To identify neurons that specifically increase blood glucose from among the diversely-functioning cell types in the ventromedial hypothalamic nucleus (VMN), we studied the cholecystokinin (CCK) receptor-B (CCKBR)-expressing VMN targets of glucose-elevating parabrachial nucleus neurons. Activating these VMNCCKBR neurons increased blood glucose. Furthermore, while silencing the broader VMN decreased energy expenditure and promoted weight gain without altering blood glucose, silencing VMNCCKBR neurons decreased hepatic glucose production (HGP), insulin-independently decreasing blood glucose without altering energy balance. Silencing VMNCCKBR neurons also impaired the counter-regulatory response (CRR) to insulin-induced hypoglycemia and glucoprivation and replicated hypoglycemia-associated autonomic failure (HAAF). Hence, VMNCCKBR cells represent a specialized subset of VMN cells that function to elevate glucose. These cells not only mediate the allostatic response to hypoglycemia, but also insulin-independently modulate the homeostatic setpoint for blood glucose, consistent with a role for the brain in the insulin-independent control of glucose homeostasis.
Jonathan N. Flak, Paulette Goforth, James Dell'Orco, Paul V. Sabatini, Chien Li, Nadejda Bozadjieva, Matthew J. Sorensen, Alec C. Valenta, Alan C. Rupp, Alison H. Affinati, Corentin Cras-Méneur, Ahsan Ansari, Jamie Sacksner, Nandan Kodur, Darleen A. Sandoval, Robert t. Kennedy, David Olson, Martin G. Myers Jr.
De novo lipogenesis is tightly regulated by insulin and nutritional signals to maintain metabolic homeostasis; excessive lipogenesis induces lipotoxicity, leading to nonalcoholic fatty liver disease (NAFLD) and type 2 diabetes. Genetic lipogenic programs have been extensively investigated, but epigenetic regulation of lipogenesis is poorly understood. Here, we identified Slug as an important epigenetic regulator of lipogenesis. Hepatic Slug levels were markedly upregulated in mice by either feeding or insulin treatment. In primary hepatocytes, insulin stimulation increased Slug expression, stability, and interactions with epigenetic enzyme lysine-specific demethylase-1 (Lsd1). Slug bound to the fatty acid synthase (Fasn) promoter where Slug-associated Lsd1 catalyzed H3K9 demethylation, thereby stimulating Fasn expression and lipogenesis. Ablation of Slug blunted insulin-stimulated lipogenesis; conversely, overexpression of Slug, but not a Lsd1 binding-defective Slug mutant, stimulated Fasn expression and lipogenesis. Lsd1 inhibitor treatment also blocked Slug-stimulated lipogenesis. Remarkably, hepatocyte-specific deletion of Slug inhibited the hepatic lipogenic program and protected against obesity-associated NAFLD, insulin resistance, and glucose intolerance in mice. Conversely, liver-restricted overexpression of Slug, but not the Lsd1 binding-defective Slug mutant, had the opposite effects. These results unveil an insulin/Slug/Lsd1/H3K9 demethylation lipogenic pathway that promotes NAFLD and type 2 diabetes.
Yan Liu, Haiyan Lin, Lin Jiang, Qingsen Shang, Lei Yin, Jiandie D. Lin, Wen-Shu Wu, Liangyou Rui
Mechanisms mediating the cardioprotective actions of glucagon-like peptide 1 (GLP-1) were unknown. Here, we show in both ex vivo and in vivo models of ischemic injury that treatment with GLP-1(28–36), a neutral endopeptidase–generated (NEP-generated) metabolite of GLP-1, was as cardioprotective as GLP-1 and was abolished by scrambling its amino acid sequence. GLP-1(28–36) enters human coronary artery endothelial cells (caECs) through macropinocytosis and acts directly on mouse and human coronary artery smooth muscle cells (caSMCs) and caECs, resulting in soluble adenylyl cyclase Adcy10–dependent (sAC-dependent) increases in cAMP, activation of protein kinase A, and cytoprotection from oxidative injury. GLP-1(28–36) modulates sAC by increasing intracellular ATP levels, with accompanying cAMP accumulation lost in sAC–/– cells. We identify mitochondrial trifunctional protein-α (MTPα) as a binding partner of GLP-1(28–36) and demonstrate that the ability of GLP-1(28–36) to shift substrate utilization from oxygen-consuming fatty acid metabolism toward oxygen-sparing glycolysis and glucose oxidation and to increase cAMP levels is dependent on MTPα. NEP inhibition with sacubitril blunted the ability of GLP-1 to increase cAMP levels in coronary vascular cells in vitro. GLP-1(28–36) is a small peptide that targets novel molecular (MTPα and sAC) and cellular (caSMC and caEC) mechanisms in myocardial ischemic injury.
