Featured ArticlesVolume 6 | No. 11 | November 2017
|The autonomic nervous system and GLP-1 receptors control heart rateGlucagon-like peptide-1 (GLP-1) is a gut hormone with pleiotropic effects that, among other actions, also controls heart rate (HR). The data available so far suggest that GLP-1 may control HR indirectly, through modulation of autonomic nervous system activity, as well as directly, through control of pacemaker activity via the atrial GLP-1 receptor (GLP-1R). To elucidate the relative importance of and inter-dependence of these pathways, Baggio, Ussher et al. have now studied the regulation of HR in control and Glp1rCM-/- mice treated with the GLP-1R agonists lixisenatide or liraglutide. Their findings reveal temporally distinct contributions from both the sympathetic nervous system and cardiac GLP-1Rs in the HR response to GLP-1R agonism in vivo. |
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Objective: Glucagon-like peptide-1 (GLP-1) is secreted from enteroendocrine cells and exerts a broad number of metabolic actions through activation of a single GLP-1 receptor (GLP-1R). The cardiovascular actions of GLP-1 have garnered increasing attention as GLP-1R agonists are used to treat human subjects with diabetes and obesity that may be at increased risk for development of heart disease. Here we studied mechanisms linking GLP-1R activation to control of heart rate (HR) in mice.
Methods: The actions of GLP-1R agonists were examined on the control of HR in wild type mice (WT) and in mice with cardiomyocyte-selective disruption of the GLP-1R (Glp1rCM−/−). Complimentary studies examined the effects of GLP-1R agonists in mice co-administered propranolol or atropine. The direct effects of GLP-1R agonism on HR and ventricular developed pressure were examined in isolated perfused mouse hearts ex vivo, and atrial depolarization was quantified in mouse hearts following direct application of liraglutide to perfused atrial preparations ex vivo.
Results: Doses of liraglutide and lixisenatide that were equipotent for acute glucose control rapidly increased HR in WT and Glp1rCM−/− mice in vivo. The actions of liraglutide to increase HR were more sustained relative to lixisenatide, and diminished in Glp1rCM−/− mice. The acute chronotropic actions of GLP-1R agonists were attenuated by propranolol but not atropine. Neither native GLP-1 nor lixisenatide increased HR or developed pressure in perfused hearts ex vivo. Moreover, liraglutide had no direct effect on sinoatrial node firing rate in mouse atrial preparations ex vivo. Despite co-localization of HCN4 and GLP-1R in primate hearts, HCN4-directed Cre expression did not attenuate levels of Glp1r mRNA transcripts, but did reduce atrial Gcgr expression in the mouse heart.
Conclusions: GLP-1R agonists increase HR through multiple mechanisms, including regulation of autonomic nervous system function, and activation of the atrial GLP-1R. Surprisingly, the isolated atrial GLP-1R does not transduce a direct chronotropic effect following exposure to GLP-1R agonists in the intact heart, or isolated atrium, ex vivo. Hence, cardiac GLP-1R circuits controlling HR require neural inputs and do not function in a heart-autonomous manner.[Hide abstract]
|Acute activation of GLP-1-expressing neurons promotes glucose homeostasis and insulin sensitivityIn response to food intake, glucagon-like peptides (GLP-1/2) are co-released from enteroendocrine L cells in the gut as well as preproglucagon (PPG) neurons in the nucleus of the solitary tract (NTS) of the brainstem, which together constitute the key nutritional signals for the control of energy balance and glucose homeostasis. Shi and colleagues determined whether activation of PPG neurons per se modulates glucose homeostasis and insulin sensitivity in vivo. They show in Gcg-Cre lean mice infected with excitatory hM3Dq virus in the brainstem NTS that acute activation of Gcg neurons (which include the PPG neurons) enhances glucose tolerance, suppresses basal endogenous glucose production, and augments hepatic insulin sensitivity.|
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Objective: Glucagon-like peptides are co-released from enteroendocrine L cells in the gut and preproglucagon (PPG) neurons in the brainstem. PPG-derived GLP-1/2 are probably key neuroendocrine signals for the control of energy balance and glucose homeostasis. The objective of this study was to determine whether activation of PPG neurons per se modulates glucose homeostasis and insulin sensitivity in vivo.
Methods: We generated glucagon (Gcg) promoter-driven Cre transgenic mice and injected excitatory hM3Dq-mCherry AAV into their brainstem NTS. We characterized the metabolic impact of PPG neuron activation on glucose homeostasis and insulin sensitivity using stable isotopic tracers coupled with hyperinsulinemic euglycemic clamp.
Results: We showed that after ip injection of clozapine N-oxide, Gcg-Cre lean mice transduced with hM3Dq in the brainstem NTS downregulated basal endogenous glucose production and enhanced glucose tolerance following ip glucose tolerance test. Moreover, acute activation of PPG neuronsNTS enhanced whole-body insulin sensitivity as indicated by increased glucose infusion rate as well as augmented insulin-suppression of endogenous glucose production and gluconeogenesis. In contrast, insulin-stimulation of glucose disposal was not altered significantly.
