Spred2 Regulates High Fat Diet-Induced Adipose Tissue Inflammation, and Metabolic Abnormalities in Mice.
ABSTRACT: Chronic low-grade inflammation in visceral adipose tissues triggers the development of obesity-related insulin resistance, leading to the metabolic syndrome, a serious health condition with higher risk of cardiovascular disease, diabetes, and stroke. In the present study, we investigated whether Sprouty-related EVH1-domain-containing protein 2 (Spred2), a negative regulator of the Ras/Raf/ERK/MAPK pathway, plays a role in the development of high fat diet (HFD)-induced obesity, adipose tissue inflammation, metabolic abnormalities, and insulin resistance. Spred2 knockout (KO) mice, fed with HFD, exhibited an augmented body weight gain, which was associated with enhanced adipocyte hypertrophy in mesenteric white adipose tissue (mWAT) and deteriorated dyslipidemia, compared with wild-type (WT) controls. The number of infiltrating macrophages with a M1 phenotype, and the crown-like structures, composed of macrophages surrounding dead or dying adipocytes, were more abundant in Spred2 KO-mWAT compared to in WT-mWAT. Exacerbated adipose tissue inflammation in Spred2 KO mice led to aggravated insulin resistance and fatty liver disease. To analyze the mechanism(s) that caused adipose tissue inflammation, cytokine response in mWAT was investigated. Stromal vascular fraction that contained macrophages from Spred2 KO-mWAT showed elevated levels of tumor necrosis factor ? (TNF?) and monocyte chemoattractant protein-1 (MCP-1/CCL2) compared with those from WT-mWAT. Upon stimulation with palmitate acid (PA), bone marrow-derived macrophages (BMDMs) derived from Spred2 KO mice secreted higher levels of TNF? and MCP-1 than those from WT mice with enhanced ERK activation. U0126, a MEK inhibitor, reduced the PA-induced cytokine response. Taken together, these results suggested that Spred2, in macrophages, negatively regulates high fat diet-induced obesity, adipose tissue inflammation, metabolic abnormalities, and insulin resistance by inhibiting the ERK/MAPK pathway. Thus, Spred2 represents a potential therapeutic tool for the prevention of insulin resistance and resultant metabolic syndrome.
Project description:GPR105, a G protein-coupled receptor for UDP-glucose, is highly expressed in several human tissues and participates in the innate immune response. Because inflammation has been implicated as a key initial trigger for type 2 diabetes, we hypothesized that GPR105 (official gene name: P2RY14) might play a role in the initiation of inflammation and insulin resistance in obesity. To this end, we investigated glucose metabolism in GPR105 knockout (KO) and wild-type (WT) mice fed a high-fat diet (HFD). We also examined whether GPR105 regulates macrophage recruitment to liver or adipose tissues by in vivo monocyte tracking and in vitro chemotaxis experiments, followed by transplantation of bone marrow from either KO or WT donors to WT recipients. Our data show that genetic deletion of GPR105 confers protection against HFD-induced insulin resistance, with reduced macrophage infiltration and inflammation in liver, and increased insulin-stimulated Akt phosphorylation in liver, muscle, and adipose tissue. By tracking monocytes from either KO or WT donors, we found that fewer KO monocytes were recruited to the liver of WT recipients. Furthermore, we observed that uridine 5-diphosphoglucose enhanced the in vitro migration of bone marrow-derived macrophages from WT but not KO mice, and that plasma uridine 5-diphosphoglucose levels were significantly higher in obese versus lean mice. Finally, we confirmed that insulin sensitivity improved in HFD mice with a myeloid cell-specific deletion of GPR105. These studies indicate that GPR105 ablation mitigates HFD-induced insulin resistance by inhibiting macrophage recruitment and tissue inflammation. Hence GPR105 provides a novel link between innate immunity and metabolism.
