Featured ArticlesVolume 11 | May 2018
|Adipocyte Xbp1s Overexpression Drives Uridine Production and Reduces Obesity Xbp1s is a transcriptional factor that is elevated in subcutaneous fat tissue from obese human subjects. Under fasting conditions, when there are high levels of lipolysis, pyrimidine biosynthesis in adipocytes is elevated. As a result, plasma uridine concentrations double, which has major implications for temperature regulation and energy expenditure. Deng et al. find that adipocyte overexpression of Xbp1s leads to profound loss of fat mass, and this loss is associated with elevation of uridine in adipose tissue and blood circulation. They demonstrate that Xbp1s triggered fat mass loss relies on pyrimidine biosynthesis. Hence, the stimulation of the adipocyte uridine synthesis pathway is a potential therapeutic target area for the treatment of obesity.|
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Objective: The spliced transcription factor Xbp1 (Xbp1s), a transducer of the unfolded protein response (UPR), regulates lipolysis. Lipolysis is stimulated by fasting when uridine synthesis is also activated in adipocytes.
Methods: Here we have examined the regulatory role Xbp1s in stimulation of uridine biosynthesis in adipocytes and triglyceride mobilization using inducible mouse models.
Results: Xbp1s is a key molecule involved in adipocyte uridine biosynthesis and release by activation of carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, dihydroorotase (CAD), the rate-limiting enzyme for UMP biosynthesis. Adipocyte Xbp1s overexpression drives energy mobilization and protects mice from obesity through activation of the pyrimidine biosynthesis pathway.
Conclusions: These observations reveal that Xbp1s is a potent stimulator of uridine production in adipocytes, enhancing lipolysis and invoking a potential anti-obesity strategy through the induction of a futile biosynthetic cycle.[Hide abstract]
|Epoxygenase inactivation exacerbates metabolic dysfunction resulting from impaired adipogenesisEpoxyeicosatrienoic acids (EET) are synthesized by epoxygenases. The biological effects of EETs are pleiotropic, ranging from anti-inflammatory and cardioprotective actions to a regulatory role in cancer, organ/tissue regeneration, and embryonic hematopoiesis. Olona, Terra et al. find that Cyp2j4, the rat orthologue of human CYP2J2, which codes for an expoxygenase, is essential for maintaining a healthy adipogenesis status. Knockout of Cyp2j4 combined with the metabolic challenges aging or cafeteria diet caused adipocyte dysfunction.|
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Objective: When molecular drivers of healthy adipogenesis are perturbed, this can cause hepatic steatosis. The role of arachidonic acid (AA) and its downstream enzymatic cascades, such as cyclooxygenase, in adipogenesis is well established. The exact contribution of the P450 epoxygenase pathway, however, remains to be established. Enzymes belonging to this pathway are mainly encoded by the CYP2J locus which shows extensive allelic expansion in mice. Here we aimed to establish the role of endogenous epoxygenase during adipogenesis under homeostatic and metabolic stress conditions.
Methods: We took advantage of the simpler genetic architecture of the Cyp2j locus in the rat and used a Cyp2j4 (orthologue of human CYP2J2) knockout rat in two models of metabolic dysfunction: physiological aging and cafeteria diet (CAF). The phenotyping of Cyp2j4−/− rats under CAF was integrated with proteomics (LC-MS/MS) and lipidomics (LC-MS) analyses in the liver and the adipose tissue.
Results: We report that Cyp2j4 deletion causes adipocyte dysfunction under metabolic challenges. This is characterized by (i) down-regulation of white adipose tissue (WAT) PPARγ and C/EBPα, (ii) adipocyte hypertrophy, (iii) extracellular matrix remodeling, and (iv) alternative usage of AA pathway. Specifically, in Cyp2j4−/− rats treated with a cafeteria diet, the dysfunctional adipogenesis is accompanied by exacerbated weight gain, hepatic lipid accumulation, and dysregulated gluconeogenesis.
Conclusions: These results suggest that AA epoxygenases are essential regulators of healthy adipogenesis. Our results uncover their synergistic role in fine-tuning AA pathway in obesity-mediated hepatic steatosis.[Hide abstract]
|Loss of GLP-1 signaling reduces BAT thermogenesis and increases adiposityThe intestine and brain both produce glucagon-like peptide-1 (GLP-1), which plays an important role in the control of food intake and glycemia. The dorsomedial nucleus of the hypothalamus (DMH), a key part of the sympathetic control network, expresses GLP-1 receptor (GLP-1R) and is densely innervated by GLP-1 fibers, suggesting a possible role of DMH GLP-1R signaling in sympathetic nervous system regulation. Lee et al. find an important link between hindbrain GLP-1 projections into the DMH and sympathetic regulation of brown adipose tissue thermogenesis and energy balance.|
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Objective: Glucagon-like peptide-1 (GLP-1) neurons in the hindbrain densely innervate the dorsomedial hypothalamus (DMH), a nucleus strongly implicated in body weight regulation and the sympathetic control of brown adipose tissue (BAT) thermogenesis. Therefore, DMH GLP-1 receptors (GLP-1R) are well placed to regulate energy balance by controlling sympathetic outflow and BAT function.
Methods: We investigate this possibility in adult male rats by using direct administration of GLP-1 (0.5 ug) into the DMH, knocking down DMH GLP-1R mRNA with viral-mediated RNA interference, and by examining the neurochemical phenotype of GLP-1R expressing cells in the DMH using in situ hybridization.
