Probiotic culture supernatant improves metabolic function through FGF21-adiponectin pathway in mice

High-fat/high-fructose diet plus intermittent hypoxia exposure (HFDIH) causes metabolic disorders such as insulin resistance, obesity, nonalcoholic fatty liver disease (NAFLD) and type 2 diabetes. The purpose of this study is to examine the effects and understand the mechanism of action of Lactobacillus rhamnosus GG culture supernatant (LGGs) on HFDIH-induced metabolic dysfunction. Mice were fed high-fat:high-fructose diet for 15 weeks. After 3 weeks of feeding, the mice were exposed to chronic intermittent hypoxia for the next 12 weeks (HFDIH), and LGGs was supplemented over the entire experiment. HFDIH exposure significantly led to metabolic disorders. LGGs treatment showed significant improvements in indices of metabolic disorders including fat mass, energy expenditure, glucose intolerance, insulin resistance, increased hepatic steatosis and liver injury. HFDIH mice markedly increased adipose inflammation and adipocyte size, and reduced circulating adiponectin, which was restored by LGGs treatment. LGGs treatment increased hepatic FGF21 mRNA expression and circulating FGF21 protein levels, which were associated with increased hepatic PPARα expression and fecal butyrate concentration. In addition, HFDIH-induced hepatic fat accumulation and apoptosis were significantly reduced by LGGs supplementation. In summary, LGGs treatment increased energy expenditure and insulin sensitivity and prevented metabolic abnormalities in HFDIH mice, and this is associated with the FGF21-adiponectin signaling pathway. LGGs may be a potential prevention/treatment strategy in subjects with the metabolic syndrome.     

Aging is associated with increased FGF21 levels but unaltered FGF21 responsiveness in adipose tissue

Fibroblast growth factor 21 (FGF21) has been proposed to be an antiaging hormone on the basis of experimental studies in rodent models. However, circulating FGF21 levels are increased with aging in rodents and humans. Moreover, despite the metabolic health‐promoting effects of FGF21, the levels of this hormone are increased under conditions such as obesity and diabetes, an apparent incongruity that has been attributed to altered tissue responsiveness to FGF21. Here, we investigated serum FGF21 levels and expression of genes encoding components of the FGF21‐response molecular machinery in adipose tissue from healthy elderly individuals (≥70 years old) and young controls. Serum FGF21 levels were increased in elderly individuals and were positively correlated with insulinemia and HOMA‐IR, indices of mildly deteriorated glucose homeostasis. Levels of β‐Klotho, the coreceptor required for cellular responsiveness to FGF21, were increased in subcutaneous adipose tissue from elderly individuals relative to those from young controls, whereas FGF receptor‐1 levels were unaltered. Moreover, total ERK1/2 protein levels were decreased in elderly individuals in association with an increase in the ERK1/2 phosphorylation ratio relative to young controls. Adipose explants from aged and young mice respond similarly to FGF21 “ex vivo”. Thus, in contrast to what is observed in obesity and diabetes, high levels of FGF21 in healthy aging are not associated with repressed FGF21‐responsiveness machinery in adipose tissue. The lack of evidence for impaired FGF21 responsiveness in adipose tissue establishes a distinction between alterations in the FGF21 endocrine system in aging and chronic metabolic pathologies.     

Impact of apolipoprotein A1- or lecithin:cholesterol acyltransferase-deficiency on white adipose tissue metabolic activity and glucose homeostasis in mice

