ABSTRACT
Glucagon signaling is essential for maintaining normoglycemia in mammals. The arrestin fold superfamily of proteins controls the trafficking, turnover, and signaling of transmembrane receptors as well as other intracellular signaling functions. Further investigation is needed to understand the in vivo functions of the arrestin domain-containing 4 (ARRDC4) protein family member and whether it is involved in mammalian glucose metabolism. Here, we show that mice with a global deletion of the ARRDC4 protein have impaired glucagon responses and gluconeogenesis at a systemic and molecular level. Mice lacking ARRDC4 exhibited lower glucose levels after fasting and could not suppress gluconeogenesis at the refed state. We also show that ARRDC4 coimmunoprecipitates with the glucagon receptor, and ARRDC4 expression is suppressed by insulin. These results define ARRDC4 as a critical regulator of glucagon signaling and glucose homeostasis and reveal a novel intersection of insulin and glucagon pathways in the liver.
Subject(s)
Glucagon , Insulin , Intracellular Signaling Peptides and Proteins , Liver , Animals , Mice , Glucagon/metabolism , Gluconeogenesis , Glucose/metabolism , Insulin/metabolism , Liver/metabolism , Mice, Inbred C57BL , Intracellular Signaling Peptides and Proteins/metabolismABSTRACT
Hepatic steatosis associated with high-fat diet, obesity, and type 2 diabetes is thought to be the major driver of severe liver inflammation, fibrosis, and cirrhosis. Cytosolic acetyl CoA (AcCoA), a central metabolite and substrate for de novo lipogenesis (DNL), is produced from citrate by ATP-citrate lyase (ACLY) and from acetate through AcCoA synthase short chain family member 2 (ACSS2). However, the relative contributions of these two enzymes to hepatic AcCoA pools and DNL rates in response to high-fat feeding are unknown. We report here that hepatocyte-selective depletion of either ACSS2 or ACLY caused similar 50% decreases in liver AcCoA levels in obese mice, showing that both pathways contribute to the generation of this DNL substrate. Unexpectedly however, the hepatocyte ACLY depletion in obese mice paradoxically increased total DNL flux measured by D2O incorporation into palmitate, whereas in contrast, ACSS2 depletion had no effect. The increase in liver DNL upon ACLY depletion was associated with increased expression of nuclear sterol regulatory element-binding protein 1c and of its target DNL enzymes. This upregulated DNL enzyme expression explains the increased rate of palmitate synthesis in ACLY-depleted livers. Furthermore, this increased flux through DNL may also contribute to the observed depletion of AcCoA levels because of its increased conversion to malonyl CoA and palmitate. Together, these data indicate that in fat diet-fed obese mice, hepatic DNL is not limited by its immediate substrates AcCoA or malonyl CoA but rather by activities of DNL enzymes.
Subject(s)
Diabetes Mellitus, Type 2 , Lipogenesis , Liver , Sterol Regulatory Element Binding Protein 1 , Animals , Mice , Acetyl Coenzyme A/metabolism , Adenosine Triphosphate/metabolism , ATP Citrate (pro-S)-Lyase/genetics , ATP Citrate (pro-S)-Lyase/metabolism , Diabetes Mellitus, Type 2/metabolism , Hepatocytes/metabolism , Liver/metabolism , Malonyl Coenzyme A/metabolism , Mice, Obese , Palmitates/metabolism , Sterol Regulatory Element Binding Protein 1/genetics , Sterol Regulatory Element Binding Protein 1/metabolismABSTRACT
RNA-guided, engineered nucleases derived from the prokaryotic adaptive immune system CRISPR-Cas represent a powerful platform for gene deletion and editing. When used as a therapeutic approach, direct delivery of Cas9 protein and single-guide RNA (sgRNA) could circumvent the safety issues associated with plasmid delivery and therefore represents an attractive tool for precision genome engineering. Gene deletion or editing in adipose tissue to enhance its energy expenditure, fatty acid oxidation, and secretion of bioactive factors through a "browning" process presents a potential therapeutic strategy to alleviate metabolic disease. Here, we developed "CRISPR-delivery particles," denoted CriPs, composed of nano-size complexes of Cas9 protein and sgRNA that are coated with an amphipathic peptide called Endo-Porter that mediates entry into cells. Efficient CRISPR-Cas9-mediated gene deletion of ectopically expressed GFP by CriPs was achieved in multiple cell types, including a macrophage cell line, primary macrophages, and primary pre-adipocytes. Significant GFP loss was also observed in peritoneal exudate cells with minimum systemic toxicity in GFP-expressing mice following intraperitoneal injection of CriPs containing Gfp-targeting sgRNA. Furthermore, disruption of a nuclear co-repressor of catabolism, the Nrip1 gene, in white adipocytes by CriPs enhanced adipocyte browning with a marked increase of uncoupling protein 1 (UCP1) expression. Of note, the CriP-mediated Nrip1 deletion did not produce detectable off-target effects. We conclude that CriPs offer an effective Cas9 and sgRNA delivery system for ablating targeted gene products in cultured cells and in vivo, providing a potential therapeutic strategy for metabolic disease.