Conclusions: We conclude that acute activation of PPG neurons in the brainstem reduces basal glucose production, enhances intraperitoneal glucose tolerance, and augments hepatic insulin sensitivity, suggesting an important physiological role of PPG neurons-mediated circuitry in promoting glycemic control and insulin sensitivity.[Hide abstract]
|Superior reductions in hepatic steatosis and fibrosis with a GLP1R agonist and obeticholic acid The current standard of care for nonalcoholic steatohepatitis (NASH) is limited to ameliorating components of the associated metabolic syndrome. Unfortunately, the long-term effectiveness of these interventions is questionable. Therefore, developing new pharmacological therapies is vital to combating the complex nature of NASH. Jouihan and colleagues investigated the use of long-acting glucagon-like peptide-1 receptor (GLP-1R) agonist IP118 and the farnesoid-X receptor (FXR) agonist obeticholic acid (OCA). They found that their combination is more effective than using either compounds separately in slowing or reversing NASH-associated defects including hepatic steatosis and fibrosis.|
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Objective: Nonalcoholic steatohepatitis (NASH) is an unmet need associated with metabolic syndrome. There are no approved therapies for NASH; however, glucagon-like peptide-1 receptor (GLP-1R) and farnesoid-X receptor (FXR) agonists are promising drug targets. We investigated the therapeutic effects of co-administration of a GLP-1R agonist, IP118, with FXR agonist obeticholic acid (OCA) in mice.
Methods: OCA and IP118 alone and in combination were sub-chronically administered to Lepob/Lepob mice with diet-induced NASH or diet-induced obese (DIO) mice. Metabolic (body weight and glucose) and liver (biochemical and histological) endpoints were assessed. NASH severity in Lepob/Lepob mice was graded using a customized integrated scoring system.
Results: OCA reduced liver weight and lipid in NASH mice (both by −17%) but had no effect on plasma ALT or AST levels. In contrast, IP118 significantly reduced liver weight (−21%), liver lipid (−15%), ALT (−29%), and AST (−27%). The combination of OCA + IP118 further reduced liver weight (−29%), liver lipid (−22%), ALT (−39%), and AST (−36%). Combination therapy was superior to monotherapies in reducing hepatic steatosis, inflammation, and fibrosis. Hepatic improvements with IP118 and OCA + IP118 were associated with reduced body weight (−4.3% and −3.5% respectively) and improved glycemic control in OCA + IP118-treated mice. In DIO mice, OCA + IP118 co-administration reduced body weight (−25.3%) to a greater degree than IP118 alone (−12.5%) and further improved glucose tolerance and reduced hepatic lipid.
Conclusions: Our data suggest a complementary or synergistic therapeutic effect of GLP-1R and FXR agonism in mouse models of metabolic disease and NASH.[Hide abstract]
|Host-microbiota interaction induces inflammation and glucose intoleranceThe gut microbiota has emerged as an important factor regulating host physiology and metabolism, in particular glucose metabolism and adiposity. Molinaro et al. did a time-resolved study on how colonization of germfree (GF) mice affects kinetics of adiposity and glucose metabolism. They found that colonization resulted in a bi-phasic glucose impairment. The first phase, occurring within 3 days of colonization (early phase) co-occurred with an inflammatory response and was independent of adiposity. The second phase, 14-28 days after colonization (delayed phase), was mostly ascribed to adipose tissue expansion and inflammation.|
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Objective: Gut microbiota modulates adiposity and glucose metabolism in humans and mice. Here we investigated how colonization of germ-free (GF) mice affects kinetics of adiposity and glucose metabolism.
Methods: Adiposity and glucose metabolism were evaluated at different time points in ex-GF and antibiotic treated mice after colonization with gut microbiota from a conventionally raised (CONV-R) mouse. Mouse physiology, microbiome configuration, serum cytokine levels, and gene expression for inflammatory markers were performed in different tissues.
Results: Colonization resulted in a bi-phasic glucose impairment: the first phase occurring within 3 days of colonization (early phase) and the second 14–28 days after colonization (delayed phase). The early phase co-occurred with an inflammatory response and was independent of adiposity, while the delayed phase was mostly ascribed to adipose tissue expansion and inflammation. Importantly, re-colonization of antibiotic treated mice displays only the delayed phase of glucose impairment and adiposity, suggesting that the early phase may be unique to colonization of the immature GF mice gut.
Conclusions: Our results provide new insights on host–microbiota interaction during colonization of GF mice and the resulting effects on adiposity and glucose metabolism in a time resolved fashion.[Hide abstract]
|Deletion of hepatic ChREBP impairs glucose homeostasis and insulin sensitivity Carbohydrate response element binding protein (ChREBP) is a key transcription factor involved in coordinating the feeding response. Association studies have linked increased ChREBP expression in liver to hepatic steatosis and insulin resistance. Jois and colleagues sought to clarify the role of hepatic ChREBP in glucose homeostasis and insulin sensitivity using a novel mouse model of hepatic ChREBP deletion. They found that liver ChREBP is vital in maintaining hepatic insulin sensitivity and coordinating the appropriate responses to fasting and feeding through glucose sensing.|
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Objective: Carbohydrate response element binding protein (ChREBP) is a transcription factor that responds to glucose and activates genes involved in the glycolytic and lipogenic pathways. Recent studies have linked adipose ChREBP to insulin sensitivity in mice. However, while ChREBP is most highly expressed in the liver, the effect of hepatic ChREBP on insulin sensitivity remains unknown. To clarify the importance of hepatic ChREBP on glucose homeostasis, we have generated a knockout mouse model that lacks this protein specifically in the liver (Liver-ChREBP KO).