High density lipoprotein (HDL) has attracted the attention of biomedical community due to its well-documented role in atheroprotection. HDL has also been recently implicated in the regulation of islets of Langerhans secretory function and in the etiology of peripheral insulin sensitivity. Indeed, data from numerous studies strongly indicate that the functions of pancreatic β-cells, skeletal muscles and adipose tissue could benefit from improved HDL functionality. To better understand how changes in HDL structure may affect diet-induced obesity and type 2 diabetes we aimed at investigating the impact of Apoa1 or Lcat deficiency, two key proteins of peripheral HDL metabolic pathway, on these pathological conditions in mouse models. We report that universal deletion of apoa1 or lcat expression in mice fed western-type diet results in increased sensitivity to body-weight gain compared to control C57BL/6 group. These changes in mouse genome correlate with discrete effects on white adipose tissue (WAT) metabolic activation and plasma glucose homeostasis. Apoa1-deficiency results in reduced WAT mitochondrial non-shivering thermogenesis. Lcat-deficiency causes a concerted reduction in both WAT oxidative phosphorylation and non-shivering thermogenesis, rendering lcat^?/? mice the most sensitive to weight gain out of the three strains tested, followed by apoa1^?/? mice. Nevertheless, only apoa1^?/? mice show disturbed plasma glucose homeostasis due to dysfunctional glucose-stimulated insulin secretion in pancreatic β-islets and insulin resistant skeletal muscles. Our analyses show that both apoa1^?/? and lcat^?/? mice fed high-fat diet have no measurable Apoa1 levels in their plasma, suggesting no direct involvement of Apoa1 in the observed phenotypic differences among groups.     

Loss of DJ-1 promotes browning of white adipose tissue in diet-induced obese mice

The seminal discovery of browning of white adipose tissue (WAT) holds great promise for the treatment of obesity and metabolic syndrome. DJ-1 is evolutionarily conserved across species, and mutations in DJ-1 have been identified in Parkinson's disease. Higher levels of DJ-1 are associated with obesity, but the underlying mechanism is less understood. Here, we report the previously unappreciated role of DJ-1 in white adipocyte biology in mature models of obesity. We used DJ-1 knockout (KO) mouse models and wild-type littermates maintained on a normal diet or high-fat diet as well as in vitro cell models to show the direct effects of DJ-1 depletion on adipocyte phenotype, thermogenic capacity, fat metabolism, and microenvironment profile. Global DJ-1 KO mice show increased sympathetic input to WAT and β3-adrenergic receptor intracellular signaling, leading to a previously unrecognized compensatory mechanism through browning of WAT with associated characteristics, including high mitochondrial contents, reduced lipid accumulation, adequate vascularization and attenuated autophagy. DJ-1 KO mice had normal body weight, energy balance, and adiposity, which were associated with protective effects on healthy WAT expansion by hyperplasia. Our findings revealed that browning of inguinal WAT occurred in DJ-1 KO mice that do not show increased predisposition to obesity and suggest that such potential mechanism may overcome the adverse metabolic consequences of obesity independent of an effect on body weight. Here, we provide the first direct evidence that targeting DJ-1 in adipocyte metabolic health may offer a unique therapeutic strategy for the treatment of obesity.     

FGF21 mediates alcohol-induced adipose tissue lipolysis by activation of systemic release of catecholamine in mice

Alcohol consumption leads to adipose tissue lipoatrophy and mobilization of FFAs, which contributes to hepatic fat accumulation in alcoholic liver disease. This study aimed to investigate the role of fibroblast growth factor (FGF)21, a metabolic regulator, in the regulation of chronic-binge alcohol-induced adipose tissue lipolysis. FGF21 KO mice were subjected to chronic-binge alcohol exposure, and epididymal white adipose tissue lipolysis and liver steatosis were investigated. Alcohol exposure caused adipose intracellular cAMP elevation and activation of lipolytic enzymes, leading to FFA mobilization in both WT and FGF21 KO mice. However, alcohol-induced systemic elevation of catecholamine, which is known to be a major player in adipose lipolysis by binding to the β-adrenergic receptor, was markedly inhibited in KO mice. Supplementation with recombinant human FGF21 to alcohol-exposed FGF21 KO mice resulted in an increase in fat loss in parallel with an increase of circulating norepinephrine concentration. Furthermore, alcohol consumption-induced fatty liver was blunted in the KO mice, indicating an inhibition of fatty acid reverse transport from adipose to the liver in the KO mice. Taken together, our studies demonstrate that FGF21 KO mice are protected from alcohol-induced adipose tissue excess-lipolysis through a mechanism involving systemic catecholamine release.     