Results: GLP-1 administered into the DMH increased BAT thermogenesis and hepatic triglyceride (TG) mobilization. On the other hand, Glp1r knockdown (KD) in the DMH increased body weight gain and adiposity, with a concomitant reduction in energy expenditure (EE), BAT temperature, and uncoupling protein 1 (UCP1) expression. Moreover, DMH Glp1r KD induced hepatic steatosis, increased plasma TG, and elevated liver specific de-novo lipogenesis, effects that collectively contributed to insulin resistance. Interestingly, DMH Glp1r KD increased neuropeptide Y (NPY) mRNA expression in the DMH. GLP-1R mRNA in the DMH, however, was found in GABAergic not NPY neurons, consistent with a GLP-1R-dependent inhibition of NPY neurons that is mediated by local GABAergic neurons. Finally, DMH Glp1r KD attenuated the anorexigenic effects of the GLP-1R agonist exendin-4, highlighting an important role of DMH GLP-1R signaling in GLP-1-based therapies.
Conclusions: Collectively, our data show that DMH GLP-1R signaling plays a key role for BAT thermogenesis and adiposity.[Hide abstract]
|The Neurochemical Signature of Beige Fat InnervationSubcutaneous white adipose tissue, particularly in the inguinal depot (iWAT) can undergo a process of “beiging” in which thermogenic, mitochondria-rich, UCP1-expressing multilocular cells develop within white adipocytes. However, the neurochemical signals from nerves targeting iWAT that induce this change are unknown. Stefanidis et al. find that there is a significant shift in the gene expression profile in neurons directly innervating iWAT, concomitant with the beiging process, to a signature which is more aligned with that of neurons innervating brown adipose tissue. The most likely candidate to mediate the beiging process is α-melanocyte stimulating hormone.
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Objective: The potential for brown adipose tissue (BAT) to be targeted as a therapeutic option to combat obesity has been heightened by the discovery of a brown–like form of inducible “beige” adipose tissue in white fat which has overlapping structural and functional properties to “classical” BAT. The likelihood that both beige and brown fat are recruited functionally by neural mechanisms, taken together with the lack of a detailed understanding of the nature of changes in the nervous system when white adipose tissue (WAT) is transformed to brown, provides the impetus for this study. Here, we aim to identify whether there is a shift in the gene expression profile in neurons directly innervating inguinal white adipose tissue (iWAT) that has undergone “beiging” to a signature that is more similar to neurons projecting to BAT.
Methods: Two groups of rats, one housed at thermoneutrality (27 °C) and the other exposed to cold (8 °C) for 7 days, were killed, and their T13/L1 ganglia, stellate ganglion (T1/T2), or superior cervical ganglion (SCG, C2/3) removed. This approach yielded ganglia containing neurons that innervate either beiged white fat (8 °C for 7 days), inguinal WAT (27 °C for 7 days), BAT (both 27 °C and 8 °C for 7 days) or non-WAT (8 °C for 7 days), the latter included to isolate changes in gene expression that were more aligned with a response to cold exposure than the transformation of white to beige adipocytes. Bioinformatics analyses of RNA sequencing data was performed followed by Ingenuity Pathway Analysis (IPA) to determine differential gene expression and recruitment of biosynthetic pathways.
Results: When iWAT is “beiged” there is a significant shift in the gene expression profile of neurons in sympathetic ganglia (T13/L1) innervating this depot toward a gene neurochemical signature that is similar to the stellate ganglion projecting to BAT. Bioinformatics analyses of “beiging” related genes revealed upregulation of genes encoding neuropeptides proopiomelanocortin (POMC) and calcitonin-gene related peptide (CGRP) within ganglionic neurons. Treatment of differentiated 3T3L1 adipocytes with αMSH, one of the products cleaved from POMC, results in an elevation in lipolysis and the beiging of these cells as indicated by changes in gene expression markers of browning (Ucp1 and Ppargc1a).
Conclusions: These data indicate that, coincident with beiging, there is a shift toward a “brown-like” neurochemical signature of postganglionic neurons projecting to inguinal white fat, an increased expression of POMC, and, consistent with a causative role for this prohormone in beiging, an αMSH-mediated increase in beige gene markers in isolated adipocytes.[Hide abstract]
|BAT derived ANGPTL4 controls glucose and lipid metabolism and thermogenesisBrown adipose tissue (BAT) is fueled by fatty acids provided by lipoprotein lipase (LPL). One of the important modulators of LPL activity is angiopoietin-like 4 (ANGPTL4). Singh et al. generated a novel mouse model lacking ANGPTL4 specifically in BAT (BAT-KO). Their results demonstrate that BAT derived ANGPTL4 is an important regulator of lipoprotein metabolism, glucose homeostasis, and insulin sensitivity. ANGPTL4 deficiency in BAT improves oxidative metabolism and the thermogenic function of brown adipocytes.|
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Objective: Brown adipose tissue (BAT) controls triglyceride-rich lipoprotein (TRL) catabolism. This process is mediated by the lipoprotein lipase (LPL), an enzyme that catalyzed the hydrolysis of triglyceride (TAG) in glycerol and fatty acids (FA), which are burned to generate heat. LPL activity is regulated by angiopoietin-like 4 (ANGPTL4), a secretory protein produced in adipose tissues (AT), liver, kidney, and muscle. While the role of ANGPTL4 in regulating lipoprotein metabolism is well established, the specific contribution of BAT derived ANGPTL4 in controlling lipid and glucose homeostasis is not well understood.
Methods and results: We generated a novel mouse model lacking ANGPTL4 specifically in brown adipose tissue (BAT-KO). Here, we report that specific deletion of ANGPTL4 in BAT results in enhanced LPL activity, circulating TAG clearance and thermogenesis. Absence of ANGPTL4 in BAT increased FA oxidation and reduced FA synthesis. Importantly, we observed that absence of ANGPTL4 in BAT leads to a remarkable improvement in glucose tolerance in short-term HFD feeding.