M. Ahsan Siraj, Dhanwantee Mundil, Sanja Beca, Abdul Momen, Eric A. Shikatani, Talat Afroze, Xuetao Sun, Ying Liu, Siavash Ghaffari, Warren Lee, Michael B. Wheeler, Gordon Keller, Peter Backx, Mansoor Husain
BACKGROUND. Beige adipose tissue is associated with improved glucose homeostasis in mice. Adipose tissue contains β3 adrenergic receptors (β3-AR), and this study was intended to determine whether the treatment of obese, insulin-resistant humans with the β3AR agonist mirabegron, which stimulates beige adipose formation in subcutaneous white adipose tissue (SC WAT), would induce other beneficial changes in fat and muscle, and improve metabolic homeostasis. METHODS. Before and after β3AR agonist treatment, oral glucose tolerance tests and euglycemic clamps were performed, and histochemistry and gene expression profiling were performed from fat and muscle biopsies. PET CT scans quantified brown adipose tissue volume and activity and we conducted in vitro studies with primary cultures of differentiated human adipocytes and muscle.RESULTS. Clinical effects of mirabegron treatment included improved oral glucose tolerance (P<0.01), reduced hemoglobin A1c (P=0.01), and improved insulin sensitivity (P=0.03) and β-cell function (P=0.01). In SC WAT, mirabegron treatment stimulated lipolysis, reduced fibrotic gene expression and increased alternatively activated macrophages. Subjects with the most SC WAT beiging demonstrated the most improvement in β-cell function. In skeletal muscle, mirabegron reduced triglycerides, increased expression of PGC1A (P<0.05), and increased type I fibers (P<0.01). Conditioned media from adipocytes treated with mirabegron stimulated muscle fiber PGC1A expression in vitro (P<0.001). CONCLUSION. Mirabegron treatment significantly improves glucose tolerance in obese, insulin resistant humans. Since β-cells and skeletal muscle do not express β3-ARs, these data suggest that the beiging of SC WAT by mirabegron reduces adipose tissue dysfunction, which enhances muscle oxidative capacity and improves β-cell function. TRIAL REGISTRATION. Clinicaltrials.gov NCT02919176.FUNDING. NIH (DK112282, P30GM127211, DK 71349, and CTSA grant UL1TR001998).
Brian S. Finlin, Hasiyet Memetimin, Beibei Zhu, Amy L. Confides, Hemendra J. Vekaria, Riham H. El Khouli, Zachary R. Johnson, Philip M. Westgate, Jianzhong Chen, Andrew J. Morris, Patrick G. Sullivan, Esther E. Dupont-Versteegden, Philip A. Kern
BACKGROUND. Mirabegron is a β3-adrenergic receptor (β3-AR) agonist approved only for the treatment of overactive bladder. Encouraging preclinical results suggest that β3-AR agonists could also improve obesity-related metabolic disease by increasing brown adipose tissue (BAT) thermogenesis, white adipose tissue (WAT) lipolysis, and insulin sensitivity. METHODS. We treated 14 healthy women of diverse ethnicity, 27.5 ± 1.1 y, BMI 25.4 ± 1.2 kg/m2, with 100 mg mirabegron (Myrbetriq extended-release tablet, Astellas Pharma) for four weeks, open-label. The primary endpoint was the change in BAT metabolic activity as measured by [18F]-2-fluoro-D-2-deoxy-D-glucose (18F-FDG) positron emission tomography/computed tomography (PET/CT). Secondary endpoints included resting energy expenditure (REE), plasma metabolites, and glucose and insulin metabolism as assessed by frequently sampled intravenous glucose tolerance test. RESULTS. Chronic mirabegron therapy increased BAT metabolic activity. Whole-body REE was higher, without changes in body weight or composition. Additionally, there were elevations in plasma levels of the beneficial lipoprotein biomarkers high-density lipoprotein (HDL) and ApoA1, as well as total bile acids. Adiponectin, a WAT-derived hormone that has anti-diabetic and anti-inflammatory capabilities, increased with acute treatment and was 35% higher at study completion. Finally, an intravenous glucose tolerance test demonstrated higher insulin sensitivity, glucose effectiveness, and insulin secretion. CONCLUSION. These findings indicate that human BAT metabolic activity can be increased after chronic pharmacological stimulation with mirabegron and support the investigation of β3-AR agonists as a treatment for metabolic disease. TRIAL REGISTRATION. Clinicaltrials.gov NCT03049462. FUNDING. This work was supported by the Intramural Research Program of the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), DK075112, DK075116, DK071013, and DK071014.