Methods: Using Liver-ChREBP KO mice, we investigated whether hepatic ChREBP deletion influences insulin sensitivity, glucose homeostasis and the development of hepatic steatosis utilizing various dietary stressors. Furthermore, we determined gene expression changes in response to fasted and fed states in liver, white, and brown adipose tissues.
Results: Liver-ChREBP KO mice had impaired insulin sensitivity as indicated by reduced glucose infusion to maintain euglycemia during hyperinsulinemic-euglycemic clamps on both chow (25% lower) and high-fat diet (33% lower) (p < 0.05). This corresponded with attenuated suppression of hepatic glucose production. Although Liver-ChREBP KO mice were protected against carbohydrate-induced hepatic steatosis, they displayed worsened glucose tolerance. Liver-ChREBP KO mice did not show the expected gene expression changes in liver in response to fasted and fed states. Interestingly, hepatic ChREBP deletion also resulted in gene expression changes in white and brown adipose tissues, suggesting inter-tissue communication. This included an almost complete abolition of BAT ChREBPβ induction in the fed state (0.15-fold) (p = 0.015) along with reduced lipogenic genes. In contrast, WAT showed inappropriate increases in lipogenic genes in the fasted state along with increased PEPCK1 in both fasted (3.4-fold) and fed (5.1-fold) states (p < 0.0001).
Conclusions: Overall, hepatic ChREBP is protective in regards to hepatic insulin sensitivity and whole body glucose homeostasis. Hepatic ChREBP action can influence other peripheral tissues and is likely essential in coordinating the body's response to different feeding states.[Hide abstract]
|FGF21 is robustly induced by ethanol and has a protective role in liver injury Chronic ethanol consumption is known to be lipotoxic and is associated with accumulation of hepatic fat. While alcoholic liver disease (ALD) and non-alcoholic fatty liver disease (NAFLD) have distinct characteristics, they also share similar pathologies. Considering this, Desai, Singhal and colleagues hypothesized that ethanol consumption might also increase expression of fibroblast growth factor 21 (FGF21). They found increased FGF21 expression after binge alcohol consumption in humans and mice. Their data suggest that FGF21 has a dual role in ethanol metabolism. Acutely, FGF21 acts centrally to inhibit ethanol consumption. Chronically, the rise in hepatic FGF21 expression may have anti-inflammatory and anti-fibrotic roles.|
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Objective: Excess ethanol consumption has serious pathologic consequences. In humans, repeated episodes of binge drinking can lead to liver damage and have adverse effects on other organs such as pancreas and brain. Long term chronic consumption of ethanol can also result in progressive alcoholic liver disease and cirrhosis. Fibroblast growth factor 21 (FGF21) is a metabolic regulator with multiple physiologic functions. FGF21 is a novel biomarker for non-alcoholic fatty liver disease (NAFLD) in humans and limits hepatotoxicity in mice. Therefore, we explored the possibility that FGF21 plays a role in response to ethanol consumption in both humans and mice.
Methods: We used a binge drinking paradigm in humans to examine the effect of acute ethanol consumption on circulating FGF21. We adapted this paradigm to evaluate the acute response to ethanol in mice. We then examined the role of FGF21 on liver pathology in two models of chronic ethanol consumption in both wild type (WT) mice and mice lacking FGF21 (FGF21-KO).
Results: Acute ethanol consumption resulted in a robust induction of serum FGF21 after 6 h in both humans and mice. Serum ethanol peaked at 1 h in both species and was cleared by 6 h. Ethanol clearance was the same in WT and FGF21-KO mice, indicating that FGF21 does not play a major role in ethanol metabolism in a binge paradigm. When FGF21-KO mice were fed the Lieber–DeCarli diet, a high fat diet supplemented with ethanol, a higher mortality was observed compared to WT mice after 16 days on the diet. When FGF21-KO mice consumed 30% ethanol in drinking water, along with a normal chow diet, there was no mortality observed even after 16 weeks, but the FGF21-KO mice had significant liver pathology compared to WT mice.
Conclusions: Acute or binge ethanol consumption significantly increases circulating FGF21 levels in both humans and mice. However, FGF21 does not play a role in acute ethanol clearance. In contrast, chronic ethanol consumption in the absence of FGF21 is associated with significant liver pathology alone or in combination with excess mortality, depending on the type of diet consumed with ethanol. This suggests that FGF21 protects against long term ethanol induced hepatic damage and may attenuate progression of alcoholic liver disease. Further study is required to assess the therapeutic potential of FGF21 in the treatment of alcoholic liver disease.[Hide abstract]
|Islet-enriched long non-coding RNAs contributing to β-cell failure in type 2 diabetes Long non-coding RNAs (lncRNAs) participate in diverse gene-regulatory mechanisms and their dysregulation has been implicated in many human diseases. Recently, lncRNAs were found to contribute to β-cell development and glucose homeostasis. Motterle et al. identified novel islet lncRNAs and investigated their role in the regulation of β-cell functions. They show that lncRNAs are modulated in islets from obese diabetic mice and individuals with type 2 diabetes (T2D) and may contribute to β-cell failure during T2D development.|
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Objective: Non-coding RNAs constitute a major fraction of the β-cell transcriptome. While the involvement of microRNAs is well established, the contribution of long non-coding RNAs (lncRNAs) in the regulation of β-cell functions and in diabetes development remains poorly understood. The aim of this study was to identify novel islet lncRNAs differently expressed in type 2 diabetes models and to investigate their role in β-cell failure and in the development of the disease.
