat fed mice, but did not cause insulin resistance. Interestingly, Mogat1 ASO treatment improved glucose tolerance in both wild-type and Mogat1 null mice, suggesting an off target effect. Inhibition of IFNAR-1 did not block the effect of Mogat1 ASO on glucose homeostasis.ConclusionThese results indicate that genetic loss of Mogat1 does not affect hepatic MGAT activity or metabolic homeostasis on HFD and show that Mogat1 ASOs improve glucose metabolism through effects independent of targeting Mogat1 or activation of IFNAR-1 signaling.Abstract FigureHighlightsMogat1 liver-specific KO or KD does not improve metabolism in HFD fed mice.Whole-body Mogat1-deletion impairs insulin tolerance in HFD fed mice.Mogat1 ASOs improves whole body metabolism independently of gene knockdown.Blockade of the INFR response does not prevent off-target effects of Mogat1 ASOs.

Macrophages reprogram their metabolism to promote appropriate responses. Pro-resolving macrophages primarily utilize fatty acid oxidation as an energy source. Metabolites generated during the catabolism of fatty acids aid in the resolution of inflammation and tissue repair, but the regulatory mechanisms that control lipid metabolism in macrophages is not fully elucidated. In this current study we show that lipin-1, a phosphatidic acid phosphatase and regulator of lipid metabolism, is required for increased oxidative phosphorylation during IL-4 mediated responses. We also show that the transcriptional coregulatory function of lipin-1 is required for β-oxidation in response to palmitate (free fatty acid) and apoptotic cell derived lipids. BMDMs lacking lipin-1 have a reduction in critical TCA cycle metabolites following IL-4 stimulation, suggesting a break in the TCA cycle that is supportive of lipid synthesis rather than lipid catabolism. Together, our data demonstrate that lipin-1 regulates intermediary metabolism within pro-resolving macrophages and highlights the importance of aligning macrophage metabolism with proper responses to stimuli.

During atherosclerosis, macrophages engulf and break down deposited modified low-density lipoproteins (modLDLs) into lipids and free fatty acids. The lipids and free fatty acids from these modLDLs either need to be stored during a process called glycerolipid synthesis or broken down during β-oxidation. In addition, free fatty acids can activate transcription factors to promote a pro-resolving macrophage phenotype. The protein lipin-1 is involved in both glycerolipid synthesis and β-oxidation. Lipin-1 enzymatic activity is a key step in the glycerolipid synthesis pathway; lipin-1 transcriptional co-regulatory activity either augments or represses various transcription factors that are activated via free fatty acids that promote β-oxidation and inhibit inflammation. Lipin-1 enzymatic activity increases pro-inflammatory macrophage phenotypes and is atherogenic. In contrast, we have also demonstrated that lipin-1 transcriptional co-regulatory activity promotes pro-resolving macrophage phenotypes leading us to the hypothesis that lipin-1 transcriptional co-regulatory activity is atheroprotective. Using a mouse model to delete lipin-1 in myeloid cells, we have demonstrated that loss of lipin-1 increases plaque size and pro-inflammatory gene expression. We have also shown mice lacking lipin-1 in myeloid cells have increased plaque collagen deposition and larger necrotic core formation. Combined, these data suggest that though lipin-1 enzymatic activity is atherogenic, lipin-1 transcriptional co-regulatory activity is atheroprotective. Overall, the results suggest that the dual activities of lipin-1 contribute to atherosclerosis progression in opposite ways.

Emerging evidence suggests that the capacity of adipocytes to appropriately store lipids and prevent their ectopic accumulation in liver and skeletal muscle may be an important regulator of systemic insulin sensitivity. Lipin 1 is a lipid phosphatase that is highly expressed in adipocytes and catalyzes the conversion of phosphatidic acid (PA) to diacylglycerol (DAG); the penultimate step in triglyceride synthesis. Adipose tissue lipin 1 expression positively correlates with insulin sensitivity in obese humans and mouse models. To test the role of adipose tissue lipin 1 in systemic insulin sensitivity, we generated mice lacking lipin 1 in adipocytes (Adn-Lpin1-/- mice). On normal chow, loss of lipin 1 in adipocytes modestly reduced white and brown adipose tissue mass, but did not affect glucose tolerance or insulin sensitivity. However, in response to 2 weeks high fat diet (HFD; 60% fat) feeding, Adn-Lpin1-/- mice remained lean, but developed hyperglycemia compared to littermate wild type (WT) mice. Lipidomic analyses demonstrated an increase in liver lipids (PA, DAG, and triglyceride) in HFD-fed Adn-Lpin1-/- mice compared to WT controls, whereas only PA accumulated in skeletal muscle. Plasma triglyceride concentration was not different, but free fatty acids and glycerol were significantly reduced in Adn-Lpin1-/- vs. WT mice fed HFD, suggesting that lipolysis may be decreased. Plasma adiponectin and resistin were significantly reduced in HFD-fed Adn-Lpin1-/- vs. WT control mice while leptin concentrations were not different. Lastly, with 5 weeks of HFD feeding, despite being leaner, Adn-Lpin1-/- mice became markedly hyperglycemic and glucose intolerant compared to littermate WT controls. Collectively, these data suggest that loss of lipin 1 in adipocytes leads to insulin resistance on HFD, likely due to an impaired capacity to store lipids appropriately in adipose tissue. Disclosure A.J. Lutkewitte: None. B.N. Finck: Advisory Panel; Self; Cirius Therapeutics. Stock/Shareholder; Self; Cirius Therapeutics.