Project description:Recent accumulating evidence suggests that innate immunity is associated with obesity-induced chronic inflammation and metabolic disorders. Here, we show that a Toll-like receptor (TLR) protein, radioprotective 105 (RP105)/myeloid differentiation protein (MD)-1 complex, contributes to high-fat diet (HFD)-induced obesity, adipose tissue inflammation, and insulin resistance. An HFD dramatically increased RP105 mRNA and protein expression in stromal vascular fraction of epididymal white adipose tissue (eWAT) in wild-type (WT) mice. RP105 mRNA expression also was significantly increased in the visceral adipose tissue of obese human subjects relative to nonobese subjects. The RP105/MD-1 complex was expressed by most adipose tissue macrophages (ATMs). An HFD increased RP105/MD-1 expression on the M1 subset of ATMs that accumulate in eWAT. Macrophages also acquired this characteristic in coculture with 3T3-L1 adipocytes. RP105 knockout (KO) and MD-1 KO mice had less HFD-induced adipose tissue inflammation, hepatic steatosis, and insulin resistance compared with wild-type (WT) and TLR4 KO mice. Finally, the saturated fatty acids, palmitic and stearic acids, are endogenous ligands for TLR4, but they did not activate RP105/MD-1. Thus, the RP105/MD-1 complex is a major mediator of adipose tissue inflammation independent of TLR4 signaling and may represent a novel therapeutic target for obesity-associated metabolic disorders.
Project description:Rapid and adequate mucosal healing is important for a remission of ulcerative colitis (UC) patients. Here, we examined whether Spred2, a member of the Sprouty-related EVH1-domain-containing proteins that inhibit the Ras/Raf/ERK pathway, plays a role in colonic mucosal homeostasis and inflammation by using Spred2 knockout (KO) mice. We first detected increased epithelial cell proliferation and cadherin 1 expression in the colon of naïve Spred2 KO mice compared to wild-type mice. Interestingly, Spred2 KO mice were resistant to dextran sulfate sodium (DSS)-induced acute colitis as indicated by lower levels of body weight loss and disease activity index. Histologically, epithelial cell injury and inflammation were milder in the colonic mucosa of Spred2 KO mice on day 3 and almost undetectable by day 8. Experiments with bone chimeric mice indicated that Spred2-deficiency in non-hematopoietic cells was responsible for the reduced sensitivity to DSS. Finally, Spred2 KO mice developed significantly fewer tumors in response to azoxymethane plus DSS. Taken together, our results demonstrate, for the first time, that Spred2 plays an important role in the regulation of colonic epithelial cell proliferation and inflammation by potentially down-regulating the activation of ERK. Thus, Spred2 may be a new therapeutic target for the treatment of UC.
Project description:The inflammasome has been recently implicated in obesity-associated dys-metabolism. However, of its products, the specific role of IL-1? was clinically demonstrated to mediate only the pancreatic beta-cell demise, and in mice mainly the intra-hepatic manifestations of obesity. Yet, it remains largely unknown if IL-1?, a cytokine believed to mainly function locally, could regulate dysfunctional inter-organ crosstalk in obesity. Here we show that High-fat-fed (HFF) mice exhibited a preferential increase of IL-1? in portal compared to systemic blood. Moreover, portally-drained mesenteric fat transplantation from IL-1?KO donors resulted in lower pyruvate-glucose flux compared to mice receiving wild-type (WT) transplant. These results raised a putative endocrine function for visceral fat-derived IL-1? in regulating hepatic gluconeogenic flux. IL-1?KO mice on HFF exhibited only a minor or no increase in adipose expression of pro-inflammatory genes (including macrophage M1 markers), Mac2-positive crown-like structures and CD11b-F4/80-double-positive macrophages, all of which were markedly increased in WT-HFF mice. Further consistent with autocrine/paracrine functions of IL-1? within adipose tissue, adipose tissue macrophage lipid content was increased in WT-HFF mice, but significantly less in IL-1?KO mice. Ex-vivo, adipose explants co-cultured with primary hepatocytes from WT or IL-1-receptor (IL-1RI)-KO mice suggested only a minor direct effect of adipose-derived IL-1? on hepatocyte insulin resistance. Importantly, although IL-1?KOs gained weight similarly to WT-HFF, they had larger fat depots with similar degree of adipocyte hypertrophy. Furthermore, adipogenesis genes and markers (pparg, cepba, fabp4, glut4) that were decreased by HFF in WT, were paradoxically elevated in IL-1?KO-HFF mice. These local alterations in adipose tissue inflammation and expansion correlated with a lower liver size, less hepatic steatosis, and preserved insulin sensitivity. Collectively, we demonstrate that by promoting adipose inflammation and limiting fat tissue expandability, IL-1? supports ectopic fat accumulation in hepatocytes and adipose-tissue macrophages, contributing to impaired fat-liver crosstalk in nutritional obesity.
