Preventing High Fat Diet-induced Obesity and Improving Insulin Sensitivity through Neuregulin 4 Gene Transfer.
ABSTRACT: Neuregulin 4 (NRG4), an epidermal growth factor-like signaling molecule, plays an important role in cell-to-cell communication during tissue development. Its function to regulate energy metabolism has recently been reported. This current study was designed to assess the preventive and therapeutic effects of NRG4 overexpression on high fat diet (HFD)-induced obesity. Using the hydrodynamic gene transfer method, we demonstrate that Nrg4 gene transfer in mice suppressed the development of diet-induced obesity, but did not affect pre-existing adiposity and body weight in obese mice. Nrg4 gene transfer curbed HFD-induced hepatic steatosis by inhibiting lipogenesis and PPARγ-mediated lipid storage. Concurrently, overexpression of NRG4 reduced chronic inflammation in both preventive and treatment studies, evidenced by lower mRNA levels of macrophage marker genes including F4/80, Cd68, Cd11b, Cd11c, and macrophage chemokine Mcp1, resulting in improved insulin sensitivity. Collectively, these results demonstrate that overexpression of the Nrg4 gene by hydrodynamic gene delivery prevents HFD-induced weight gain and fatty liver, alleviates obesity-induced chronic inflammation and insulin resistance, and supports the health benefits of NRG4 in managing obesity and obesity-associated metabolic disorders.
Project description:High-fat diet (HFD) induced obesity is associated with low-grade inflammation, insulin resistance (IR), and glucose intolerance. The objective of this study is to assess the effect of interleukin 10 (IL10), an anti-inflammatory cytokine, on blocking HFD-induced obesity and obesity-associated metabolic disorders by hydrodynamic delivery of IL10-containing plasmid. Animals fed a regular chow or HFD received two injections (one on day 1 and the other on day 31) of plasmids containing green fluorescence protein (GFP) or mouse IL10 (mIL10) gene. Blood concentration of mIL10 reached ~200?ng/ml on day 7 in animals receiving mIL10 plasmid DNA. The transfection efficiency of liver cells was the same in animals fed a regular chow or HFD. No difference was seen in animals on regular chow when injected with plasmids containing either gfp or mIL10 gene. Overexpression of mIL10 prevented weight gain of animals on HFD. Intraperitoneal glucose tolerance test (IPGTT) and insulin tolerance tests (ITT) showed that mIL10 maintained insulin sensitivity and prevented glucose intolerance. The mechanistic study reveals that mIL10 suppressed macrophage infiltration and reduced the development of crown-like structures in adipose tissue (AT). Collectively, these results suggest that maintaining a higher level of IL10 through gene transfer could be an effective strategy in preventing diet-induced obesity.
Project description:BACKGROUND:The growth differentiation factor 11 (GDF11) was shown to reverse age-related hypertrophy on cardiomyocytes and considered as anti-aging rejuvenation factor. The role of GDF11 in regulating metabolic homeostasis is unclear. In this study, we investigated the functions of GDF11 in regulating metabolic homeostasis and energy balance. METHODS:Using a hydrodynamic injection approach, plasmids carrying a mouse Gdf11 gene were delivered into mice and generated the sustained Gdf11 expression in the liver and its protein level in the blood. High fat diet (HFD)-induced obesity was employed to examine the impacts of Gdf11 gene transfer on HFD-induced adiposity, hyperglycemia, insulin resistance, and hepatic lipid accumulation. The impacts of GDF11 on metabolic homeostasis of obese and diabetic mice were examined using HFD-induced obese and STZ-induced diabetic models. RESULTS:Gdf11 gene transfer alleviates HFD-induced obesity, hyperglycemia, insulin resistance, and fatty liver development. In obese and STZ-induced diabetic mice, Gdf11 gene transfer restores glucose metabolism and improves insulin resistance. Mechanism study reveals that Gdf11 gene transfer increases the energy expenditure of mice, upregulates the expression of genes responsible for thermoregulation in brown adipose tissue, downregulates the expression of inflammatory genes in white adipose tissue and those involved in hepatic lipid and glucose metabolism. Overexpression of GDF11 also activates TGF-?/Smad2, PI3K/AKT/FoxO1, and AMPK signaling pathways in white adipose tissue. CONCLUSIONS:These results demonstrate that GDF11 plays an important role in regulating metabolic homeostasis and energy balance and could be a target for pharmacological intervention to treat metabolic disease.