Methods: Novel transcripts dysregulated in the islets of diet-induced obese mice were identified by high throughput RNA-sequencing coupled with de novo annotation. Changes in the level of the lncRNAs were assessed by real-time PCR. The functional role of the selected lncRNAs was determined by modifying their expression in MIN6 cells and primary islet cells.
Results: We identified about 1500 novel lncRNAs, a number of which were differentially expressed in obese mice. The expression of two lncRNAs highly enriched in β-cells, βlinc2, and βlinc3, correlated to body weight gain and glycemia levels in obese mice and was also modified in diabetic db/db mice. The expression of both lncRNAs was also modulated in vitro in isolated islet cells by glucolipotoxic conditions. Moreover, the expression of the human orthologue of βlinc3 was altered in the islets of type 2 diabetic patients and was associated to the BMI of the donors. Modulation of the level of βlinc2 and βlinc3 by overexpression or downregulation in MIN6 and mouse islet cells did not affect insulin secretion but increased β-cell apoptosis.
Conclusions: Taken together, the data show that lncRNAs are modulated in a model of obesity-associated type 2 diabetes and that variations in the expression of some of them may contribute to β-cell failure during the development of the disease.[Hide abstract]
|Disruption of orthopedia homeobox (Otp) is associated with obesity and anxiety The development of neuroendocrine cell lineages in the hypothalamus requires a number of transcription factors including orthopedia homeobox (OTP). Moir, Bochukova and colleagues have carried out a high throughput dominant N-ethyl-N-nitrosourea (ENU) mutagenesis screen in mice with the objective of identifying novel obesity models. They report an obese model with a novel hypomorphic mutation in the orthopedia homeobox (Otp) gene.|
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Objective: Genetic studies in obese rodents and humans can provide novel insights into the mechanisms involved in energy homeostasis.
Methods: In this study, we genetically mapped the chromosomal region underlying the development of severe obesity in a mouse line identified as part of a dominant N-ethyl-N-nitrosourea (ENU) mutagenesis screen. We characterized the metabolic and behavioral phenotype of obese mutant mice and examined changes in hypothalamic gene expression. In humans, we examined genetic data from people with severe early onset obesity.
Results: We identified an obese mouse heterozygous for a missense mutation (pR108W) in orthopedia homeobox (Otp), a homeodomain containing transcription factor required for the development of neuroendocrine cell lineages in the hypothalamus, a region of the brain important in the regulation of energy homeostasis. OtpR108W/+ mice exhibit increased food intake, weight gain, and anxiety when in novel environments or singly housed, phenotypes that may be partially explained by reduced hypothalamic expression of oxytocin and arginine vasopressin. R108W affects the highly conserved homeodomain, impairs DNA binding, and alters transcriptional activity in cells. We sequenced OTP in 2548 people with severe early-onset obesity and found a rare heterozygous loss of function variant in the homeodomain (Q153R) in a patient who also had features of attention deficit disorder.
Conclusions: OTP is involved in mammalian energy homeostasis and behavior and appears to be necessary for the development of hypothalamic neural circuits. Further studies will be needed to investigate the contribution of rare variants in OTP to human energy homeostasis.[Hide abstract]
|Four microRNAs are regulators of skeletal muscle mitochondrial metabolism MicroRNAs (miRNAs) are small non-coding RNA strands of approximately 20-22 nucleotides regulate the stability and translation of conventional messenger RNAs. Dahlmans et al. performed an unbiased, hypothesis-free screening approach in C2C12 myoblasts, using specific miRNA inhibitors, leading to the identification of 19 specific miRNAs as positive modulators of mitochondrial metabolism when silenced. The expression of four of these miRNAs showed a strong relationship with in vivo mitochondrial function in humans.|
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Objective: Strategies improving skeletal muscle mitochondrial capacity are commonly paralleled by improvements in (metabolic) health. We and others previously identified microRNAs regulating mitochondrial oxidative capacity, but data in skeletal muscle are limited. Therefore, the present study aimed to identify novel microRNAs regulating skeletal muscle mitochondrial metabolism.
Methods and results: We conducted an unbiased, hypothesis-free microRNA silencing screen in C2C12 myoblasts, using >700 specific microRNA inhibitors, and investigated a broad panel of mitochondrial markers. After subsequent validation in differentiated C2C12 myotubes, and exclusion of microRNAs without a human homologue or with an adverse effect on mitochondrial metabolism, 19 candidate microRNAs remained. Human clinical relevance of these microRNAs was investigated by measuring their expression in human skeletal muscle of subject groups displaying large variation in skeletal muscle mitochondrial capacity.