Conclusions: Our findings demonstrate an important role of BAT derived ANGPTL4 in regulating lipoprotein metabolism, whole-body lipid and glucose metabolism, and thermogenesis during acute cold exposure.[Hide abstract]
|Enterochromaffin 5-HT Cells - a major Target for GLP-1 and Gut Microbial MetabolitesThe enterochromaffin (EC) cell is the most abundant cell type among the enteroendocrine cells and its main secretory product is the biogenic amine serotonin (5-HT). Lund and colleagues characterize the expression of receptors that function as sensors of luminal stimuli in EC cells from the small intestine and the colon. They find that the receptor for the gut hormone glucagon-like peptide-1 (GLP-1) is particularly highly expressed and enriched in EC cells and that GLP-1 efficiently stimulates 5-HT release. Also, they find the colonic EC cells to express a large repertoire of known and proposed sensors of microbial metabolites including several receptors for short chain fatty acids.|
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Objective: 5-HT storing enterochromaffin (EC) cells are believed to respond to nutrient and gut microbial components, and 5-HT receptor-expressing afferent vagal neurons have been described to be the major sensors of nutrients in the GI-tract. However, the molecular mechanism through which EC cells sense nutrients and gut microbiota is still unclear.
Methods and results: TPH1, the 5-HT generating enzyme, and chromogranin A, an acidic protein responsible for secretory granule storage of 5-HT, were highly enriched in FACS-purified EC cells from both small intestine and colon using a 5-HT antibody-based method. Surprisingly, EC cells from the small intestine did not express GPCR sensors for lipid and protein metabolites, such as FFAR1, GPR119, GPBAR1 (TGR5), CaSR, and GPR142, in contrast to the neighboring GLP-1 storing enteroendocrine cell. However, the GLP-1 receptor was particularly highly expressed and enriched in EC cells as judged both by qPCR and by immunohistochemistry using a receptor antibody. GLP-1 receptor agonists robustly stimulated 5-HT secretion from intestinal preparations using both HPLC and a specific amperometric method. Colonic EC cells expressed many different types of known and potential GPCR sensors of microbial metabolites including three receptors for SCFAs, i.e. FFAR2, OLF78, and OLF558 and receptors for aromatic acids, GPR35; secondary bile acids GPBAR1; and acyl-amides and lactate, GPR132.
Conclusions: Nutrient metabolites apparently do not stimulate EC cells of the small intestine directly but through a paracrine mechanism involving GLP-1 secreted from neighboring enteroendocrine cells. In contrast, colonic EC cells are able to sense a multitude of different metabolites generated by the gut microbiota as well as gut hormones, including GLP-1.[Hide abstract]
|Bile acids are important regulators of appetite- and metabolism-regulating hormones Recent studies have indicated that bile acids (BA), in addition to their well-known role in fat absorption, may stimulate the secretion of a number of appetite- and metabolism-regulating peptide hormones from the gut and pancreas. Kuhre et al. sought to evaluate the rat as a model for BA effects by examining the effects of BAs on the secretion of the most important appetite- and metabolism-regulating hormones. Their study shows that BAs have marked effects on the secretion of these hormones and, therefore, in addition to their role as fat emulsifiers, should be regarded as important regulators of blood glucose and metabolism.|
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Objective: Bile acids (BAs) facilitate fat absorption and may play a role in glucose and metabolism regulation, stimulating the secretion of gut hormones. The relative importance and mechanisms involved in BA-stimulated secretion of appetite and metabolism regulating hormones from the gut and pancreas is not well described and was the purpose of this study.
Methods: The effects of bile acids on the secretion of gut and pancreatic hormones was studied in rats and compared to the most well described nutritional secretagogue: glucose. The molecular mechanisms that underlie the secretion was studied by isolated perfused rat and mouse small intestine and pancreas preparations and supported by immunohistochemistry, expression analysis, and pharmacological studies.
Results: Bile acids robustly stimulate secretion of not only the incretin hormones, glucose-dependent insulinotropic peptide (GIP), and glucagon-like peptide-1 (GLP-1), but also glucagon and insulin in vivo, to levels comparable to those resulting from glucose stimulation. The mechanisms of GLP-1, neurotensin, and peptide YY (PYY) secretion was secondary to intestinal absorption and depended on activation of basolateral membrane Takeda G-protein receptor 5 (TGR5) receptors on the L-cells in the following order of potency: Lithocholic acid (LCA) >Deoxycholicacid (DCA)>Chenodeoxycholicacid (CDCA)> Cholic acid (CA). Thus BAs did not stimulate secretion of GLP-1 and PYY from perfused small intestine in TGR5 KO mice but stimulated robust responses in wild type littermates. TGR5 is not expressed on α-cells or β-cells, and BAs had no direct effects on glucagon or insulin secretion from the perfused pancreas.
Conclusions: BAs should be considered not only as fat emulsifiers but also as important regulators of appetite- and metabolism-regulating hormones by activation of basolateral intestinal TGR5.[Hide abstract]
|FGF21 increases after acute alcohol ingestion and sustained binge drinking at OktoberfestIn mice, alcohol increases circulating fibroblast growth factor 21 (FGF21) levels and FGF21 overexpression or continuous infusion reduces alcohol and sweet intake. Søberg, Andersen et al. tested the hypothesis that in humans, alcohol ingestion increases hepatic secretion of FGF21, which then suppresses further alcohol intake. To do this, they assessed plasma FGF21 levels in fasted humans after acute and sub-chronic alcohol ingestion and measured the effects of bolus human FGF21 administration on alcohol preference in mice. They further correlated fasting FGF21 levels with self-reported alcohol-related behaviors, emotional responses, and problems in a cohort of 49 healthy human subjects. Their findings support the notion that FGF21 may be a liver-derived inhibitor of alcohol intake.|
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Objective: Excessive alcohol consumption is a leading cause of global morbidity and mortality. However, knowledge of the biological factors that influence ad libitum alcohol intake may be incomplete. Two large studies recently linked variants in the KLB locus with levels of alcohol intake in humans. KLB encodes β-klotho, co-receptor for the liver-derived hormone fibroblast growth factor 21 (FGF21). In mice, FGF21 reduces alcohol intake, and human Fgf21 variants are enriched among heavy drinkers. Thus, the liver may limit alcohol consumption by secreting FGF21. However, whether full-length, active plasma FGF21 (FGF21 (1–181)) levels in humans increase acutely or sub-chronically in response to alcohol ingestion is uncertain.