Project description:IMPACT STATEMENT:Due to high-fat and high-sugar diets accompanied by sedentary lifestyles, diabetes has become a global epidemic. Literature findings suggest a potential therapeutic effect of Nrg4 on treating obesity-related metabolic disorders including type 2 diabetes (T2D). Adipose tissue-derived MSCs (ADSCs) were used in our study as they are abundant and can be harvested with minimally invasive procedures. In the end, our study reveals that ADSC transplantation improves glucose tolerance and metabolic balance in HFD-fed mice by multiple mechanisms, including upregulating GLUT4 expression and suppressing inflammation. More importantly, our study shows that Nrg4 overexpression could improve the efficacy of ADSCs in ameliorating insulin resistance (IR) and other obesity-related metabolic disorders, given the function of Nrg4 in attenuating hepatic lipogenesis. It would provide a new therapeutic strategy for the treatment of obesity, IR, and T2D.
Project description:Brown and white adipose tissue exerts pleiotropic effects on systemic energy metabolism in part by releasing endocrine factors. Neuregulin 4 (Nrg4) was recently identified as a brown fat-enriched secreted factor that ameliorates diet-induced metabolic disorders, including insulin resistance and hepatic steatosis. However, the physiological mechanisms through which Nrg4 regulates energy balance and glucose and lipid metabolism remain incompletely understood. The aims of the current study were: i) to investigate the regulation of adipose Nrg4 expression during obesity and the physiological signals involved, ii) to elucidate the mechanisms underlying Nrg4 regulation of energy balance and glucose and lipid metabolism, and iii) to explore whether Nrg4 regulates adipose tissue secretome gene expression and adipokine secretion.We examined the correlation of adipose Nrg4 expression with obesity in a cohort of diet-induced obese mice and investigated the upstream signals that regulate Nrg4 expression. We performed metabolic cage and hyperinsulinemic-euglycemic clamp studies in Nrg4 transgenic mice to dissect the metabolic pathways regulated by Nrg4. We investigated how Nrg4 regulates hepatic lipid metabolism in the fasting state and explored the effects of Nrg4 on adipose tissue gene expression, particularly those encoding secreted factors.Adipose Nrg4 expression is inversely correlated with adiposity and regulated by pro-inflammatory and anti-inflammatory signaling. Transgenic expression of Nrg4 increases energy expenditure and augments whole body glucose metabolism. Nrg4 protects mice from diet-induced hepatic steatosis in part through activation of hepatic fatty acid oxidation and ketogenesis. Finally, Nrg4 promotes a healthy adipokine profile during obesity.Nrg4 exerts pleiotropic beneficial effects on energy balance and glucose and lipid metabolism to ameliorate obesity-associated metabolic disorders. Biologic therapeutics based on Nrg4 may improve both type 2 diabetes and non-alcoholic fatty liver disease (NAFLD) in patients.
Project description:Adipose tissue macrophages have been proposed as a link between obesity and insulin resistance. However, the mechanisms underlying these processes are not completely defined. Calpains are calcium-dependent neutral cysteine proteases that modulate cellular function and have been implicated in various inflammatory diseases. To define whether activated calpains influence diet-induced obesity and adipose tissue macrophage accumulation, mice that were either wild type (WT) or overexpressing calpastatin (CAST Tg), the endogenous inhibitor of calpains were fed with high (60% kcal) fat diet for 16 weeks. CAST overexpression did not influence high fat diet-induced body weight and fat mass gain throughout the study. Calpain inhibition showed a transient improvement in glucose tolerance at 5 weeks of HFD whereas it lost this effect on glucose and insulin tolerance at 16 weeks HFD in obese mice. However, CAST overexpression significantly reduced adipocyte apoptosis, adipose tissue collagen and macrophage accumulation as detected by TUNEL, Picro Sirius and F4/80 immunostaining, respectively. CAST overexpression significantly attenuated obesity-induced inflammatory responses in adipose tissue. Furthermore, calpain inhibition suppressed macrophage migration to adipose tissue in vitro. The present study demonstrates a pivotal role for calpains in mediating HFD-induced adipose tissue remodeling by influencing multiple functions including apoptosis, fibrosis and inflammation.