Leptin selectively reduces white adipose tissue in mice via a UCP1-dependent mechanism in brown adipose tissue

We tested the hypothesis that leptin, in addition to reducing body fat by restraining food intake, reduces body fat through a peripheral mechanism requiring uncoupling protein 1 (UCP1). Leptin was administered to wild-type (WT) mice and mice with a targeted disruption of the UCP1 gene (UCP1 deficient), while vehicle-injected control animals of each genotype were pair-fed to each leptin-treated group. Leptin reduced the size of white adipose tissue (WAT) depots in WT mice but not in UCP1-deficient animals. This was accompanied by a threefold increase in the amount of UCP1 protein and mRNA in the brown adipose tissue (BAT) of WT mice. Leptin also increased UCP2 mRNA in WAT of both WT and UCP1-deficient mice but increased UCP2 and UCP3 mRNA only in BAT from UCP1-deficient mice. These results indicate that leptin reduces WAT through a peripheral mechanism requiring the presence of UCP1, with little or no involvement of UCP2 or UCP3.     

Central leptin regulates the UCP1 and ob genes in brown and white adipose tissue via different beta-adrenoceptor subtypes

The three known subtypes of beta-adrenoreceptors (beta(1)-AR, beta(2)-AR, and beta(3)-AR) are differentially expressed in brown and white adipose tissue and mediate peripheral responses to central modulation of sympathetic outflow by leptin. To assess the relative roles of the beta-AR subtypes in mediating leptin's effects on adipocyte gene expression, mice with a targeted disruption of the beta(3)-adrenoreceptor gene (beta(3)-AR KO) were treated with vehicle or the beta(1)/beta(2)-AR selective antagonist, propranolol (20 microgram/g body weight/day) prior to intracerebroventricular (ICV) injections of leptin (0.1 microgram/g body weight/day). Leptin produced a 3-fold increase in UCP1 mRNA in brown adipose tissue of wild type (FVB/NJ) and beta(3)-AR KO mice. The response was unaltered by propranolol in wild type mice, but was completely blocked by this antagonist in beta(3)-AR KO mice. In contrast, ICV leptin had no effect on leptin mRNA in either epididymal or retroperitoneal white adipose tissue (WAT) from beta(3)-AR KOs. Moreover, propranolol did not block the ability of exogenous leptin to reduce leptin mRNA in either WAT depot site of wild type mice. These results demonstrate that the beta(3)-AR is required for leptin-mediated regulation of ob mRNA expression in WAT, but is interchangeable with the beta(1)/beta(2)-ARs in mediating leptin's effect on UCP1 mRNA expression in brown adipose tissue.     

Deficiency of heat shock protein A12A promotes browning of white adipose tissues in mice

Browning of white adipose tissues (WAT) is critical for a variety of physiological and pathophysiological events. Given the limited understanding in molecular control of WAT browning, further research is needed. Heat shock protein A12A (HSPA12A) is a new member of multigene Hsp70 family. This study investigated the effect of HSPA12A on the browning of WAT. WAT Browning in mice was induced by cold exposure for 5 days. We observed that nuclear HSPA12A content was increased in WAT after cold exposure, while deficiency of HSPA12A (Hspa12a^−/−) promoted the cold-induced browning of WAT in mice compared to wild type (WT) littermates. Accordingly, Hspa12a^−/− mice showed attenuation of body temperature drop and increase of thermogenic gene expression compared to WT mice after cold exposure. However, in vitro experiments demonstrated that HSPA12A deficiency in primary white adipocytes did not affect their browning and thermogenic gene expression. Further loss- and gain-of-HSPA12A functional studies revealed that HSPA12A deficiency promoted whereas HSPA12A overexpression impeded M2 macrophage polarization. Importantly, the conditioned medium (CM) from Hspa12a^−/− bone marrow-derived macrophages (BMDMs) enhanced the browning of primary white adipocytes when compared to the CM from WT BMDMs. The data identified macrophage HSPA12A as a novel regulator of WAT browning through a paracrine mechanism. Targeting HSPA12A might provide meaningful advances for the management of browning-associated physiological events such as hypothermia adaptation and pathophysiological disorders such as obesity and cancer-related cachexia.     