Alana E. O'Mara, James W. Johnson, Joyce D. Linderman, Robert J. Brychta, Suzanne McGehee, Laura A. Fletcher, Yael A. Fink, Devika Kapuria, Thomas M. Cassimatis, Nathan Kelsey, Cheryl Cero, Zahraa Abdul-Sater, Francesca Piccinini, Alison S. Baskin, Brooks P. Leitner, Hongyi Cai, Corina M. Millo, William Dieckmann, Mary Walter, Norman B. Javitt, Yaron Rotman, Peter J. Walter, Marilyn Ader, Richard N. Bergman, Peter Herscovitch, Kong Y. Chen, Aaron M. Cypess
Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a complex disease with no known cause or mechanism. There is an increasing appreciation for the role of immune and metabolic dysfunction in the disease. ME/CFS has historically presented in outbreaks, often has a flu-like onset, and results in inflammatory symptoms. Patients suffer from severe fatigue and post-exertional malaise. There is little known about the metabolism of specific immune cells in ME/CFS patients. To investigate immune metabolism in ME/CFS, we isolated CD4+ and CD8+ T cells from 53 ME/CFS patients and 45 healthy controls. We analyzed glycolysis and mitochondrial respiration in resting and activated T cells, along with markers related to cellular metabolism, and plasma cytokines. We found that ME/CFS CD8+ T cells have reduced mitochondrial membrane potential compared to healthy controls. Both CD4+ and CD8+ T cells from ME/CFS patients had reduced glycolysis at rest, while CD8+ T cells also had reduced glycolysis following activation. ME/CFS patients had significant correlations between measures of T cell metabolism and plasma cytokine abundance that differed from healthy control subjects. Our data indicate that patients have impaired T cell metabolism consistent with ongoing immune alterations in ME/CFS that may illuminate the mechanism behind this disease.
Alexandra H. Mandarano, Jessica Maya, Ludovic Giloteaux, Daniel L. Peterson, Marco Maynard, C. Gunnar Gottschalk, Maureen R. Hanson
Background. An increase in intrahepatic triglyceride (IHTG) is the hallmark feature of nonalcoholic fatty liver disease (NAFLD) and is decreased by weight loss. Hepatic de novo lipogenesis (DNL) contributes to steatosis in people with NAFLD. The physiological factors that stimulate hepatic DNL and the effect of weight loss on hepatic DNL are not clear.Methods. Hepatic DNL, 24-h integrated plasma insulin and glucose concentrations, and both liver and whole-body insulin sensitivity were determined in people who were lean (n = 14), obese with normal IHTG content (Obese, n = 26) and obese with NAFLD (Obese-NAFLD, n = 27). Hepatic DNL was assessed by using the deuterated water method corrected for the potential confounding contribution of adipose tissue DNL. Liver and whole-body insulin sensitivity were assessed by using the hyperinsulinemic-euglycemic clamp procedure in conjunction with glucose tracer infusion. Six subjects in the Obese-NAFLD group were also evaluated before and after 10% diet-induced weight loss.Results. The contribution of hepatic DNL to IHTG-palmitate was 11%, 19% and 38% in the Lean, Obese and Obese-NAFLD groups, respectively. Hepatic DNL was inversely correlated with hepatic and whole-body insulin sensitivity, but directly correlated with 24-h plasma glucose and insulin concentrations. Weight loss decreased IHTG content, in conjunction with a decrease in hepatic DNL and 24-h plasma glucose and insulin concentrations. Conclusions. These data suggest hepatic DNL is an important regulator of IHTG content, and that increases in circulating glucose and insulin stimulate hepatic DNL in people with NAFLD. Weight loss decreases IHTG content, at least in part, by decreasing hepatic DNL.