Project description:The objective of this study was to assess the activity of anti-inflammatory cytokine IL-13 (interleukin-13) in blocking high-fat diet-induced obesity and obesity-associated insulin resistance and liver steatosis.C57BL/6 mice were fed a high-fat diet and received hydrodynamic delivery of plasmids carrying the mouse Il-13 or Gfp (control) gene. IL-13 blood protein levels, food consumption and body weight of mice were continuously monitored for 8 weeks. Fat and lean masses of treated and control animals were determined at the end of the experiment. Serum concentrations of glucose, insulin and lipids were determined, and mRNA levels of macrophage marker genes in adipose tissue and genes involved in energy metabolism were examined using real-time PCR. Glucose tolerance and insulin sensitivity tests were performed to determine glucose homeostasis. Histochemistry and lipid assays were performed to determine the hepatic lipid accumulation.Blood concentration of IL-13 was 20?ng?ml(-1) 1 week after gene delivery and declined with time. Overexpression of Il-13 prevented high-fat diet-induced weight gain without affecting food consumption. Mice that underwent Il-13 gene transfer showed regular body weight and normal serum concentrations of glucose and insulin, and less lipid accumulation in the liver. Overexpression of Il-13 blocked macrophage infiltration in adipose tissue and suppressed high-fat diet-induced expression of inflammatory F4/80, Cd68 and Mcp1, and elevated the expression of Ucp1 (uncoupling protein 1 gene) responsible for energy expenditure.These results suggest that suppression of diet-induced inflammation by IL-13 is an effective strategy in preventing diet-induced obesity and obesity-associated insulin resistance and fatty liver.
Project description:C-type natriuretic peptide (CNP) is expressed in diverse tissues, including adipose and endothelium, and exerts its effects by binding to and activating its receptor, guanylyl cyclase B. Natriuretic peptides regulate intracellular cGMP and phosphorylated vasodilator-stimulated phosphoprotein (VASP). We recently revealed that overexpression of CNP in endothelial cells protects against high-fat diet (HFD)-induced obesity in mice. Given that endothelial CNP affects adipose tissue during obesity, CNP in adipocytes might directly regulate adipocyte function during obesity. Therefore, to elucidate the effect of CNP in adipocytes, we assessed 3T3-L1 adipocytes and transgenic (Tg) mice that overexpressed CNP specifically in adipocytes (A-CNP). We found that CNP activates the cGMP-VASP pathway in 3T3-L1 adipocytes. Compared with Wt mice, A-CNP Tg mice showed decreases in fat weight and adipocyte hypertrophy and increases in fatty acid ?-oxidation, lipolysis-related gene expression, and energy expenditure during HFD-induced obesity. These effects led to decreased levels of the macrophage marker F4/80 in the mesenteric fat pad and reduced inflammation. Furthermore, A-CNP Tg mice showed improved glucose tolerance and insulin sensitivity, which were associated with enhanced insulin-stimulated Akt phosphorylation. Our results suggest that CNP overexpression in adipocytes protects against adipocyte hypertrophy, excess lipid metabolism, inflammation, and decreased insulin sensitivity during HFD-induced obesity.