Subject(s)
Adipose Tissue, White/metabolism , Energy Metabolism , Gene Targeting/methods , Nuclear Receptor Interacting Protein 1/genetics , Adipocytes/metabolism , Adipose Tissue, White/cytology , Animals , CRISPR-Cas Systems , Cell Line , Clustered Regularly Interspaced Short Palindromic Repeats , Gene Editing , Genes, Reporter , Humans , Mice, Inbred C57BL , Nuclear Receptor Interacting Protein 1/metabolism , Plasmids/genetics , Plasmids/metabolism , Uncoupling Protein 1/genetics , Uncoupling Protein 1/metabolismABSTRACT
Stearoyl-CoA desaturase 1 (SCD1), a lipogenic enzyme that adds a double bond at the delta 9 position of stearate (C18: 0) and palmitate (C16: 0), has been proven to be important in the development of obesity. Mice with skin-specific deficiency of SCD1 (SKO) display increased whole-body energy expenditure, which is protective against adiposity from a high-fat diet because it improves glucose clearance, insulin sensitivity, and hepatic steatosis. Of note, these mice also display elevated levels of the "pro-inflammatory" plasma interleukin-6 (IL-6). In whole skin of SKO mice, IL-6 mRNA levels are increased, and protein expression is evident in hair follicle cells and in keratinocytes. Recently, the well-known role of IL-6 in causing white adipose tissue lipolysis has been linked to indirectly activating the gluconeogenic enzyme pyruvate carboxylase 1 in the liver, thereby increasing hepatic glucose production. In this study, we suggest that skin-derived IL-6 leads to white adipose tissue lipolysis, which contributes to the lean phenotype of SKO mice without the incidence of meta-inflammation that is associated with IL-6 signaling.
Subject(s)
Interleukin-6/metabolism , Skin/metabolism , Stearoyl-CoA Desaturase/deficiency , Adipose Tissue, White/metabolism , Adiposity , Animals , Diet, High-Fat/adverse effects , Fatty Acids/metabolism , Gluconeogenesis , Hair Follicle/cytology , Hair Follicle/metabolism , Interleukin-6/genetics , Keratinocytes/metabolism , Lipolysis , Liver/metabolism , Macrophages/cytology , Male , Mice , Mice, Knockout , RNA, Messenger/genetics , RNA, Messenger/metabolism , Skin/cytology , Stearoyl-CoA Desaturase/genetics , Thinness/genetics , Thinness/metabolism , Tissue DistributionABSTRACT
Obesity-mediated inflammation is a major cause of insulin resistance, and macrophages play an important role in this process. The 78-kDa glucose-regulated protein (GRP78) is a major endoplasmic reticulum chaperone that modulates unfolded protein response (UPR), and mice with GRP78 heterozygosity were resistant to diet-induced obesity. Here, we show that mice with macrophage-selective ablation of GRP78 (Lyz- GRP78-/-) are protected from skeletal muscle insulin resistance without changes in obesity compared with wild-type mice after 9 wk of high-fat diet. GRP78-deficient macrophages demonstrated adapted UPR with up-regulation of activating transcription factor (ATF)-4 and M2-polarization markers. Diet-induced adipose tissue inflammation was reduced, and bone marrow-derived macrophages from Lyz- GRP78-/- mice demonstrated a selective increase in IL-6 expression. Serum IL-13 levels were elevated by >4-fold in Lyz- GRP78-/- mice, and IL-6 stimulated the myocyte expression of IL-13 and IL-13 receptor. Lastly, recombinant IL-13 acutely increased glucose metabolism in Lyz- GRP78-/- mice. Taken together, our data indicate that GRP78 deficiency activates UPR by increasing ATF-4, and promotes M2-polarization of macrophages with a selective increase in IL-6 secretion. Macrophage-derived IL-6 stimulates the myocyte expression of IL-13 and regulates muscle glucose metabolism in a paracrine manner. Thus, our findings identify a novel crosstalk between macrophages and skeletal muscle in the modulation of obesity-mediated insulin resistance.-Kim, J. H., Lee, E., Friedline, R. H., Suk, S., Jung, D. Y., Dagdeviren, S., Hu, X., Inashima, K., Noh, H. L., Kwon, J. Y., Nambu, A., Huh, J. R., Han, M. S., Davis, R. J., Lee, A. S., Lee, K. W., Kim, J. K. Endoplasmic reticulum chaperone GRP78 regulates macrophage function and insulin resistance in diet-induced obesity.