Methods: We recruited 10 healthy, fasted male subjects to receive an oral water or alcohol bolus with concurrent blood sampling for FGF21 (1–181) measurement in plasma. In addition, we measured circulating FGF21 (1–181) levels, liver stiffness, triglyceride, and other metabolic parameters in three healthy Danish men before and after consuming an average of 22.6 beers/person/day (4.4 g/kg/day of ethanol) for three days during Oktoberfest 2017 in Munich, Germany. We further correlated fasting FGF21 (1–181) levels in 49 healthy, non-alcoholic subjects of mixed sex with self-reports of alcohol-related behaviors, emotional responses, and problems. Finally, we characterized the effect of recombinant human FGF21 injection on ad libitum alcohol intake in mice.
Results: We show that alcohol ingestion (25.3 g or ∼2.5 standard drinks) acutely increases plasma levels of FGF21 (1–181) 3.4-fold in fasting humans. We also find that binge drinking for three days at Oktoberfest is associated with a 2.1-fold increase in baseline FGF21 (1–181) levels, in contrast to minor deteriorations in metabolic and hepatic biomarkers. However, basal FGF21 (1–181) levels were not correlated with differences in alcohol-related behaviors, emotional responses, or problems in our non-alcoholic subjects. Finally, we show that once-daily injection of recombinant human FGF21 reduces ad libitum alcohol intake by 21% in mice.
Conclusions: FGF21 (1–181) is markedly increased in circulation by both acute and sub-chronic alcohol intake in humans, and reduces alcohol intake in mice. These observations are consistent with a role for FGF21 as an endocrine inhibitor of alcohol appetite in humans.[Hide abstract]
|E2F1 promotes gluconeogenesis and contributes to hyperglycemia during diabetesAberrant regulation of hepatic glucose production is a major contributor to the hyperglycemia observed in type 2 diabetes (T2D). The transcription factor E2F1 is essential for controlling liver metabolism, regulating cholesterol uptake and promoting lipid synthesis through transcriptional regulation of key lipogenic enzymes. Giralt, Denechaud et al. demonstrate that E2F1 also contributes to mammalian glucose homeostasis through the control of hepatic glucose production. Their results indicate that E2F1 is critical for inducing hyperglycemia in T2D. This suggests that reducing E2F1 activity could protect against obesity-induced hyperglycemia.|
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Objective: Aberrant hepatic glucose production contributes to the development of hyperglycemia and is a hallmark of type 2 diabetes. In a recent study, we showed that the transcription factor E2F1, a component of the cell cycle machinery, contributes to hepatic steatosis through the transcriptional regulation of key lipogenic enzymes. Here, we investigate if E2F1 contributes to hyperglycemia by regulating hepatic gluconeogenesis.
Methods: We use different genetic models to investigate if E2F1 regulates gluconeogenesis in primary hepatocytes and in vivo. We study the impact of depleting E2F1 or inhibiting E2F1 activity in diabetic mouse models to evaluate if this transcription factor contributes to hyperglycemia during insulin resistance. We analyze E2F1 mRNA levels in the livers of human diabetic patients to assess the relevance of E2F1 in human pathophysiology.
Results: Lack of E2F1 impaired gluconeogenesis in primary hepatocytes. Conversely, E2F1 overexpression increased glucose production in hepatocytes and in mice. Several genetic models showed that the canonical CDK4-RB1-E2F1 pathway is directly involved in this regulation. E2F1 mRNA levels were increased in the livers from human diabetic patients and correlated with the expression of the gluconeogenic enzyme Pck1. Genetic invalidation or pharmacological inhibition of E2F1 improved glucose homeostasis in diabetic mouse models.
Conclusions: Our study unveils that the transcription factor E2F1 contributes to mammalian glucose homeostasis by directly controlling hepatic gluconeogenesis. Together with our previous finding that E2F1 promotes hepatic steatosis, the data presented here show that E2F1 contributes to both hyperlipidemia and hyperglycemia in diabetes, suggesting that specifically targeting E2F1 in the liver could be an interesting strategy for therapies against type 2 diabetes.[Hide abstract]
|GHR-deficient pigs resemble the pathophysiology of human Laron syndrome and reveal altered liver signalingLaron syndrome (LS) is a rare, autosomal recessive, hereditary disorder caused by loss-of-function mutations in the growth hormone receptor (GHR) gene. Although mechanistic studies have been performed in cell lines derived from LS patients and healthy controls, animal models are of pivotal importance for understanding the pathophysiology of LS in vivo. Hinrichs and colleagues developed a GHR-deficient (GHR-KO) pig model and show that it resembles important aspects of LS pathophysiology and features altered activation of signaling cascades in the liver.|
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Objective: Laron syndrome (LS) is a rare, autosomal recessive disorder in humans caused by loss-of-function mutations of the growth hormone receptor (GHR) gene. To establish a large animal model for LS, pigs with GHR knockout (KO) mutations were generated and characterized.
Methods: CRISPR/Cas9 technology was applied to mutate exon 3 of the GHR gene in porcine zygotes. Two heterozygous founder sows with a 1-bp or 7-bp insertion in GHR exon 3 were obtained, and their heterozygous F1 offspring were intercrossed to produce GHR-KO, heterozygous GHR mutant, and wild-type pigs. Since the latter two groups were not significantly different in any parameter investigated, they were pooled as the GHR expressing control group. The characterization program included body and organ growth, body composition, endocrine and clinical-chemical parameters, as well as signaling studies in liver tissue.