Project description:The proinflammatory consequences of obesity are thought to be due, in part, to macrophage infiltration into adipose tissue. There are, however, potential antiinflammatory consequences of obesity that include obesity-associated up-regulation of IL-1 receptor antagonist (IL-1RA). Here we show that obesity-associated up-regulation of IL-1RA speeds recovery from hypoxia. We found that high-fat diet-fed (HFD) mice recovered from acute hypoxia 5 times faster than normal-diet-fed (ND) mice. HFD mice had a 10-fold increase in serum IL-1RA when compared with ND mice. White adipose tissue (WAT) was a significant source of IL-RA, generating 330 +/- 77 pg/mg protein in HFD mice as compared with 15 +/- 5 pg/mg protein in ND mice. Peritoneal macrophages isolated from HFD mice showed little difference in IL-1RA production when compared with ND mice, but WAT macrophages from HFD mice generated 11-fold more IL-1RA than those from ND mice. When ND mice were given an ip transfer of the stromal vascular fraction portion of WAT from HFD mice, serum IL-1RA increased 836% and recovery from acute hypoxia was faster than in mice that did not receive a stromal vascular fraction transfer. To determine whether IL-1RA was important to this accelerated recovery, ND mice were administered exogenous IL-1RA prior to hypoxia, and their recovery matched that of HFD mice. Inversely, when IL-1RA was immunoabsorbed in HFD mice with IL-1RA antiserum, recovery from acute hypoxia was attenuated. Taken together these data demonstrate that HFD-induced obesity speeds recovery from hypoxia due to obesity-associated up-regulation of IL-1RA.
Project description:Activation of brown adipose tissue-mediated thermogenesis is a strategy for tackling obesity and promoting metabolic health. BMP8b is secreted by brown/beige adipocytes and enhances energy dissipation. Here we show that adipocyte-secreted BMP8b contributes to adrenergic-induced remodeling of the neuro-vascular network in adipose tissue (AT). Overexpression of bmp8b in AT enhances browning of the subcutaneous depot and maximal thermogenic capacity. Moreover, BMP8b-induced browning, increased sympathetic innervation and vascularization of AT were maintained at 28 °C, a condition of low adrenergic output. This reinforces the local trophic effect of BMP8b. Innervation and vascular remodeling effects required BMP8b signaling through the adipocytes to 1) secrete neuregulin-4 (NRG4), which promotes sympathetic axon growth and branching in vitro, and 2) induce a pro-angiogenic transcriptional and secretory profile that promotes vascular sprouting. Thus, BMP8b and NRG4 can be considered as interconnected regulators of neuro-vascular remodeling in AT and are potential therapeutic targets in obesity.
Project description:Obesity induces accumulation of adipose tissue macrophages (ATMs) and ATM-driven inflammatory responses that promote the development of glucose and lipid metabolism disorders. ClC-3 chloride channel/antiporter, encoded by the Clcn3, is critical for some basic cellular functions. Our previous work has shown significant alleviation of type 2 diabetes in Clcn3 knockout (Clcn3-/-) mice. In the present study we investigated the role of Clcn3 in high-fat diet (HFD)-induced obesity and ATM inflammation. To establish the mouse obesity model, both Clcn3-/- mice and wild-type mice were fed a HFD for 4 or 16 weeks. The metabolic parameters were assessed and the abdominal total adipose tissue was scanned using computed tomography. Their epididymal fat pad tissue and adipose tissue stromal vascular fraction (SVF) cells were isolated for analyses. We found that the HFD-fed Clcn3-/- mice displayed a significant decrease in obesity-induced body weight gain and abdominal visceral fat accumulation as well as an improvement of glucose and lipid metabolism as compared with HFD-fed wild-type mice. Furthermore, the Clcn3 deficiency significantly attenuated HFD-induced ATM accumulation, HFD-increased F4/80+ CD11c+ CD206- SVF cells as well as HFD-activated TLR-4/NF-?B signaling in epididymal fat tissue. In cultured human THP-1 macrophages, adenovirus-mediated transfer of Clcn3 specific shRNA inhibited, whereas adenovirus-mediated cDNA overexpression of Clcn3 enhanced lipopolysaccharide-induced activation of NF-?B and TLR-4. These results demonstrate a novel role for Clcn3 in HFD-induced obesity and ATM inflammation.