Subject(s)
Heat-Shock Proteins/metabolism , Insulin Resistance , Macrophages/metabolism , Obesity/metabolism , Activating Transcription Factor 4/metabolism , Animals , Cell Line , Cells, Cultured , Diet, High-Fat/adverse effects , Endoplasmic Reticulum Chaperone BiP , Glucose/metabolism , Heat-Shock Proteins/genetics , Interleukin-13/genetics , Interleukin-13/metabolism , Interleukin-6/genetics , Interleukin-6/metabolism , Macrophages/cytology , Male , Mice , Mice, Inbred C57BL , Muscle Cells/metabolism , Obesity/etiology , Unfolded Protein ResponseABSTRACT
Macrophages are phagocytes that play important roles in health and diseases. Acyl-CoA:cholesterol acyltransferase 1 (ACAT1) converts cellular cholesterol to cholesteryl esters and is expressed in many cell types. Unlike global Acat1 knockout (KO), myeloid-specific Acat1 KO ( Acat1-) does not cause overt abnormalities in mice. Here, we performed analyses in age- and sex-matched Acat1-M/-M and wild-type mice on chow or Western diet and discovered that Acat1-M/-M mice exhibit resistance to Western diet-induced obesity. On both chow and Western diets, Acat1-M/-M mice display decreased adipocyte size and increased insulin sensitivity. When fed with Western diet, Acat1-M/-M mice contain fewer infiltrating macrophages in white adipose tissue (WAT), with significantly diminished inflammatory phenotype. Without Acat1, the Ly6Chi monocytes express reduced levels of integrin-ß1, which plays a key role in the interaction between monocytes and the inflamed endothelium. Adoptive transfer experiment showed that the appearance of leukocytes from Acat1-M/-M mice to the inflamed WAT of wild-type mice is significantly diminished. Under Western diet, Acat1-M/-M causes suppression of multiple proinflammatory genes in WAT. Cell culture experiments show that in RAW 264.7 macrophages, inhibiting ACAT1 with a small-molecule ACAT1-specific inhibitor reduces inflammatory responses to lipopolysaccharide. We conclude that under Western diet, blocking ACAT1 in macrophages attenuates inflammation in WAT. Other results show that Acat1-M/-M does not compromise antiviral immune response. Our work reveals that blocking ACAT1 suppresses diet-induced obesity in part by slowing down monocyte infiltration to WAT as well as by reducing the inflammatory responses of adipose tissue macrophages.
Subject(s)
Acetyl-CoA C-Acetyltransferase/genetics , Acetyl-CoA C-Acetyltransferase/physiology , Diet , Inflammation/genetics , Inflammation/pathology , Insulin Resistance/genetics , Macrophages/pathology , Obesity/genetics , Sterol O-Acyltransferase/genetics , Sterol O-Acyltransferase/physiology , Adipocytes/pathology , Adipose Tissue, White/metabolism , Adipose Tissue, White/pathology , Animals , Cell Size , Female , Glucose Intolerance/genetics , Glucose Intolerance/metabolism , Inflammation/immunology , Integrin beta1/metabolism , Macrophages/immunology , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Obesity/physiopathology , RAW 264.7 CellsABSTRACT
Altered energy balance and insulin resistance are important characteristics of aging. Skeletal muscle is a major site of glucose disposal, and the role of aging-associated inflammation in skeletal muscle insulin resistance remains unclear. To investigate, we examined glucose metabolism in 18-mo-old transgenic mice with muscle-specific overexpression of IL-10 (MIL10) and in wild-type mice during hyperinsulinemic-euglycemic clamping. Despite similar fat mass and energy balance, MIL10 mice were protected from aging-associated insulin resistance with significant increases in glucose infusion rates, whole-body glucose turnover, and skeletal muscle glucose uptake (â¼60%; P < 0.05), as compared to age-matched WT mice. This protective effect was associated with decreased muscle inflammation, but no changes in adipose tissue inflammation in aging MIL10 mice. These results demonstrate the importance of skeletal muscle inflammation in aging-mediated insulin resistance, and our findings further implicate a potential therapeutic role of anti-inflammatory cytokine in the treatment of aging-mediated insulin resistance.-Dagdeviren, S., Jung, D. Y., Friedline, R. H., Noh, H. L., Kim, J. H., Patel, P. R., Tsitsilianos, N., Inashima, K., Tran, D. A., Hu, X., Loubato, M. M., Craige, S. M., Kwon, J. Y., Lee, K. W., Kim, J. K. IL-10 prevents aging-associated inflammation and insulin resistance in skeletal muscle.