Project description:Obesity is a major risk factor for the development of insulin resistance and type 2 diabetes. However, the mechanisms that trigger the underlying adipose tissues inflammation are not completely understood. Here, we show that the E3 ubiquitin ligase March1 controls the phenotypic and functional properties of CD8+ T cells in mice white adipose tissue. In a diet-induced obesity model, mice lacking March1 [March1 knockout (KO)] show increased insulin resistance compared to their WT counterparts. Also, in obese March1 KO mice, the proportions of effector/memory (Tem) and resident/memory (Trm) CD8+ T cells were higher in the visceral adipose tissue, but not in the spleen. The effect of March1 on insulin resistance and on the phenotype of adipose tissue CD8+ T cells was independent of major histocompatibility complex class II ubiquitination. Interestingly, we adoptively transferred either WT or March1 KO splenic CD8+ T cells into obese WT chimeras that had been reconstituted with Rag1-deficient bone marrow. We observed an enrichment of Tem and Trm cells and exacerbated insulin resistance in mice that received March1 KO CD8 T cells. Mechanistically, we found that March1 deficiency alters the metabolic activity of CD8+ T cells. Our results provide additional evidence of the involvement of CD8+ T cells in adipose tissue inflammation and suggest that March1 controls the metabolic reprogramming of these cells.
Project description:Obesity-induced inflammation is a key component of systemic insulin resistance, which is a hallmark of type 2 diabetes. A major driver of this inflammation/insulin resistance syndrome is the accumulation of proinflammatory macrophages in adipose tissue and liver. We found that the orphan GPCR Gpr21 was highly expressed in the hypothalamus and macrophages of mice and that whole-body KO of this receptor led to a robust improvement in glucose tolerance and systemic insulin sensitivity and a modest lean phenotype. The improvement in insulin sensitivity in the high-fat diet-fed (HFD-fed) Gpr21 KO mouse was traced to a marked reduction in tissue inflammation caused by decreased chemotaxis of Gpr21 KO macrophages into adipose tissue and liver. Furthermore, mice lacking macrophage expression of Gpr21 were protected from HFD-induced inflammation and displayed improved insulin sensitivity. Results of in vitro chemotaxis studies in human monocytes suggested that the defect in chemotaxis observed ex vivo and in vivo in mice is also translatable to humans. Cumulatively, our data indicate that GPR21 has a critical function in coordinating macrophage proinflammatory activity in the context of obesity-induced insulin resistance.
Project description:OBJECTIVE:Chronic inflammation of adipose tissues contributes to obesity-triggered insulin resistance. Unfortunately, the potential molecular mechanisms regarding obesity-associated systemic inflammation and metabolic disorder remain complicated. Here, we report that inactive rhomboid-like protein 2 (iRhom2) was increased in overweight mice with adipose inflammation. METHODS:Mice with deletion of iRhom2 on a C57BL/6J background, mice without deletion of this gene (controls), and mice with deficiency of iRhom2 only in myeloid cells were fed a standard chow diet (SCD) or a high-fat diet (HFD; 60% fat calories). Then the adipose tissues or bone marrow cells were isolated for the further detection. RESULTS:After 16 weeks on a high-fat diet (HFD), obesity, chronic inflammation in adipose tissues, and insulin resistance were markedly mitigated in iRhom2 knockout (iRhom2 KO) mice, whereas these parameters were exaggerated in iRhom2 overactivated mice. The adverse influences of iRhom2 on adipose inflammation and associated pathologies were determined in db/db mice. We further demonstrated that, in response to an HFD, iRhom2 KO mice and mice with deletion only in the myeloid cells showed less severe adipose inflammation and insulin resistance than control groups. Conversely, transplantation of bone marrow cells from normal mice to iRhom2 KO mice unleashed severe systemic inflammation and metabolic dysfunction after HFD ingestion. CONCLUSION:We identified iRhom2 as a key regulator that promotes obesity-associated metabolic disorders. Loss of iRhom2 from macrophages in adipose tissues may indirectly restrain inflammation and insulin resistance via blocking crosslinks between macrophages and adipocytes. Hence, iRhom2 may be a therapeutic target for obesity-induced metabolic dysfunction.