Gordon I. Smith, Mahalakshmi Shankaran, Mihoko Yoshino, George G. Schweitzer, Maria Chondronikola, Joseph W. Beals, Adewole L. Okunade, Bruce W. Patterson, Edna Nyangau, Tyler Field, Claude B. Sirlin, Saswata Talukdar, Marc K. Hellerstein, Samuel Klein
Background: Ceramides are sphingolipids that play causative roles in diabetes and heart disease, with their serum levels measured clinically as biomarkers of cardiovascular disease (CVD). Methods: We performed targeted lipidomics on serum samples of individuals with familial coronary artery disease (CAD) (n = 462) and population-based controls (n = 212) to explore the relationship between serum sphingolipids and CAD, employing unbiased machine learning to identify sphingolipid species positively associated with CAD. Results: Nearly every sphingolipid measured (n = 30 of 32) was significantly elevated in subjects with CAD compared with population controls. We generated a novel Sphingolipid Inclusive CAD risk score, termed SIC, that demarcates CAD patients independently and more effectively than conventional clinical CVD biomarkers including LDL-cholesterol and serum triglycerides. This new metric comprises several minor lipids which likely serve as measures of flux through the ceramide biosynthesis pathway, rather than the abundant deleterious ceramide species that are incorporated in other ceramide-based scores. Conclusion: This study validates serum ceramides as candidate biomarkers of cardiovascular disease and suggests that comprehensive sphingolipid panels be considered as measures of CVD.
Annelise M. Poss, J. Alan Maschek, James E. Cox, Benedikt J. Hauner, Paul N. Hopkins, Steven C. Hunt, William L. Holland, Scott A. Summers, Mary C. Playdon
Gout is caused by deposition of monosodium urate crystals in joints when plasma uric acid levels are chronically elevated beyond the saturation threshold, mostly due to renal underexcretion of uric acid. Although molecular pathways of this underexcretion have been elucidated, its etiology remains mostly unknown. We demonstrate that gout can be caused by a mutation in LDHD within the putative catalytic site of the encoded d-lactate dehydrogenase, resulting in augmented blood levels of d-lactate, a stereoisomer of l-lactate, which is normally present in human blood in miniscule amounts. Consequent excessive renal secretion of d-lactate in exchange for uric acid reabsorption culminated in hyperuricemia and gout. We showed that LDHD expression is enriched in tissues with a high metabolic rate and abundant mitochondria and that d-lactate dehydrogenase resides in the mitochondria of cells overexpressing the human LDHD gene. Notably, the p.R370W mutation had no effect on protein localization. In line with the human phenotype, injection of d-lactate into naive mice resulted in hyperuricemia. Thus, hyperuricemia and gout can result from the accumulation of metabolites whose renal excretion is coupled to uric acid reabsorption.
Max Drabkin, Yuval Yogev, Lior Zeller, Raz Zarivach, Ran Zalk, Daniel Halperin, Ohad Wormser, Evgenia Gurevich, Daniel Landau, Rotem Kadir, Yonatan Perez, Ohad S. Birk
Brown adipose tissue (BAT), as the main site of adaptive thermogenesis, exerts beneficial metabolic effects on obesity and insulin resistance. BAT has been previously assumed to contain a homogeneous population of brown adipocytes. Utilizing multiple mouse models capable of genetically labeling different cellular populations, as well as single-cell RNA sequencing, and 3D tissue profiling, we discovered a new brown adipocyte subpopulation with low thermogenic activity co-existing with the classical high thermogenic brown adipocytes within the BAT. These low thermogenic brown adipocytes had significantly lower Ucp1 and Adipoq expression, larger lipid droplets, and lower mitochondrial content. Functional analyses showed that the low thermogenic brown adipocytes have significant lower basal mitochondrial respiration, and they are specialized in fatty acid uptake. Upon changes in environmental temperature, the two brown adipocyte subpopulations underwent dynamic inter-conversions. Cold exposure converted low thermogenic brown adipocytes into high thermogenic cells, and a thermoneutral environment had the opposite effect. This recruitment of high thermogenic brown adipocytes by cold stimulation is not affected by high fat diet feeding, but significantly declined with age. Our results revealed a high degree of functional heterogeneity of brown adipocytes.
Anying Song, Wenting Dai, Min Jee Jang, Leonard Medrano, Zhuo Li, Hu Zhao, Mengle Shao, Jiayi Tan, Aimin Li, Tinglu Ning, Marcia M. Miller, Brian Armstrong, Janice M. Huss, Yi Zhu, Yong Liu, Viviana Gradinaru, Xiwei Wu, Lei Jiang, Philipp E. Scherer, Qiong A. Wang