Subject(s)
Aging/physiology , Inflammation/metabolism , Insulin Resistance/physiology , Interleukin-10/metabolism , Muscle, Skeletal/metabolism , Animals , Creatine Kinase, MM Form , Energy Metabolism , Interleukin-10/genetics , Male , Mice , Mice, TransgenicABSTRACT
Obesity is characterized by a dysregulated immune system, which may causally associate with insulin resistance and type 2 diabetes. Despite widespread use of nonobese diabetic (NOD) mice, NOD with severe combined immunodeficiency (scid) mutation (SCID) mice, and SCID bearing a null mutation in the IL-2 common γ chain receptor (NSG) mice as animal models of human diseases including type 1 diabetes, the underlying metabolic effects of a genetically altered immune system are poorly understood. For this, we performed a comprehensive metabolic characterization of these mice fed chow or after 6 wk of a high-fat diet. We found that NOD mice had â¼50% less fat mass and were 2-fold more insulin sensitive, as measured by hyperinsulinemic-euglycemic clamp, than C57BL/6 wild-type mice. SCID mice were also more insulin sensitive with increased muscle glucose metabolism and resistant to diet-induced obesity due to increased energy expenditure (â¼10%) and physical activity (â¼40%) as measured by metabolic cages. NSG mice were completely protected from diet-induced obesity and insulin resistance with significant increases in glucose metabolism in peripheral organs. Our findings demonstrate an important role of genetic background, lymphocytes, and cytokine signaling in diet-induced obesity and insulin resistance.
Subject(s)
Insulin Resistance/physiology , Interleukin-2/metabolism , Lymphocytes/metabolism , Mice, Inbred NOD/metabolism , Obesity/metabolism , Animals , Diabetes Mellitus, Experimental/metabolism , Diabetes Mellitus, Experimental/physiopathology , Diabetes Mellitus, Type 2/metabolism , Diabetes Mellitus, Type 2/physiopathology , Diet, High-Fat/adverse effects , Dietary Fats/adverse effects , Energy Metabolism/physiology , Glucose/metabolism , Glucose Clamp Technique/methods , Insulin/metabolism , Lymphocytes/physiology , Male , Mice , Mice, Inbred C57BL , Mice, Inbred NOD/physiology , Muscle, Skeletal/metabolism , Muscle, Skeletal/physiology , Obesity/physiopathology , Signal Transduction/physiologyABSTRACT
Understanding the function of individual microRNA (miRNA) species in mice would require the production of hundreds of loss-of-function strains. To accelerate analysis of miRNA biology in mammals, we combined recombinant adeno-associated virus (rAAV) vectors with miRNA 'tough decoys' (TuDs) to inhibit specific miRNAs. Intravenous injection of rAAV9 expressing anti-miR-122 or anti-let-7 TuDs depleted the corresponding miRNA and increased its mRNA targets. rAAV producing anti-miR-122 TuD but not anti-let-7 TuD reduced serum cholesterol by >30% for 25 weeks in wild-type mice. High-throughput sequencing of liver miRNAs from the treated mice confirmed that the targeted miRNAs were depleted and revealed that TuDs induced miRNA tailing and trimming in vivo. rAAV-mediated miRNA inhibition thus provides a simple way to study miRNA function in adult mammals and a potential therapy for dyslipidemia and other diseases caused by miRNA deregulation.
Subject(s)
Dependovirus/genetics , Genetic Vectors/genetics , MicroRNAs/antagonists & inhibitors , MicroRNAs/genetics , Animals , Base Sequence , Binding Sites , Cell Line , Cholesterol/metabolism , Liver/metabolism , Mice , Mice, Inbred C57BL , MicroRNAs/metabolism , Molecular Sequence Data , RNA, Antisense/genetics , RNA, Antisense/metabolism , Recombinant Proteins/geneticsABSTRACT
The cytoplasmic polyadenylation element binding protein CPEB1 (CPEB) regulates germ cell development, synaptic plasticity, and cellular senescence. A microarray analysis of mRNAs regulated by CPEB unexpectedly showed that several encoded proteins are involved in insulin signaling. An investigation of Cpeb1 knockout mice revealed that the expression of two particular negative regulators of insulin action, PTEN and Stat3, were aberrantly increased. Insulin signaling to Akt was attenuated in livers of CPEB-deficient mice, suggesting that they might be defective in regulating glucose homeostasis. Indeed, when the Cpeb1 knockout mice were fed a high-fat diet, their livers became insulin-resistant. Analysis of HepG2 cells, a human liver cell line, depleted of CPEB demonstrated that this protein directly regulates the translation of PTEN and Stat3 mRNAs. Our results show that CPEB regulated translation is a key process involved in insulin signaling.