Conclusions: The results show that that microRNA-320a, microRNA-196b-3p, microRNA-150-5p, and microRNA-34c-3p are tightly related to in vivo skeletal muscle mitochondrial function in humans and identify these microRNAs as targets for improving mitochondrial metabolism.[Hide abstract]
|Absence of the kinase S6k1 mimics the effect of chronic endurance exercise Physical exercise training is associated with increased insulin sensitivity, improved glycemic control, and reduced risk for developing type 2 diabetes mellitus. However, the regulatory network responsible for the diabetes-protective effect of exercise is not well understood. Activation of the mammalian target of rapamycin complex 1(mTORC1)/ Ribosomal protein S6 Kinase-1 (S6k1) pathway may play an important role in metabolic adaptation to endurance exercise training that relates to improvements in glucose homeostasis. Binsch, Jelenik et al. report that the absence of S6k1 upregulates ketogenesis and improves oxidative substrate utilization and conservation of carbohydrate reserves under a high fat diet consumption, thus mimicking the condition of chronic endurance exercise.|
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Objective: Ribosomal protein S6 Kinase-1 (S6K1) has been linked to resistance exercise-mediated improvements in glycemia. We hypothesized that S6K1 may also play a role in regulating glycemic control in response to endurance exercise training.
Methods: S6k1-knockout (S6K1KO) and WT mice on a 60 cal% high-fat diet were trained for 4 weeks on treadmills, metabolically phenotyped, and compared to sedentary controls.
Results: WT mice showed improved glucose tolerance after training. In contrast, S6K1KO mice displayed equally high glucose tolerance already in the sedentary state with no further improvement after training. Similarly, training decreased mitochondrial ROS production in skeletal muscle of WT mice, whereas ROS levels were already low in the sedentary S6K1KO mice with no further decrease after training. Nevertheless, trained S6K1KO mice displayed an increased running capacity compared to trained WT mice, as well as substantially reduced triglyceride contents in liver and skeletal muscle. The improvements in glucose handling and running endurance in S6K1KO mice were associated with markedly increased ketogenesis and a higher respiratory exchange ratio.
Conclusions: In high-fat fed mice, loss of S6K1 mimics endurance exercise training by reducing mitochondrial ROS production and upregulating oxidative utilization of ketone bodies. Pharmacological targeting of S6K1 may improve the outcome of exercise-based interventions in obesity and diabetes.[Hide abstract]
|FGF21 mimetic antibody stimulates UCP1-independent brown fat thermogenesis Fibroblast Growth Factor 21 (FGF21) is an endocrine member of the FGF super family that has been identified as a regulator of brown adipose tissue (BAT) thermogenesis and nutrient metabolism. Although pre-clinical and clinical studies suggest that FGF21 analogs may become an effective therapy for obesity related disorders, the mechanism that leads to BAT thermogenesis remains elusive. Chen et al. demonstrate that uncoupling protein 1 is not essential for the antibody bFKB1 to stimulate BAT thermogenesis in obese mice. They show that bFKB1 acts as an FGF21 mimetic protein whose in vivo metabolic activity originates outside of the adipocytes.|
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Objective: Fibroblast Growth Factor 21 (FGF21) is a potent stimulator of brown fat thermogenesis that improves insulin sensitivity, ameliorates hepatosteatosis, and induces weight loss by engaging the receptor complex comprised of Fibroblast Growth Factor Receptor 1 (FGFR1) and the requisite coreceptor βKlotho. Previously, recombinant antibody proteins that activate the FGFR1/βKlotho complex were proposed to act as an FGF21-mimetic; however, in vivo action of these engineered proteins has not been well studied.
Methods: We investigated the mechanism by which anti-FGFR1/βKlotho bispecific antibody (bFKB1) stimulates thermogenesis in UCP1-expressing brown adipocytes using genetically engineered mice. Anti-FGFR1 agonist antibody was also used to achieve brown adipose tissue restricted activation in transgenic mice.
Results: Studies with global Ucp1-deficient mice and adipose-specific Fgfr1 deficient mice demonstrated that bFKB1 acts on targets distal to adipocytes and indirectly stimulates brown adipose thermogenesis in a UCP1-independent manner. Using a newly developed transgenic system, we also show that brown adipose tissue restricted activation of a transgenic FGFR1 expressed under the control of Ucp1 promoter does not stimulate energy expenditure. Finally, consistent with its action as a FGF21 mimetic, bFBK1 suppresses intake of saccharin-containing food and alcohol containing water in mice.
Conclusions: Collectively, we propose that FGFR1/βKlotho targeted therapy indeed mimics the action of FGF21 in vivo and stimulates UCP1-independent brown fat thermogenesis through receptors outside of adipocytes and likely in the nervous system.[Hide abstract]
|The mitochondrial pyruvate carrier mediates increases in hepatic TCA cycle capacity Hepatocyte mitochondria provide energy for the whole body by supporting the synthesis of new glucose during fasting Hepatocyte mitochondrial pyruvate carrier (MPC) activity may play a fundamental role in the aberrant metabolism underlying type 2 diabetes (T2D). Rauckhorst, Gray, and colleagues demonstrate that high-fat diet increases TCA cycle capacity and that this increase is MPC-dependent. Thus, by contributing to chronic hyperglycemia, fibrosis, and TCA cycle expansion, the hepatocyte MPC is a key mediator of the pathophysiology induced in the HFD model of T2D.|
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Objective: Excessive hepatic gluconeogenesis is a defining feature of type 2 diabetes (T2D). Most gluconeogenic flux is routed through mitochondria. The mitochondrial pyruvate carrier (MPC) transports pyruvate from the cytosol into the mitochondrial matrix, thereby gating pyruvate-driven gluconeogenesis. Disruption of the hepatocyte MPC attenuates hyperglycemia in mice during high fat diet (HFD)-induced obesity but exerts minimal effects on glycemia in normal chow diet (NCD)-fed conditions. The goal of this investigation was to test whether hepatocyte MPC disruption provides sustained protection from hyperglycemia during long-term HFD and the differential effects of hepatocyte MPC disruption on TCA cycle metabolism in NCD versus HFD conditions.