Gluconeogenesis from amino acids contributes significantly to hepatic glucose production and hyperglycemia in diabetes. Alanine transaminases (ALT) catalyze the bi-directional interconversion of alanine to pyruvate with concomitant interconversion of α-ketoglutarate to glutamate. ALT1 is cytosolic while ALT2 is localized to the mitochondrial matrix. The mRNA expression and protein abundance of ALT1 and 2 was significantly increased in liver of db/db or diet induced obese mice compared to lean controls. The expression of ALT2 was also increased in obese humans compared to lean controls and was downregulated following bariatric surgery. The increase in hepatic ALT2 expression in diabetic mice was caused by endoplasmic reticulum stress and mediated by activating transcription factor 4 (ATF4). Indeed, inhibition of ATF4 reduced ALT2 expression in liver of db/db mice. Consistent with enhanced use of amino acids in gluconeogenesis, db/db and diet-induced obese mice had elevated blood glucose concentrations following IP administration of L-alanine or glutamine. ALT2 silencing in liver had no effect on glucose concentrations in lean mice, but alleviated alanine-induced hyperglycemia in db/db mice; likely by reducing the incorporation of alanine and/or glutamine into newly synthesized glucose. Plasma alanine tended to rise and glutamate significantly increased in db/db mice with ALT2 silencing whereas concentrations of the branched chain amino acids isoleucine, leucine, and valine decreased or tended to decrease. In summary, our results are consistent with a significant role for ALT2 in hepatic gluconeogenesis from amino acids and in the regulation of blood glucose levels in obesity and diabetes. Disclosure M.R. Martino: None. M. Gutierrez Aguilar: None. K.S. McCommis: None. J. Yoshino: None. B.N. Finck: Advisory Panel; Self; Cirius Therapeutics. Stock/Shareholder; Self; Cirius Therapeutics. Funding National Institutes of Health (R01DK117657)

Summary Many metabolic diseases disrupt endoplasmic reticulum (ER) homeostasis, but little is known about how metabolic activity is communicated to the ER. Here, we show in hepatocytes and other metabolically active cells that decreasing the availability of substrate for the tricarboxylic acid (TCA) cycle diminished NADPH production, elevated glutathione oxidation, led to altered oxidative maturation of ER client proteins, and attenuated ER stress. This attenuation was prevented when glutathione oxidation was disfavored. ER stress was also alleviated by inhibiting either TCA-dependent NADPH production or Glutathione Reductase. Conversely, stimulating TCA activity increased NADPH production, glutathione reduction, and ER stress. Validating these findings, deletion of the Mitochondrial Pyruvate Carrier—which is known to decrease TCA cycle activity and protect the liver from steatohepatitis—also diminished NADPH, elevated glutathione oxidation, and alleviated ER stress. Together, our results demonstrate a novel pathway by which mitochondrial metabolic activity is communicated to the ER through the relay of redox metabolites., Graphical Abstract, Highlights • Inhibiting nutrient catabolism alleviates ER stress in metabolically active cells • NADPH production and glutathione redox link TCA activity to ER homeostasis • ER client protein oxidation, maturation, and ERAD respond to metabolic activity • Cells lacking the Mitochondrial Pyruvate Carrier are resistant to ER stress, biological sciences; cell biology; functional aspects of cell biology

The myocardium is metabolically flexible and can use fatty acids, glucose, lactate/pyruvate, ketones, or amino acids to fuel mechanical work. However, impaired metabolic flexibility is associated with cardiac dysfunction in conditions including diabetes and heart failure. The mitochondrial pyruvate carrier (MPC) is required for pyruvate metabolism and is composed of a hetero-oligomer of two proteins known as MPC1 and MPC2. Interestingly, MPC1 and MPC2 expression is downregulated in failing human hearts and in a mouse model of heart failure. Mice with cardiac-specific deletion of MPC2 (CS-MPC2-/-) exhibited loss of both MPC2 and MPC1 proteins and reduced pyruvate-stimulated mitochondrial respiration. CS-MPC2-/- mice exhibited normal cardiac size and function at 6-weeks old, but progressively developed cardiac dilation and contractile dysfunction thereafter. Feeding CS-MPC2-/- mice a ketogenic diet (KD) completely prevented or reversed the cardiac remodeling and dysfunction. Other diets with higher fat content and enough carbohydrate to limit ketosis also improved heart failure in CS-MPC2-/- mice, but direct ketone body provisioning provided only minor improvements in cardiac remodeling. Finally, KD was also able to prevent further remodeling in an ischemic, pressure-overload mouse model of heart failure. In conclusion, loss of mitochondrial pyruvate utilization leads to dilated cardiomyopathy that can be corrected by a ketogenic diet.