Subject(s)
Insulin/metabolism , Liver/metabolism , PTEN Phosphohydrolase/genetics , Protein Biosynthesis/genetics , STAT3 Transcription Factor/genetics , Transcription Factors/genetics , mRNA Cleavage and Polyadenylation Factors/genetics , Animals , Cell Line , Diet, High-Fat , Fibroblasts/cytology , Fibroblasts/metabolism , Gene Expression Regulation , Glucose/metabolism , Humans , Insulin/genetics , Insulin Resistance/genetics , Metabolic Networks and Pathways , Mice , Mice, Knockout , PTEN Phosphohydrolase/metabolism , Polyribosomes/genetics , STAT3 Transcription Factor/metabolism , Transcription Factors/deficiency , Transcription Factors/metabolism , mRNA Cleavage and Polyadenylation Factors/deficiency , mRNA Cleavage and Polyadenylation Factors/metabolismABSTRACT
To investigate the role of GRP78 in adipogenesis and metabolic homeostasis, we knocked down GRP78 in mouse embryonic fibroblasts and 3T3-L1 preadipocytes induced to undergo differentiation into adipocytes. We also created an adipose Grp78-knockout mouse utilizing the aP2 (fatty acid binding protein 4) promoter-driven Cre-recombinase. Adipogenesis was monitored by molecular markers and histology. Tissues were analyzed by micro-CT and electron microscopy. Glucose homeostasis and cytokine analysis were performed. Our results indicate that GRP78 is essential for adipocyte differentiation in vitro. aP2-cre-mediated GRP78 deletion leads to lipoatrophy with â¼90% reduction in gonadal and subcutaneous white adipose tissue and brown adipose tissue, severe growth retardation, and bone defects. Despite severe abnormality in adipose mass and function, adipose Grp78-knockout mice showed normal plasma triglyceride levels, and plasma glucose and insulin levels were reduced by 40-60% compared to wild-type mice, suggesting enhanced insulin sensitivity. The endoplasmic reticulum is grossly expanded in the residual mutant white adipose tissue. Thus, these studies establish that GRP78 is required for adipocyte differentiation, glucose homeostasis, and balanced secretion of adipokines. Unexpectedly, the phenotypes and metabolic parameters of the mutant mice, which showed early postnatal mortality, are uniquely distinct from previously characterized lipodystrophic mouse models.
Subject(s)
Adipogenesis/physiology , Adipose Tissue, Brown/metabolism , Adipose Tissue, White/metabolism , Glucose/metabolism , Heat-Shock Proteins/metabolism , Homeostasis/physiology , 3T3-L1 Cells , Adipocytes/cytology , Adipocytes/metabolism , Adipokines/genetics , Adipokines/metabolism , Adipose Tissue, Brown/cytology , Adipose Tissue, White/cytology , Animals , Cell Differentiation/physiology , Disease Models, Animal , Endoplasmic Reticulum/genetics , Endoplasmic Reticulum/metabolism , Endoplasmic Reticulum Chaperone BiP , Fatty Acid-Binding Proteins/genetics , Fatty Acid-Binding Proteins/metabolism , Gene Deletion , Glucose/genetics , Heat-Shock Proteins/genetics , Lipodystrophy/genetics , Lipodystrophy/metabolism , Lipodystrophy/pathology , Mice , Mice, Knockout , Triglycerides/blood , Triglycerides/geneticsABSTRACT
Obesity is a major cause of metabolic dysfunction-associated steatohepatitis (MASH) and is characterized by inflammation and insulin resistance. Interferon-γ (IFNγ) is a pro-inflammatory cytokine elevated in obesity and modulating macrophage functions. Here, we show that male mice with loss of IFNγ signaling in myeloid cells (Lyz-IFNγR2-/-) are protected from diet-induced insulin resistance despite fatty liver. Obesity-mediated liver inflammation is also attenuated with reduced interleukin (IL)-12, a cytokine primarily released by macrophages, and IL-12 treatment in vivo causes insulin resistance by impairing hepatic insulin signaling. Following MASH diets, Lyz-IFNγR2-/- mice are rescued from developing liver fibrosis, which is associated with reduced fibroblast growth factor (FGF) 21 levels. These results indicate critical roles for IFNγ signaling in macrophages and their release of IL-12 in modulating obesity-mediated insulin resistance and fatty liver progression to MASH. In this work, we identify the IFNγ-IL12 axis in regulating intercellular crosstalk in the liver and as potential therapeutic targets to treat MASH.