Methods: We utilized long-term high fat feeding, serial measurements of postabsorptive blood glucose and metabolomic profiling and 13C-lactate/13C-pyruvate tracing to investigate the contribution of the MPC to hyperglycemia and altered hepatic TCA cycle metabolism during HFD-induced obesity.
Results: Hepatocyte MPC disruption resulted in long-term attenuation of hyperglycemia induced by HFD. HFD increased hepatic mitochondrial pyruvate utilization and TCA cycle capacity in an MPC-dependent manner. Furthermore, MPC disruption decreased progression of fibrosis and levels of transcript markers of inflammation.
Conclusions: By contributing to chronic hyperglycemia, fibrosis, and TCA cycle expansion, the hepatocyte MPC is a key mediator of the pathophysiology induced in the HFD model of T2D.[Hide abstract]
|Amino acid sensing in hypothalamic tanycytes via umami taste receptors One population of hypothalamic cells that is potentially a key player in energy homeostasis is the tanycyte. As amino acids are important signals of satiety, determination of the ability of tanycytes to detect amino acids would be an important advance in understanding the possible functions of these cells. Lazutkaite et al. show that hypothalamic tanycytes are directly sensitive to a range of amino acids. Amino acids act on tanycytes via two receptors. These data warrant investigation as to whether tanycytes may be physiological mediators of satiety signals and act to reduce food intake.|
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Objective: Hypothalamic tanycytes are glial cells that line the wall of the third ventricle and contact the cerebrospinal fluid (CSF). While they are known to detect glucose in the CSF we now show that tanycytes also detect amino acids, important nutrients that signal satiety.
Methods: Ca2+ imaging and ATP biosensing were used to detect tanycyte responses to l-amino acids. The downstream pathway of the responses was determined using ATP receptor antagonists and channel blockers. The receptors were characterized using mice lacking the Tas1r1 gene, as well as an mGluR4 receptor antagonist.
Results: Amino acids such as Arg, Lys, and Ala evoke Ca2+ signals in tanycytes and evoke the release of ATP via pannexin 1 and CalHM1, which amplifies the signal via a P2 receptor dependent mechanism. Tanycytes from mice lacking the Tas1r1 gene had diminished responses to lysine and arginine but not alanine. Antagonists of mGluR4 greatly reduced the responses to alanine and lysine.
Conclusions: Two receptors previously implicated in taste cells, the Tas1r1/Tas1r3 heterodimer and mGluR4, contribute to the detection of a range of amino acids by tanycytes in CSF.[Hide abstract]
|The FGF21 response to fructose predicts metabolic health Concerns about the consumption of fructose have been raised as high fructose intake may contribute to the current epidemics of obesity and its metabolic complications. Fructose is preferentially metabolized by the liver and can increase hepatic expression of fibroblast growth factor 21 (FGF21). ter Horst, Gilijamse et al. find that fructose-FGF21 responsiveness is exaggerated in subjects with poor metabolic health as reflected by the associations with elevated endogenous glucose production, increased lipolysis, and insulin resistance. They also demonstrate that the FGF21 response to fructose persists in post-bariatric subjects.|
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Objective: Fructose consumption has been implicated in the development of obesity and insulin resistance. Emerging evidence shows that fibroblast growth factor 21 (FGF21) has beneficial effects on glucose, lipid, and energy metabolism and may also mediate an adaptive response to fructose ingestion. Fructose acutely stimulates circulating FGF21 consistent with a hormonal response. We aimed to evaluate whether fructose-induced FGF21 secretion is linked to metabolic outcomes in obese humans before and after bariatric surgery-induced weight loss.
Methods: We recruited 40 Roux-en-Y gastric bypass patients and assessed the serum FGF21 response to fructose (75-g fructose tolerance test) and basal and insulin-mediated glucose and lipid fluxes during a 2-step hyperinsulinemic-euglycemic clamp with infusion of [6,6-2H2] glucose and [1,1,2,3,3-2H5] glycerol. Liver biopsies were obtained during bariatric surgery. Nineteen subjects underwent the same assessments at 1-year follow-up.
Results: Serum FGF21 increased 3-fold at 120 min after fructose ingestion and returned to basal levels at 300 min. Neither basal FGF21 nor the fructose-FGF21 response correlated with liver fat content or liver histopathology, but increased levels were associated with elevated endogenous glucose production, increased lipolysis, and peripheral/muscle insulin resistance. At 1-year follow-up, subjects had lost 28 ± 6% of body weight and improved in all metabolic outcomes, but fructose-stimulated FGF21 dynamics did not markedly differ from the pre-surgical state. The association between increased basal and stimulated FGF21 levels with poor metabolic health was no longer present after weight loss.