Results: GHR-KO pigs lacked GHR and had markedly reduced serum insulin-like growth factor 1 (IGF1) levels and reduced IGF-binding protein 3 (IGFBP3) activity but increased IGFBP2 levels. Serum GH concentrations were significantly elevated compared with control pigs. GHR-KO pigs had a normal birth weight. Growth retardation became significant at the age of five weeks. At the age of six months, the body weight of GHR-KO pigs was reduced by 60% compared with controls. Most organ weights of GHR-KO pigs were reduced proportionally to body weight. However, the weights of liver, kidneys, and heart were disproportionately reduced, while the relative brain weight was almost doubled. GHR-KO pigs had a markedly increased percentage of total body fat relative to body weight and displayed transient juvenile hypoglycemia along with decreased serum triglyceride and cholesterol levels. Analysis of insulin receptor related signaling in the liver of adult fasted pigs revealed increased phosphorylation of IRS1 and PI3K. In agreement with the loss of GHR, phosphorylation of STAT5 was significantly reduced. In contrast, phosphorylation of JAK2 was significantly increased, possibly due to the increased serum leptin levels and increased hepatic leptin receptor expression and activation in GHR-KO pigs. In addition, increased mTOR phosphorylation was observed in GHR-KO liver samples, and phosphorylation studies of downstream substrates suggested the activation of mainly mTOR complex 2.
Conclusions: GHR-KO pigs resemble the pathophysiology of LS and are an interesting model for mechanistic studies and treatment trials.[Hide abstract]
|VEGF and GLUT1 are highly heritable, inversely correlated and affected by dietary fat Cognitive function is affected by macronutrient intake, with high fat diets and obesity being particularly deleterious. The central nervous system depends on glucose as its energy substrate, and brain glucose uptake is mediated by the glucose transporter GLUT1. A high fat diet in mice was shown to reduce GLUT1 expression in brain endothelia, leading to cognitive impairment. This reaction was counterbalanced by an increased production of vascular endothelial growth factor (VEGF), which increased GLUT1 expression again. Schüler et al. investigated the response of 92 human mono- and dizygous twins to an acute shift from low fat to high fat diet with respect to serum levels of VEGF and the expression of GLUT1. Their data indicate VEGF as a determining factor linking dietary fat intake to cognitive function in humans.|
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Objective: Reduction of brain glucose transporter GLUT1 results in severe neurological dysfunction. VEGF is required to restore and maintain brain glucose uptake across the blood brain barrier via GLUT1, which was shown to be acutely diminished in response to a high fat diet (HFD) in mice. The genetic and HFD-related regulation and association of VEGF and GLUT1 (SLC2A1) in humans was investigated in the NUtriGenomic Analysis in Twins (NUGAT) study.
Methods: 92 healthy and non-obese twins were standardized to a high-carbohydrate low-fat diet for 6 weeks before switched to a 6-week HFD under isocaloric conditions. Three clinical investigation days were conducted: after 6 weeks of low-fat diet and after 1 and 6 weeks of HFD. Serum VEGF and other cytokine levels were measured using ELISA. Gene expression in subcutaneous adipose tissue was assessed by quantitative Real-Time PCR. Genotyping was performed using microarray. The Auditory Verbal Learning Task was conducted to measure cognitive performance.
Results: In this human study, we showed that the environmental regulation of SLC2A1 expression and serum VEGF by HFD was inversely correlated and both factors showed strong heritability (>90%). In response to the HFD containing 45% fat, serum VEGF levels increased (P = 0.002) while SLC2A1 mRNA expression in adipose tissue decreased (P = 0.001). Higher BMI was additionally associated with lower SLC2A1 expression. AA-genotypes of the rs9472159 polymorphism, which explained ∼39% of the variation in circulating VEGF concentrations, showed significantly reduced serum VEGF levels (P = 6.4 × 10−11) but higher SLC2A1 expression (P = 0.009) in adipose tissue compared to CC/CA-genotypes after 6 weeks of HFD. Memory performance in AA-genotypes declined in response to the HFD compared to CC- and CA-genotypes.
Conclusions: The results provide evidence to suggest the translatability of the dietary regulation of VEGF and GLUT1 from mouse models to humans. Our data demonstrate that HFD induces a genetically determined and correlated decrease of GLUT1 and increase of VEGF which may affect memory performance.[Hide abstract]
|Statin Dose Reduction With Complementary Diet Therapy: A Pilot Study Of Personalized MedicineThe first line treatment for cardiovascular disease (CVD) prevention is LDL-cholesterol lowering by statins. A critical issue that undermines CVD prevention is the high prevalence of statin undertreatment due to real or perceived adverse effects. The combination of nutraceuticals with statin treatment could help overcome these limitations. Scolaro and colleagues evaluated the effects of daily consumption of a combined supplementation of omega-3 fatty acids, plant sterols, and polyphenols on biomarkers of inflammation, lipidemia, and oxidative stress in type 2 diabetic patients treated with statins. They then conducted a pilot study to evaluate the effects of statin dose reduction. The results suggest a potential for adding nutraceuticals to statin therapy for primary prevention.|
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Objective: Statin intolerance, whether real or perceived, is a growing issue in clinical practice. Our aim was to evaluate the effects of reduced-dose statin therapy complemented with nutraceuticals.
Methods: First phase: Initially, 53 type 2 diabetic statin-treated patients received a supplementation with fish oil (1.7 g EPA + DHA/day), chocolate containing plant sterols (2.2 g/day), and green tea (two sachets/day) for 6 weeks. Second phase: “Good responders” to supplementation were identified after multivariate analysis (n = 10), and recruited for a pilot protocol of statin dose reduction. “Good responders” were then provided with supplementation for 12 weeks: standard statin therapy was kept during the first 6 weeks and reduced by 50% from weeks 6–12.