Subject(s)
Fatty Liver , Insulin Resistance , Interferon-gamma , Interleukin-12 , Liver , Macrophages , Mice, Knockout , Obesity , Signal Transduction , Animals , Interferon-gamma/metabolism , Interleukin-12/metabolism , Male , Obesity/metabolism , Mice , Fatty Liver/metabolism , Fatty Liver/pathology , Macrophages/metabolism , Liver/metabolism , Liver/pathology , Mice, Inbred C57BL , Diet, High-Fat/adverse effects , Receptors, Interferon/metabolism , Receptors, Interferon/genetics , Interferon gamma Receptor , Liver Cirrhosis/metabolism , Liver Cirrhosis/pathology , Liver Cirrhosis/geneticsABSTRACT
The pathophysiology of obesity and type 2 diabetes in rodents and humans is characterized by low-grade inflammation in adipose tissue and liver. The CD40 receptor and its ligand CD40L initiate immune cell signaling promoting inflammation, but conflicting data on CD40L-null mice confound its role in obesity-associated insulin resistance. Here, we demonstrate that CD40 receptor-deficient mice on a high-fat diet display the expected decrease in hepatic cytokine levels but paradoxically exhibit liver steatosis, insulin resistance, and glucose intolerance compared with their age-matched wild-type controls. Hyperinsulinemic-euglycemic clamp studies also demonstrated insulin resistance in glucose utilization by the CD40-null mice compared with wild-type mice. In contrast to liver, adipose tissue in CD40-deficient animals harbors elevated cytokine levels and infiltration of inflammatory cells, particularly macrophages and CD8(+) effector T cells. In addition, ex vivo explants of epididymal adipose tissue from CD40(-/-) mice display elevated basal and isoproterenol-stimulated lipolysis, suggesting a potential increase of lipid efflux from visceral fat to the liver. These findings reveal that 1) CD40-null mice represent an unusual model of hepatic steatosis with reduced hepatic inflammation, and 2) CD40 unexpectedly functions in adipose tissue to attenuate its inflammation in obesity, thereby protecting against hepatic steatosis.
Subject(s)
Adipose Tissue/pathology , CD40 Antigens/deficiency , Fatty Liver/genetics , Fatty Liver/pathology , Inflammation/genetics , Inflammation/pathology , Insulin Resistance/genetics , Obesity/genetics , Obesity/pathology , Adipocytes/metabolism , Animals , Blotting, Western , Diet , Disease Progression , Enzyme-Linked Immunosorbent Assay , Flow Cytometry , Glucose Clamp Technique , Glucose Tolerance Test , Lipid Metabolism/genetics , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , RNA/biosynthesis , RNA/genetics , Real-Time Polymerase Chain ReactionABSTRACT
Obesity is a major cause of insulin resistance, and weight loss is shown to improve glucose homeostasis. But the underlying mechanism and the role of inflammation remain unclear. Male C57BL/6 mice were fed a high-fat diet (HFD) for 12 wk. After HFD, weight loss was induced by changing to a low-fat diet (LFD) or exercise with continuous HFD. The weight loss effects on energy balance and insulin sensitivity were determined using metabolic cages and hyperinsulinemic euglycemic clamps in awake mice. Diet and exercise intervention for 3 wk caused a modest weight loss and improved glucose homeostasis. Weight loss dramatically reduced local inflammation in skeletal muscle, liver, and heart but not in adipose tissue. Exercise-mediated weight loss increased muscle glucose metabolism without affecting Akt phosphorylation or lipid levels. LFD-mediated weight loss reduced lipid levels and improved insulin sensitivity selectively in liver. Both weight loss interventions improved cardiac glucose metabolism. These results demonstrate that a short-term weight loss with exercise or diet intervention attenuates obesity-induced local inflammation and selectively improves insulin sensitivity in skeletal muscle and liver. Our findings suggest that local factors, not adipose tissue inflammation, are involved in the beneficial effects of weight loss on glucose homeostasis.