Conclusions: Fructose ingestion in obese humans stimulates FGF21 secretion, and this response is related to systemic metabolism. Further studies are needed to establish if FGF21 signaling is (patho)physiologically involved in fructose metabolism and metabolic health.[Hide abstract]
|Maternal obesity alters fatty acid oxidation, AMPK activity, and associated DNA methylation Epidemiological data indicate that obesity during pregnancy is an important contributor to adiposity and metabolic disease risk in the offspring. Boyle et al. investigated cellular lipid metabolism and AMP-activated protein kinase (AMPK) activity in infant mesenchymal stem cells (MSCs) undergoing myogenesis in vitro. They identified maternal obesity-associated reductions in offspring MSC lipid metabolism and AMPK activity. The authors will continue to follow these children longitudinally to be better able to address whether umbilical cord MSC metabolism is predictive of future weight gain or adiposity patterns.|
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Objective: Infants born to mothers with obesity have greater adiposity, ectopic fat storage, and are at increased risk for childhood obesity and metabolic disease compared with infants of normal weight mothers, though the cellular mechanisms mediating these effects are unclear.
Methods: We tested the hypothesis that human, umbilical cord-derived mesenchymal stem cells (MSCs) from infants born to obese (Ob-MSC) versus normal weight (NW-MSC) mothers demonstrate altered fatty acid metabolism consistent with adult obesity. In infant MSCs undergoing myogenesis in vitro, we measured cellular lipid metabolism and AMPK activity, AMPK activation in response to cellular nutrient stress, and MSC DNA methylation and mRNA content of genes related to oxidative metabolism.
Results: We found that Ob-MSCs exhibit greater lipid accumulation, lower fatty acid oxidation (FAO), and dysregulation of AMPK activity when undergoing myogenesis in vitro. Further experiments revealed a clear phenotype distinction within the Ob-MSC group where more severe MSC metabolic perturbation corresponded to greater neonatal adiposity and umbilical cord blood insulin levels. Targeted analysis of DNA methylation array revealed Ob-MSC hypermethylation in genes regulating FAO (PRKAG2, ACC2, CPT1A, SDHC) and corresponding lower mRNA content of these genes. Moreover, MSC methylation was positively correlated with infant adiposity.
Conclusions: These data suggest that greater infant adiposity is associated with suppressed AMPK activity and reduced lipid oxidation in MSCs from infants born to mothers with obesity and may be an important, early marker of underlying obesity risk.[Hide abstract]
|Decreasing CB1 receptor signaling improves insulin sensitivity Liver resident macrophages, called Kupffer cells (KCs), are thought to be the major source of hepatic inflammation. Cannabinoid 1 receptor (CB1R) has a proinflammatory function in macrophages and CB1R signaling is strongly involved in the development of fatty liver and insulin resistance. Jourdan et al. demonstrate that knock-down of CB1R in Kupffer cells leads to improved global insulin sensitivity by reducing inflammation and reactive oxygen species production and by promoting mitochondria uncoupling through an increase in uncoupling protein 2 activity.|
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Objective: Obesity-induced accumulation of ectopic fat in the liver is thought to contribute to the development of insulin resistance, and increased activity of hepatic CB1R has been shown to promote both processes. However, lipid accumulation in liver can be experimentally dissociated from insulin resistance under certain conditions, suggesting the involvement of additional mechanisms. Obesity is also associated with pro-inflammatory changes which, in turn, can promote insulin resistance. Kupffer cells (KCs), the liver's resident macrophages, are the major source of pro-inflammatory cytokines in the liver, such as TNF-α, which has been shown to inhibit insulin signaling in multiple cell types, including hepatocytes. Here, we sought to identify the role of CB1R in KCs in obesity-induced hepatic insulin resistance.
Methods: We used intravenously administered β-D-glucan-encapsulated siRNA to knock-down CB1R gene expression selectively in KCs.
Results: We demonstrate that a robust knock-down of the expression of Cnr1, the gene encoding CB1R, results in improved glucose tolerance and insulin sensitivity in diet-induced obese mice, without affecting hepatic lipid content or body weight. Moreover, Cnr1 knock-down in KCs was associated with a shift from pro-inflammatory M1 to anti-inflammatory M2 cytokine profile and improved insulin signaling as reflected by increased insulin-induced Akt phosphorylation.
Conclusions: These findings suggest that CB1R expressed in KCs plays a critical role in obesity-related hepatic insulin resistance via a pro-inflammatory mechanism.[Hide abstract]
|KLK5 induces shedding of DPP4 from circulatory Th17 cells Increasing plasma levels and activity of dipeptidyl peptidase-4 DPP4 are associated with rapid progression of metabolic syndrome to overt type 2 diabetes mellitus (T2DM). While DPP4 inhibitors are increasingly used as anti-hyperglycemic agents, the reason for the increase in plasma DPP4 activity in T2DM patients remains elusive. Nargis and colleagues show that in T2DM patients, circulating CD4+ T cells, specifically cells having the Th17 phenotype, shed cleaved DPP4 protein into plasma due to the enzymatic action of kallikrein-related peptidase (KLK5). Thus, they uncovered a hitherto unknown link between T cell inflammation and aberrant plasma DPP4 abundance in T2DM.|
Abstract | PDF
Objective: Increasing plasma levels and activity of dipeptidyl peptidase-4 (DPP4 or CD26) are associated with rapid progression of metabolic syndrome to overt type 2 diabetes mellitus (T2DM). While DPP4 inhibitors are increasingly used as anti-hyperglycemic agents, the reason for the increase in plasma DPP4 activity in T2DM patients remains elusive.