Project description:Individuals who are obese are frequently insulin resistant, putting them at increased risk of developing type 2 diabetes and its associated adverse health conditions. The accumulation in adipose tissue of macrophages in an inflammatory state is a hallmark of obesity-induced insulin resistance. Here, we reveal a role for AMPK ?1 in protecting macrophages from inflammation under high lipid exposure. Genetic deletion of the AMPK ?1 subunit in mice (referred to herein as ?1(-/-) mice) reduced macrophage AMPK activity, acetyl-CoA carboxylase phosphorylation, and mitochondrial content, resulting in reduced rates of fatty acid oxidation. ?1(-/-) macrophages displayed increased levels of diacylglycerol and markers of inflammation, effects that were reproduced in WT macrophages by inhibiting fatty acid oxidation and, conversely, prevented by pharmacological activation of AMPK ?1-containing complexes. The effect of AMPK ?1 loss in macrophages was tested in vivo by transplantation of bone marrow from WT or ?1(-/-) mice into WT recipients. When challenged with a high-fat diet, mice that received ?1(-/-) bone marrow displayed enhanced adipose tissue macrophage inflammation and liver insulin resistance compared with animals that received WT bone marrow. Thus, activation of AMPK ?1 and increasing fatty acid oxidation in macrophages may represent a new therapeutic approach for the treatment of insulin resistance.
Project description:The mitogen-activated protein kinase (MAPK) pathways are involved in many cellular processes, including the development of fibrosis. Here, we examined the role of Sprouty-related EVH-1-domain-containing protein (Spred) 2, a negative regulator of the MAPK-ERK pathway, in the development of bleomycin (BLM)-induced pulmonary fibrosis (PF). Compared to WT mice, Spred2?/? mice developed milder PF with increased proliferation of bronchial epithelial cells. Spred2?/? lung epithelial cells or MLE-12 cells treated with spred2 siRNA proliferated faster than control cells in vitro. Spred2?/? and WT macrophages produced similar levels of TNF? and MCP-1 in response to BLM or lipopolysaccharide and myeloid cell-specific deletion of Spred2 in mice had no effect. Spred2?/? fibroblasts proliferated faster and produced similar levels of MCP-1 compared to WT fibroblasts. Spred2 mRNA was almost exclusively detected in bronchial epithelial cells of naïve WT mice and it accumulated in approximately 50% of cells with a characteristic of Clara cells, 14 days after BLM treatment. These results suggest that Spred2 is involved in the regulation of tissue repair after BLM-induced lung injury and increased proliferation of lung bronchial cells in Spred2?/? mice may contribute to faster tissue repair. Thus, Spred2 may present a new therapeutic target for the treatment of PF.
Project description:OBJECTIVE: The study objective was to evaluate the possible role of the macrophage molecule CD14 in insulin resistance. RESEARCH DESIGN AND METHODS: The effects of recombinant human soluble CD14 (rh-sCD14) on insulin sensitivity (clamp procedure) and adipose tissue gene expression were evaluated in wild-type (WT) mice, high fat-fed mice, ob/ob mice, and CD14 knockout (KO) mice. We also studied WT mice grafted with bone marrow stem cells from WT donor mice and CD14 KO mice. Finally, CD14 was evaluated in human adipose tissue and during differentiation of human preadipocytes. RESULTS: rh-sCD14 led to increased insulin action in WT mice, high-fat-fed mice, and ob/ob mice, but not in CD14 KO mice, in parallel to a marked change in the expression of 3,479 genes in adipose tissue. The changes in gene families related to lipid metabolism were most remarkable. WT mice grafted with bone marrow stem cells from WT donor mice became insulin resistant after a high-fat diet. Conversely, WT mice grafted with cells from CD14 KO mice resisted the occurrence of insulin resistance in parallel to decreased mesenteric adipose tissue inflammatory gene expression. Glucose intolerance did not worsen in CD14 KO mice grafted with bone marrow stem cells from high fat-fed WT mice when compared with recipient KO mice grafted with cells from CD14 KO donor mice. CD14 gene expression was increased in whole adipose tissue and adipocytes from obese humans and further increased after tumor necrosis factor-?. CONCLUSIONS: CD14 modulates adipose tissue inflammatory activity and insulin resistance.