SCD1 promotes lipid mobilization in subcutaneous white adipose tissue

Beiging of white adipose tissue (WAT) has beneficial effects on metabolism. Although it is known that beige adipocytes are active in lipid catabolism and thermogenesis, how they are regulated deserves more explorations. In this study, we demonstrate that stearoyl-CoA desaturase 1 (SCD1) in subcutaneous WAT (scWAT) responded to cold stimulation and was able to promote mobilization of triacylglycerol [TAG (triglyceride)]. In vitro studies showed that SCD1 promoted lipolysis in C3H10T1/2 white adipocytes. The lipolytic effect was contributed by one of SCD1’s products, oleic acid (OA). OA upregulated adipose TAG lipase and hormone-sensitive lipase expression. When SCD1 was overexpressed in the scWAT of mice, lipolysis was enhanced, and oxygen consumption and heat generation were increased. These effects were also demonstrated by the SCD1 knockdown experiments in mice. In conclusion, our study suggests that SCD1, known as an enzyme for lipid synthesis, plays a role in upregulating lipid mobilization through its desaturation product, OA.     

EPO对高脂喂养小鼠棕色脂肪组织PRDM16、FGF21表达及STAT3磷酸化水平的影响

目的：探索促红细胞生成素（EPO）对高脂饲料（HFD）喂养小鼠血糖和血浆胰岛素水平、胰岛素抵抗指数（HOMA-IR）、糖耐量,以及棕色脂肪组织中含PR结构域蛋白16（PRDM16）、信号转导与转录激活因子3（STAT3）磷酸化水平（p-STAT3/STAT3）、成纤维细胞生长因子21（FGF21）mRNA以及蛋白质表达的影响,为肥胖及其并发症的发生机制提供线索。方法：20只高脂饲料喂养的C57BL/6J雄性小鼠随机分为对照组（HFD-Con）和EPO组（HFD-EPO）,两组分别腹腔注射生理盐水和EPO（200 IU/kg）,每周3次,连续4周。4周后检测两组动物的体重、血糖与血浆胰岛素水平、HOMA-IR及糖耐量的变化;分别使用实时定量PCR法和Western blot法检测棕色脂肪组织中PRDM16、STAT3、FGF21 mRNA和蛋白质水平。结果：腹腔注射EPO 4周后,HFD-EPO组小鼠体重为（26.65±0.85）g,HFD-Con组体重为（31.50±1.60）g,P<0.01。HFD-Con组血糖为（91.06±9.86）mg/dl,HFD-EPO组为（62.79±8.09）mg/dl,P<0.01;HFD-EPO组小鼠血浆胰岛素水平为（10.56±1.06）μU/ml,HFD-Con组为（13.2±1.1）μU/ml,P<0.01。与HFD-Con组比较,HFD-EPO组的糖耐量水平显著改善,胰岛素抵抗指数下降;HFD-EPO组动物棕色脂肪组织中PRDM16、FGF21mRNA以及蛋白质表达,p-STAT/STAT3水平均显著增加,两组小鼠肝脏中FGF21 mRNA含量、血浆中FGF21含量无明显差异。结论：EPO可能通过增加棕色脂肪组织中PRDM16表达促进棕色脂肪组织的分化,降低高脂喂养小鼠的血糖水平、改善高脂喂养小鼠的糖代谢状态。     

The P2X7 ion channel is dispensable for energy and metabolic homeostasis of white and brown adipose tissues

Purinergic Signalling - Several studies suggest a role of extracellular adenine nucleotides in regulating adipose tissue functions via the purinergic signaling network. Metabolic studies in mice...     