Methods: We looked into the source of plasma DPP4 activity in a cohort of 135 treatment naive nonobese (BMI < 30) T2DM patients. A wide array of ex vivo, in vitro, and in silico methods were employed to study enzyme activity, gene expression, subcellular localization, protease identification, surface expression, and protein–protein interactions.
Results: We show that circulating immune cells, particularly CD4+ T cells, served as an important source for the increase in plasma DPP4 activity in T2DM. Moreover, we found kallikrein-related peptidase 5 (KLK5) as the enzyme responsible for cleaving DPP4 from the cell surface by directly interacting with the extracellular loop. Expression and secretion of KLK5 is induced in CD4+ T cells of T2DM patients. In addition, KLK5 shed DPP4 from circulating CD4+ T helper (Th)17 cells and shed it into the plasma of T2DM patients. Similar cleavage and shedding activities were not seen in controls.
Conclusions: Our study provides mechanistic insights into the molecular interaction between KLK5 and DPP4 as well as CD4+ T cell derived KLK5 mediated enzymatic cleavage of DPP4 from cell surface. Thus, our study uncovers a hitherto unknown cellular source and mechanism behind enhanced plasma DPP4 activity in T2DM.[Hide abstract]
|Bombesin-like receptor 3 expression in glutamatergic neurons is required for regulation of energy metabolismBombesin-like receptor 3 (BRS-3) is an orphan G protein-coupled receptor. Insights into the function of BRS-3 come from a Brs3 knockout (KO) mouse. The null phenotype includes obesity, increased food intake and meal size, and reductions in metabolic rate, resting body temperature, and resting heart rate. Xiao and colleagues developed mice that allow selective, conditional deletion or re-expression of Brs3. They used these mice to investigate the necessity and sufficiency of Brs3 in glutamatergic and GABAergic neurons for regulation of energy homeostasis and identified a role for Brs3 in glutamatergic but not GABAergic neurons.|
Abstract | PDF
Objective: Bombesin-like receptor 3 (BRS-3) is an orphan G protein-coupled receptor. Brs3 null mice have reduced resting metabolic rate and body temperature, increased food intake, and obesity. Here we study the role of Brs3 in different neuron types.
Methods: Mice able to undergo Cre recombinase-dependent inactivation or re-expression of Brs3 were generated, respectively Brs3fl/y and Brs3loxTB/y. We then studied four groups of mice with Brs3 selectively inactivated or re-expressed in cells expressing Vglut2-Cre or Vgat-Cre.
Results: Deletion of Brs3 in glutamatergic neurons expressing Vglut2 reproduced the global null phenotype for regulation of food intake, metabolic rate, body temperature, adiposity, and insulin resistance. These mice also no longer responded to a BRS-3 agonist, MK-5046. In contrast, deletion of Brs3 in GABAergic neurons produced no detectable phenotype. Conversely, the wild type phenotype was restored by selective re-expression of Brs3 in glutamatergic neurons, with no normalization achieved by re-expressing Brs3 in GABAergic neurons.
Conclusions: Brs3 expression in glutamatergic neurons is both necessary and sufficient for full Brs3 function in energy metabolism. In these experiments, no function was identified for Brs3 in GABAergic neurons. The data suggest that the anti-obesity pharmacologic actions of BRS-3 agonists occur via agonism of receptors on glutamatergic neurons.[Hide abstract]
|Adult neural stem cell fate is determined by activation of mitochondrial metabolismNeurogenesis persists in the brain of adult mammals in two well-defined niches, the hippocampus and the sub-ventricular zone. Neural stem cells (NSCs) principally rely on aerobic glycolysis before differentiation, contrasting with mature cells, in which metabolism is mainly based on mitochondrial oxidative phosphorylation (OXPHOS). Gothié et al. hypothesized that thyroid hormone (TH) determination of NSC fate could implicate modulation of the glycolysis to OXPHOS metabolism transition. Their data show that TH signaling increases mitochondrial dynamics and activates mitochondrial respiration during NSC differentiation, thus directing adult NSC determination to a neuronal fate.|
Abstract | PDF
Objective: In the adult brain, neural stem cells (NSCs) located in the subventricular zone (SVZ) produce both neuronal and glial cells. Thyroid hormones (THs) regulate adult NSC differentiation towards a neuronal phenotype, but also have major roles in mitochondrial metabolism. As NSC metabolism relies mainly on glycolysis, whereas mature cells preferentially use oxidative phosphorylation, we studied how THs and mitochondrial metabolism interact on NSC fate determination.
Methods: We used a mitochondrial membrane potential marker in vivo to analyze mitochondrial activity in the different cell types in the SVZ of euthyroid and hypothyroid mice. Using primary adult NSC cultures, we analyzed ROS production, SIRT1 expression, and phosphorylation of DRP1 (a mitochondrial fission mediator) as a function of TH availability.
Results: We observed significantly higher mitochondrial activity in cells adopting a neuronal phenotype in vivo in euthyroid mice. However, prolonged hypothyroidism reduced not only neuroblast numbers but also their mitochondrial activity. In vitro studies showed that TH availability favored a neuronal phenotype and that blocking mitochondrial respiration abrogated TH-induced neuronal fate determination. DRP1 phosphorylation was preferentially activated in cells within the neuronal lineage and was stimulated by TH availability.
Conclusions: These results indicate that THs favor NSC fate choice towards a neuronal phenotype in the adult mouse SVZ through effects on mitochondrial metabolism.[Hide abstract]