Results: First phase: After 6 weeks of supplementation, plasma LDL-C (−13.7% ± 3.7, P = .002) and C-reactive protein (−35.5% ± 5.9, P = .03) were reduced. Analysis of lathosterol and campesterol in plasma suggested that intensity of LDL-C reduction was influenced by cholesterol absorption rate rather than its synthesis. Second phase: no difference was observed for plasma lipids, inflammation, cholesterol efflux capacity, or HDL particles after statin dose reduction when compared to standard therapy.
Conclusions: Although limited by the small sample size, our study demonstrates the potential for a new therapeutic approach combining lower statin dose and specific dietary compounds. Further studies should elucidate “good responders” profile as a tool for personalized medicine. This may be particularly helpful in the many patients with or at risk for CVD who cannot tolerate high dose statin therapy.[Hide abstract]
|Computational genome-wide screen connects miRNAs to obesity and type 2 diabetesMiRNAs are small non-coding RNAs which are known to bind to the 3’UTR region of specific mRNAs leading to an altered expression or translation of the corresponding target gene(s). Most miRNAs have hundreds of targets involved in different signaling cascades. Thus, they are potent regulators and suitable candidates in polygenic diseases like obesity or type 2 diabetes (T2D). Gottmann et al. introduce a computational framework miR-QTL-Scan that integrates miRNAs located in quantitative trait loci (QTL), target-prediction tools and databases of experimentally validated targets, transcriptome profiles, and pathway enrichment analysis to identify miRNAs that can be related to obesity and T2D.|
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Objective: Obesity and type 2 diabetes (T2D) arise from the interplay between genetic, epigenetic, and environmental factors. The aim of this study was to combine bioinformatics and functional studies to identify miRNAs that contribute to obesity and T2D.
Methods: A computational framework (miR-QTL-Scan) was applied by combining QTL, miRNA prediction, and transcriptomics in order to enhance the power for the discovery of miRNAs as regulative elements. Expression of several miRNAs was analyzed in human adipose tissue and blood cells and miR-31 was manipulated in a human fat cell line.
Results: In 17 partially overlapping QTL for obesity and T2D 170 miRNAs were identified. Four miRNAs (miR-15b, miR-30b, miR-31, miR-744) were recognized in gWAT (gonadal white adipose tissue) and six (miR-491, miR-455, miR-423-5p, miR-132-3p, miR-365-3p, miR-30b) in BAT (brown adipose tissue). To provide direct functional evidence for the achievement of the miR-QTL-Scan, miR-31 located in the obesity QTL Nob6 was experimentally analyzed. Its expression was higher in gWAT of obese and diabetic mice and humans than of lean controls. Accordingly, 10 potential target genes involved in insulin signaling and adipogenesis were suppressed. Manipulation of miR-31 in human SGBS adipocytes affected the expression of GLUT4, PPARγ, IRS1, and ACACA. In human peripheral blood mononuclear cells (PBMC) miR-15b levels were correlated to baseline blood glucose concentrations and might be an indicator for diabetes.
Conclusions: Thus, miR-QTL-Scan allowed the identification of novel miRNAs relevant for obesity and T2D.[Hide abstract]
|Skeletal muscle O-GlcNAc transferase is important for energy homeostasis and insulin sensitivityProtein modification by O-linked β-D-N-acetylglucosamine (O-GlcNAc) is a post-translational event that has been suggested to serve as an integrated cellular nutrient sensor. Dysregulated O-GlcNAc signaling in skeletal muscle is associated with the development of several metabolic diseases. Shi, Munk and colleagues show that skeletal muscle from type 2 diabetic humans has elevated O-GlcNAcylation levels compared with matched controls. Moreover, they show that knockout of O-GlcNAc transferase (OGT) in skeletal muscle in mice improves whole-body insulin sensitivity and increases energy expenditure.|
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Objective: Given that cellular O-GlcNAcylation levels are thought to be real-time measures of cellular nutrient status and dysregulated O-GlcNAc signaling is associated with insulin resistance, we evaluated the role of O-GlcNAc transferase (OGT), the enzyme that mediates O-GlcNAcylation, in skeletal muscle.
Methods: We assessed O-GlcNAcylation levels in skeletal muscle from obese, type 2 diabetic people, and we characterized muscle-specific OGT knockout (mKO) mice in metabolic cages and measured energy expenditure and substrate utilization pattern using indirect calorimetry. Whole body insulin sensitivity was assessed using the hyperinsulinemic euglycemic clamp technique and tissue-specific glucose uptake was subsequently evaluated. Tissues were used for histology, qPCR, Western blot, co-immunoprecipitation, and chromatin immunoprecipitation analyses.
Results: We found elevated levels of O-GlcNAc-modified proteins in obese, type 2 diabetic people compared with well-matched obese and lean controls. Muscle-specific OGT knockout mice were lean, and whole body energy expenditure and insulin sensitivity were increased in these mice, consistent with enhanced glucose uptake and elevated glycolytic enzyme activities in skeletal muscle. Moreover, enhanced glucose uptake was also observed in white adipose tissue that was browner than that of WT mice. Interestingly, mKO mice had elevated mRNA levels of Il15 in skeletal muscle and increased circulating IL-15 levels. We found that OGT in muscle mediates transcriptional repression of Il15 by O-GlcNAcylating Enhancer of Zeste Homolog 2 (EZH2).