Restoration of leptin responsiveness in diet‐induced obese mice using an optimized leptin analog in combination with exendin‐4 or FGF21

The identification of leptin as a mediator of body weight regulation provided much initial excitement for the treatment of obesity. Unfortunately, leptin monotherapy is insufficient in reversing obesity in rodents or humans. Recent findings suggest that amylin is able to restore leptin sensitivity and when used in combination with leptin enhances body weight loss in obese rodents and humans. However, as the uniqueness of this combination therapy remains unclear, we assessed whether co‐administration of leptin with other weight loss‐inducing hormones equally restores leptin responsiveness in diet‐induced obese (DIO) mice. Accordingly, we report here the design and characterization of a series of site‐specifically enhanced leptin analogs of high potency and sustained action that, when administered in combination with exendin‐4 or fibroblast growth factor 21 (FGF21), restores leptin responsiveness in DIO mice after an initial body weight loss of 30%. Using either combination, body weight loss was enhanced compared with either exendin‐4 or FGF21 monotherapy, and leptin alone was sufficient to maintain the reduced body weight. In contrast, leptin monotherapy proved ineffective when identical weight loss was induced by caloric restriction alone over a comparable time. Accordingly, we find that a hypothalamic counter‐regulatory response to weight loss, assessed using changes in hypothalamic agouti related peptide (AgRP) levels, is triggered by caloric restriction, but blunted by treatment with exendin‐4. We conclude that leptin re‐sensitization requires pharmacotherapy but does not appear to be restricted to a unique signaling pathway. Our findings provide preclinical evidence that high activity, long‐acting leptin analogs are additively efficacious when used in combination with other weight‐lowering agents. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd.     

Intracerebroventricular leptin infusion improves glucose homeostasis in lean type 2 diabetic MKR mice via hepatic vagal and non-vagal mechanisms

MKR mice, lacking insulin-like growth factor 1 receptor (IGF-1R) signaling in skeletal muscle, are lean yet hyperlipidemic, hyperinsulinemic, and hyperglycemic, with severe insulin resistance and elevated hepatic and skeletal muscle levels of triglycerides. We have previously shown that chronic peripheral administration of the adipokine leptin improves hepatic insulin sensitivity in these mice independently of its effects on food intake. As central leptin signaling has been implicated in the control of peripheral glucose homeostasis, here we examined the ability of central intracerebroventricular leptin administration to affect energy balance and peripheral glucose homeostasis in non-obese diabetic male MKR mice. Central leptin significantly reduced food intake, body weight gain and adiposity, as well as serum glucose, insulin, leptin, free fatty acid and triglyceride levels relative to ACSF treated controls. These reductions were accompanied by increased fat oxidation as measured by indirect calorimetry, as well as increased oxygen consumption. Central leptin also improved glucose tolerance and hepatic insulin sensitivity determined using the euglycemic-hyperinsulinemic clamps relative to pair fed vehicle treated controls, as well as increasing the rate of glucose disappearance. Hepatic vagotomy only partially reversed the ability of central leptin to improve glucose tolerance. These results demonstrate that central leptin dramatically improves insulin sensitivity independently of its effects on food intake, in a lean mouse model of type 2 diabetes. The findings also suggest that: 1) both hepatic vagal and non-vagal pathways contribute to this improvement, and 2) central leptin alters glucose disposal in skeletal muscle in this model.     

Targeting FGF21 in cardiovascular and metabolic diseases: from mechanism to medicine

Cardiovascular and metabolic disease (CVMD) is becoming increasingly prevalent in developed and developing countries with high morbidity and mortality. In recent years, fibroblast growth factor 21 (FGF21) has attracted intensive research interest due to its purported role as a potential biomarker and critical player in CVMDs, including atherosclerosis, coronary artery disease, myocardial infarction, hypoxia/reoxygenation injury, heart failure, type 2 diabetes, obesity, and nonalcoholic steatohepatitis. This review summarizes the recent developments in investigating the role of FGF21 in CVMDs and explores the mechanism whereby FGF21 regulates the development of CVMDs. Novel molecular targets and related pathways of FGF21 (adenosine 5''-monophosphate-activated protein kinase, silent information regulator 1, autophagy-related molecules, and gut microbiota-related molecules) are highlighted in this review. Considering the poor pharmacokinetics and biophysical properties of native FGF21, the development of new generations of FGF21-based drugs has tremendous therapeutic potential. Related preclinical and clinical studies are also summarized in this review to foster clinical translation. Thus, our review provides a timely and insightful overview of the physiology, biomarker potential, molecular targets, and therapeutic potential of FGF21 in CVMDs.