Subject(s)
Adipose Tissue/pathology , Inflammation/pathology , Insulin Resistance/physiology , Obesity/pathology , Weight Loss/physiology , Animals , Antigens, CD/metabolism , Antigens, Differentiation, Myelomonocytic/metabolism , Blotting, Western , Body Composition/physiology , Diet , Energy Metabolism/physiology , Interleukins/blood , Liver/pathology , Mice , Mice, Inbred C57BL , Mice, Obese , Muscle, Skeletal/pathology , Myocardium/pathology , Physical Conditioning, Animal/physiology , Signal Transduction/physiologyABSTRACT
The liver plays a central role in regulating glucose and lipid metabolism. Aberrant insulin action in the liver is a major driver of selective insulin resistance, in which insulin fails to suppress glucose production but continues to activate lipogenesis in the liver, resulting in hyperglycemia and hypertriglyceridemia. The underlying mechanisms of selective insulin resistance are not fully understood. Here It is shown that hepatic membrane phospholipid composition controlled by lysophosphatidylcholine acyltransferase 3 (LPCAT3) regulates insulin signaling and systemic glucose and lipid metabolism. Hyperinsulinemia induced by high-fat diet (HFD) feeding augments hepatic Lpcat3 expression and membrane unsaturation. Loss of Lpcat3 in the liver improves insulin resistance and blunts lipogenesis in both HFD-fed and genetic ob/ob mouse models. Mechanistically, Lpcat3 deficiency directly facilitates insulin receptor endocytosis, signal transduction, and hepatic glucose production suppression and indirectly enhances fibroblast growth factor 21 (FGF21) secretion, energy expenditure, and glucose uptake in adipose tissue. These findings identify hepatic LPCAT3 and membrane phospholipid composition as a novel regulator of insulin sensitivity and provide insights into the pathogenesis of selective insulin resistance.
Subject(s)
Insulin Resistance , Mice , Animals , Insulin Resistance/genetics , Phospholipids/metabolism , Liver/metabolism , Glucose/metabolism , Insulin/metabolism , 1-Acylglycerophosphocholine O-Acyltransferase/metabolismABSTRACT
OBJECTIVES: Nuclear receptor interacting protein 1 (NRIP1) suppresses energy expenditure via repression of nuclear receptors, and its depletion markedly elevates uncoupled respiration in mouse and human adipocytes. We tested whether NRIP1 deficient adipocytes implanted into obese mice would enhance whole body metabolism. Since ß-adrenergic signaling through cAMP strongly promotes adipocyte thermogenesis, we tested whether the effects of NRIP1 knock-out (NRIP1KO) require the cAMP pathway. METHODS: NRIP1KO adipocytes were implanted in recipient high-fat diet (HFD) fed mice and metabolic cage studies conducted. The Nrip1 gene was disrupted by CRISPR in primary preadipocytes isolated from control vs adipose selective GsαKO (cAdGsαKO) mice prior to differentiation to adipocytes. Protein kinase A inhibitor was also used. RESULTS: Implanting NRIP1KO adipocytes into HFD fed mice enhanced whole-body glucose tolerance by increasing insulin sensitivity, reducing adiposity, and enhancing energy expenditure in the recipients. NRIP1 depletion in both control and GsαKO adipocytes was equally effective in upregulating uncoupling protein 1 (UCP1) and adipocyte beiging, while ß-adrenergic signaling by CL 316,243 was abolished in GsαKO adipocytes. Combining NRIP1KO with CL 316,243 treatment synergistically increased Ucp1 gene expression and increased the adipocyte subpopulation responsive to beiging. Estrogen-related receptor α (ERRα) was dispensable for UCP1 upregulation by NRIPKO. CONCLUSIONS: The thermogenic effect of NRIP1 depletion in adipocytes causes systemic enhancement of energy expenditure when such adipocytes are implanted into obese mice. Furthermore, NRIP1KO acts independently but cooperatively with the cAMP pathway in mediating its effect on adipocyte beiging.
Subject(s)
Adipocytes , Signal Transduction , Mice , Humans , Animals , Nuclear Receptor Interacting Protein 1/metabolism , Mice, Obese , Adipocytes/metabolism , Obesity/metabolism , Thermogenesis/geneticsABSTRACT
TRPM2 Ca(2+)-permeable cation channel is widely expressed and activated by markers of cellular stress. Since inflammation and stress play a major role in insulin resistance, we examined the role of TRPM2 Ca(2+) channel in glucose metabolism. A 2-h hyperinsulinemic euglycemic clamp was performed in TRPM2-deficient (KO) and wild-type mice to assess insulin sensitivity. To examine the effects of diet-induced obesity, mice were fed a high-fat diet for 4-10 mo, and metabolic cage and clamp studies were conducted in conscious mice. TRPM2-KO mice were more insulin sensitive partly because of increased glucose metabolism in peripheral organs. After 4 mo of high-fat feeding, TRPM2-KO mice were resistant to diet-induced obesity, and this was associated with increased energy expenditure and elevated expressions of PGC-1α, PGC-1ß, PPARα, ERRα, TFAM, and MCAD in white adipose tissue. Hyperinsulinemic euglycemic clamps showed that TRPM2-KO mice were more insulin sensitive, with increased Akt and GSK-3ß phosphorylation in heart. Obesity-mediated inflammation in adipose tissue and liver was attenuated in TRPM2-KO mice. Overall, TRPM2 deletion protected mice from developing diet-induced obesity and insulin resistance. Our findings identify a novel role of TRPM2 Ca(2+) channel in the regulation of energy expenditure, inflammation, and insulin resistance.