Conclusions: Elevated muscle O-GlcNAc levels paralleled insulin resistance and type 2 diabetes in humans. Moreover, OGT-mediated signaling is necessary for proper skeletal muscle metabolism and whole-body energy homeostasis, and our data highlight O-GlcNAcylation as a potential target for ameliorating metabolic disorders.[Hide abstract]
|NAMPT-mediated NAD+ biosynthesis is indispensable for adipose tissue plasticity and development of obesityThe expression of nicotinamide phosphoribosyltransferase (NAMPT) is associated with obesity in humans. However, divergent patterns of NAMPT association suggest differing depot-specific NAMPT contributions in human fat and make it difficult to assign a positive or negative role of NAMPT in metabolic syndrome. Nørgaard Nielsen, Peics, Ma et al. sought to determine whether NAMPT plays a causal role in adipose tissue dynamics and the development of obesity. Their findings indicate that adipose NAMPT plays an essential role in handling dietary lipid to modulate fat tissue plasticity, food intake, and systemic glucose homeostasis.|
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Objective: The ability of adipose tissue to expand and contract in response to fluctuations in nutrient availability is essential for the maintenance of whole-body metabolic homeostasis. Given the nutrient scarcity that mammals faced for millions of years, programs involved in this adipose plasticity were likely evolved to be highly efficient in promoting lipid storage. Ironically, this previously advantageous feature may now represent a metabolic liability given the caloric excess of modern society. We speculate that nicotinamide adenine dinucleotide (NAD+) biosynthesis exemplifies this concept. Indeed NAD+/NADH metabolism in fat tissue has been previously linked with obesity, yet whether it plays a causal role in diet-induced adiposity is unknown. Here we investigated how the NAD+ biosynthetic enzyme nicotinamide phosphoribosyltransferase (NAMPT) supports adipose plasticity and the pathological progression to obesity.
Methods: We utilized a newly generated Nampt loss-of-function model to investigate the tissue-specific and systemic metabolic consequences of adipose NAD+ deficiency. Energy expenditure, glycemic control, tissue structure, and gene expression were assessed in the contexts of a high dietary fat burden as well as the transition back to normal chow diet.
Results: Fat-specific Nampt knockout (FANKO) mice were completely resistant to high fat diet (HFD)-induced obesity. This was driven in part by reduced food intake. Furthermore, HFD-fed FANKO mice were unable to undergo healthy expansion of adipose tissue mass, and adipose depots were rendered fibrotic with markedly reduced mitochondrial respiratory capacity. Yet, surprisingly, HFD-fed FANKO mice exhibited improved glucose tolerance compared to control littermates. Removing the HFD burden largely reversed adipose fibrosis and dysfunction in FANKO animals whereas the improved glucose tolerance persisted.
Conclusions: These findings indicate that adipose NAMPT plays an essential role in handling dietary lipid to modulate fat tissue plasticity, food intake, and systemic glucose homeostasis.[Hide abstract]
|Vertical Sleeve Gastrectomy Corrects Metabolic Perturbations in a Rat Model Bariatric surgery is currently the most effective strategy for weight loss. Vertical sleeve gastrectomy (VSG) is a procedure in which about 80% of the stomach along the greater curvature is removed. It results in sustained weight loss and rapid improvements in glucose and lipid metabolism. However, the mechanism underlying this success is unclear. Wood et al. tested the effects of VSG in a rat model selectively bred for low running capacity. They found that VSG caused weight loss and some metabolic improvements independent of any changes in exercise capacity. However, it seems that basal energy expenditure and fat and carbohydrate oxidation was reduced by VSG making these animals more efficient at storing energy, an effect that would limit weight loss.|
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Objective: Bariatric surgery is currently our most effective strategy at weight loss, yet the mechanisms for its success remain unknown. Low exercise capacity, in humans and rodents, predicts poor metabolic outcome. The objective of this manuscript was to determine if bariatric surgery could restore metabolic perturbations in rats with low intrinsic exercise capacity.
Methods: We performed vertical sleeve gastrectomy (VSG) or sham surgery in high fat-fed rats selectively bred for low running capacity.
Results: We found that VSG reduced body mass through a reduction in fat mass, caused early reductions in food intake, and shifted macronutrient preference away from fat and toward carbohydrates. VSG had no impact on basal glucose but did improve the return to baseline after an oral glucose load. As has been shown previously, VSG increased postprandial insulin, GLP-1, and bile acids. There was no significant impact of VSG on plasma triglycerides, hepatic triglycerides, or cholesterol. Interestingly, the brown adipose tissue to white adipose tissue ratio tended to be greater in VSG compared to sham surgery animals. While VSG positively impacted several aspects of metabolism, it did not enhance maximal oxygen capacity and seemed to lower metabolic efficiency as indicated by lower resting oxygen consumption and fat and carbohydrate oxidation.
Conclusions: VSG can improve the metabolic status of animals with a low exercise capacity independently of exercise capacity.[Hide abstract]
|Phosphate-dependent FGF23 secretion is modulated by PiT2/Slc20a2Fibroblast growth factor 23 (FGF23) is a bone-derived hormone which regulates the serum inorganic phosphate (Pi) concentration and vitamin D metabolism. The presence of regulatory feedback loops operating between FGF23 and Pi/vitamin D has been suggested. However, no Pi-receptor or sensor has yet been identified in mammals. Bon and colleagues provide experimental evidence that the Pi transporter PiT2 regulates synthesis and secretion of FGF23 in response to high Pi load. These findings may identify PiT2 as a target for novel therapies to improve the excessive FGF23 secretion in hyperphosphatemic disorders such as chronic kidney disease.|
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Objective: The canonical role of the bone-derived fibroblast growth factor 23 (Fgf23) is to regulate the serum inorganic phosphate (Pi) level. As part of a feedback loop, serum Pi levels control Fgf23 secretion through undefined mechanisms. We recently showed in vitro that the two high-affinity Na+-Pi co-transporters PiT1/Slc20a1 and PiT2/Slc20a2 were required for mediating Pi-dependent signaling. Here, we addressed the contribution of PiT1 and PiT2 to the regulation of Fgf23 secretion.