Subject(s)
Energy Metabolism/physiology , Glucose/metabolism , TRPM Cation Channels/physiology , Animals , Blotting, Western , Body Composition/physiology , Body Weight/physiology , Calmodulin/metabolism , Calorimetry, Indirect , Dietary Fats/pharmacology , Glucose Clamp Technique , Glucose Tolerance Test , Immunoprecipitation , Inflammation/metabolism , Insulin Receptor Substrate Proteins/metabolism , Insulin Resistance/physiology , Mice , Mice, Knockout , Myocardium/enzymology , Myocardium/metabolism , Oxygen Consumption/physiology , Phosphorylation , RNA/biosynthesis , RNA/genetics , Superoxide Dismutase/metabolismABSTRACT
Estrogens protect against weight gain and metabolic disruption in women and female rodents. Aberrations in the gut microbiota composition are linked to obesity and metabolic disorders. Furthermore, estrogen-mediated protection against diet-induced metabolic disruption is associated with modifications in gut microbiota. In this study, we tested if estradiol (E2)-mediated protection against obesity and metabolic disorders in female mice is dependent on gut microbiota. Specifically, we tested if fecal microbiota transplantation (FMT) from E2-treated lean female mice, supplemented with or without Akkermansia muciniphila, prevented high fat diet (HFD)-induced body weight gain, fat mass gain, and hyperglycemia in female recipients. FMT from, and cohousing with, E2-treated lean donors was not sufficient to transfer the metabolic benefits to the E2-deficient female recipients. Moreover, FMT from lean donors supplemented with A. muciniphila exacerbated HFD-induced hyperglycemia in E2-deficient recipients, suggesting its detrimental effect on the metabolic health of E2-deficient female rodents fed a HFD. Given that A. muciniphila attenuates HFD-induced metabolic insults in males, the present findings suggest a sex difference in the impact of this microbe on metabolic health.
Subject(s)
Diet, High-Fat , Hyperglycemia , Female , Mice , Male , Animals , Diet, High-Fat/adverse effects , Akkermansia , Fecal Microbiota Transplantation , Mice, Inbred C57BL , Obesity/etiology , Obesity/therapy , Obesity/metabolism , Weight GainABSTRACT
The signaling mechanisms underlying adipose thermogenesis have not been fully elucidated. Particularly, the involvement of adipokines that are selectively expressed in brown adipose tissue (BAT) and beige adipocytes remains to be investigated. Here we show that a previously uncharacterized adipokine (UPF0687 protein / human C20orf27 homolog) we named as Adissp (Adipose-secreted signaling protein) is a key regulator for white adipose tissue (WAT) thermogenesis and glucose homeostasis. Adissp expression is adipose-specific and highly BAT-enriched, and its secretion is stimulated by ß3-adrenergic activation. Gain-of-functional studies collectively showed that secreted Adissp promotes WAT thermogenesis, improves glucose homeostasis, and protects against obesity. Adipose-specific Adissp knockout mice are defective in WAT browning, and are susceptible to high fat diet-induced obesity and hyperglycemia. Mechanistically, Adissp binds to a putative receptor on adipocyte surface and activates protein kinase A independently of ß-adrenergic signaling. These results establish BAT-enriched Adissp as a major upstream signaling component in thermogenesis and offer a potential avenue for the treatment of obesity and diabetes.
Subject(s)
Adipokines , Adipose Tissue, Brown , Mice , Animals , Humans , Adipose Tissue, Brown/metabolism , Thermogenesis , Adipose Tissue, White/metabolism , Obesity/metabolism , Glucose/metabolism , Adrenergic Agents/metabolism , Adipocytes, Brown/metabolism , Energy MetabolismABSTRACT
FOXP3-expressing regulatory T (Treg) cells are vital for maintaining peripheral T cell tolerance and homeostasis. The mechanisms by which FOXP3 target genes orchestrate context-dependent Treg cell function are largely unknown. In this study we show that in mouse peripheral lymphocytes the Drosophila Disabled-2 (Dab2) homolog, a gene that is involved in enhancing TGFbeta responses, is exclusively expressed in FOXP3+ regulatory T cells. Dab2 is a direct target of FOXP3, and regulatory T cells lacking DAB2 are functionally impaired in vitro and in vivo. However, not all aspects of Treg cell function are perturbed, and DAB2 appears to be dispensable for Treg cell function in maintaining naive T cell homeostasis.