Methods: To this aim, we used PiT2 KO and DMP1Cre; PiT1lox/lox fed Pi-modified diets, as well as ex vivo isolated long bone shafts. Fgf23 secretion and expression of Pi homeostasis-related genes were assessed.
Results: In vivo, PiT2 KO mice responded inappropriately to low-Pi diets, displaying abnormally normal serum levels of intact Fgf23. Despite the high iFgf23 level, serum Pi levels remained unaffected, an effect that may relate to lower αKlotho expression in the kidney. Moreover, consistent with a role of PiT2 as a possible endocrine Pi sensor, the iFGF23/cFGF23 ratios were suppressed in PiT2 KO mice, irrespective of the Pi loads. While deletion of PiT1 in osteocytes using the DMP1-Cre mice was inefficient, adenovirus-mediated deletion of PiT1 in isolated long bone shafts suggested that PiT1 does not contribute to Pi-dependent regulation of Fgf23 secretion. In contrast, using isolated bone shafts from PiT2 KO mice, we showed that PiT2 was necessary for the appropriate Pi-dependent secretion of Fgf23, independently from possible endocrine regulatory loops.
Conclusions: Our data provide initial mechanistic insights underlying the Pi-dependent regulation of Fgf23 secretion in identifying PiT2 as a potential player in this process, at least in high Pi conditions. Targeting PiT2, therefore, could improve excess FGF23 in hyperphosphatemic conditions such as chronic kidney disease.[Hide abstract]
|GPR142 Prompts GLP-1 Release to Improve β Cell FunctionGPR142 is a tryptophan-activated Gαq-coupled receptor with enriched expression in pancreatic islets. Both natural and synthetic ligands for this receptor were shown to enhance glucose-dependent insulin secretion and improve in vivo glucose homeostasis in animals. However, which specific cell types in the pancreatic islet express this receptor and what mechanisms mediate augmented insulin release remains poorly understood. Lin et al. show that in addition to its localization in β cells, GPR142 is measurably expressed in α cells in both human and mouse pancreas. They demonstrate that GPR142 activation stimulates glucagon secretion as well as glucagon-like peptide-1 production and release from the islet. The latter determines GPR142’s insulinotropic, β cell proliferative, and pro-survival effects.|
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Objective: GPR142 agonists are being pursued as novel diabetes therapies by virtue of their insulin secretagogue effects. But it is undetermined whether GPR142's functions in pancreatic islets are limited to regulating insulin secretion. The current study expands research on its action.
Methods and results: We demonstrated by in situ hybridization and immunostaining that GPR142 is expressed not only in β cells but also in a subset of α cells. Stimulation of GPR142 by a selective agonist increased glucagon secretion in both human and mouse islets. More importantly, the GPR142 agonist also potentiated glucagon-like peptide-1 (GLP-1) production and its release from islets through a mechanism that involves upregulation of prohormone convertase 1/3 expression. Strikingly, stimulation of insulin secretion and increase in insulin content via GPR142 engagement requires intact GLP-1 receptor signaling. Furthermore, GPR142 agonist increased β cell proliferation and protected both mouse and human islets against stress-induced apoptosis.
Conclusions: Collectively, we provide here evidence that local GLP-1 release from α cells defines GPR142's beneficial effects on improving β cell function and mass, and we propose that GPR142 agonism may have translatable and durable efficacy for the treatment of type 2 diabetes.[Hide abstract]
|Overexpression of ST5 has no effect on β-cell proliferation Identifying methods to promote β-cell proliferation, especially in adults, has been an attractive therapeutic strategy for the treatment of diabetes. Suppression of Tumorigenicity 5 (ST5) plays a role in one of the pathways thought to promote cell proliferation. Ou and colleagues tested the hypothesis that overexpression of the long isoform of ST5, an activator of the Ras/ERK pathway, is able to promote adult β-cell proliferation. However, they found that that overexpressing ST5 under both basal metabolic or challenged states is not sufficient to enhance β-cell proliferation.
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Objective: Both Type I and Type II diabetes mellitus result from insufficient functional β-cell mass. Efforts to increase β-cell proliferation as a means to restore β-cell mass have been met with limited success. Suppression of Tumorigenicity 5 (ST5) activates Ras/Erk signaling in the presence of Epidermal Growth Factor (EGF). In the pancreatic islet, Ras/Erk signaling is required for augmented β-cell proliferation during pregnancy, suggesting that ST5 is an appealing candidate to enhance adult β-cell proliferation. We aimed to test the hypothesis that overexpression of ST5 drives adult β-cell proliferation.
Methods: We utilized a doxycycline-inducible bitransgenic mouse model to activate β-cell-specific expression of human ST5 in adult mice at will. Islet morphology, β-cell proliferation, and β-cell mass in control and ST5-overexpressing (ST5 OE) animals were analyzed by immunofluorescent staining, under basal and two stimulated metabolic states: pregnancy and streptozotocin (STZ)-induced β-cell loss.
Results: Doxycycline treatment resulted in robust ST5 overexpression in islets from 12-16 week-old ST5 OE animals compared to controls, without affecting the islet morphology and identity of the β-cells. Under both basal and metabolically stimulated pregnancy states, β-cell proliferation and mass were comparable in ST5 OE and control animals. Furthermore, there was no detectable difference in β-cell proliferation between ST5 OE and control animals in response to STZ-induced β-cell loss.
Conclusions: We successfully derived an inducible bitransgenic mouse model to overexpress ST5 specifically in β-cells. However, our findings demonstrate that ST5 overexpression by itself has no mitogenic effect on the adult β-cell under basal and metabolically challenged states.[Hide abstract]