The Dual-Specificity Phosphatase 2 (DUSP2) Does Not Regulate Obesity-Associated Inflammation or Insulin Resistance in Mice

Alterations in the immune cell profile and the induction of inflammation within adipose tissue are a hallmark of obesity in mice and humans. Dual-specificity phosphatase 2 (DUSP2) is widely expressed within the immune system and plays a key role promoting immune and inflammatory responses dependent on mitogen-activated protein kinase (MAPK) activity. We hypothesised that the absence of DUSP2 would protect mice against obesity-associated inflammation and insulin resistance. Accordingly, male and female littermate mice that are either wild-type (wt) or homozygous for a germ-line null mutation of the dusp2 gene (dusp2^−/−) were fed either a standard chow diet (SCD) or high fat diet (HFD) for 12 weeks prior to metabolic phenotyping. Compared with mice fed the SCD, all mice consuming the HFD became obese, developed glucose intolerance and insulin resistance, and displayed increased macrophage recruitment and markers of inflammation in epididymal white adipose tissue. The absence of DUSP2, however, had no effect on the development of obesity or adipose tissue inflammation. Whole body insulin sensitivity in male mice was unaffected by an absence of DUSP2 in response to either the SCD or HFD; however, HFD-induced insulin resistance was slightly, but significantly, reduced in female dusp2^−/− mice. In conclusion, DUSP2 plays no role in regulating obesity-associated inflammation and only a minor role in controlling insulin sensitivity following HFD in female, but not male, mice. These data indicate that rather than DUSP2 being a pan regulator of MAPK dependent immune cell mediated inflammation, it appears to differentially regulate inflammatory responses that have a MAPK component.     

12/15-Lipoxygenase Is Required for the Early Onset of High Fat Diet-Induced Adipose Tissue Inflammation and Insulin Resistance in Mice

Background

Recent understanding that insulin resistance is an inflammatory condition necessitates searching for genes that regulate inflammation in insulin sensitive tissues. 12/15-lipoxygenase (12/15LO) regulates the expression of proinflammatory cytokines and chemokines and is implicated in the early development of diet-induced atherosclerosis. Thus, we tested the hypothesis that 12/15LO is involved in the onset of high fat diet (HFD)-induced insulin resistance.

Methodology/Principal Findings

Cells over-expressing 12/15LO secreted two potent chemokines, MCP-1 and osteopontin, implicated in the development of insulin resistance. We assessed adipose tissue inflammation and whole body insulin resistance in wild type (WT) and 12/15LO knockout (KO) mice after 2–4 weeks on HFD. In adipose tissue from WT mice, HFD resulted in recruitment of CD11b⁺, F4/80⁺ macrophages and elevated protein levels of the inflammatory markers IL-1β, IL-6, IL-10, IL-12, IFNγ, Cxcl1 and TNFα. Remarkably, adipose tissue from HFD-fed 12/15LO KO mice was not infiltrated by macrophages and did not display any increase in the inflammatory markers compared to adipose tissue from normal chow-fed mice. WT mice developed severe whole body (hepatic and skeletal muscle) insulin resistance after HFD, as measured by hyperinsulinemic euglycemic clamp. In contrast, 12/15LO KO mice exhibited no HFD-induced change in insulin-stimulated glucose disposal rate or hepatic glucose output during clamp studies. Insulin-stimulated Akt phosphorylation in muscle tissue from HFD-fed mice was significantly greater in 12/15LO KO mice than in WT mice.

Conclusions

These results demonstrate that 12/15LO mediates early stages of adipose tissue inflammation and whole body insulin resistance induced by high fat feeding.     

Macrophage Migration Inhibitory Factor Deficiency Ameliorates High-Fat Diet Induced Insulin Resistance in Mice with Reduced Adipose Inflammation and Hepatic Steatosis

Macrophage infiltration is a critical determinant of high-fat diet induced adipose tissue inflammation and insulin resistance. The precise mechanisms underpinning the initiation of macrophage recruitment and activation are unclear. Macrophage migration inhibitory factor (MIF), a pro-inflammatory cytokine, displays chemokine-like properties. Circulating MIF levels are elevated during obesity however its role in high-fat diet induced adipose inflammation and insulin resistance remains elusive. Wildtype and MIF^−/− C57Bl\6J mice were fed chow or high-fat diet. Body weight and food intake was assessed. Glucose homeostasis was monitored by glucose and insulin tolerance tests. Adipose tissue macrophage recruitment and adipose tissue insulin sensitivity was evaluated. Cytokine secretion from stromal vascular fraction, adipose explants and bone marrow macrophages was measured. Inflammatory signature and insulin sensitivity of 3T3-L1-adipocytes co-cultured with wildtype and MIF^−/− macrophage was quantified. Hepatic triacylglyceride levels were assessed. MIF^−/− exhibited reduced weight gain. Age and weight-matched obese MIF^−/− mice exhibited improved glucose homeostasis coincident with reduced adipose tissue M1 macrophage infiltration. Obese MIF^−/− stromal vascular fraction secreted less TNFα and greater IL-10 compared to wildtype. Activation of JNK was impaired in obese MIF^−/−adipose, concomitant with pAKT expression. 3T3-L1-adipocytes cultured with MIF^−/− macrophages had reduced pro-inflammatory cytokine secretion and improved insulin sensitivity, effects which were also attained with MIF inhibitor ISO-1. MIF^−/− liver exhibited reduced hepatic triacyglyceride accumulation, enhanced pAKT expression and reduced NFκB activation. MIF deficiency partially protects from high-fat diet induced insulin resistance by attenuating macrophage infiltration, ameliorating adipose inflammation, which improved adipocyte insulin resistance ex vivo. MIF represents a potential therapeutic target for treatment of high-fat diet induced insulin resistance.     

Cannabinoid CB2 Receptor Potentiates Obesity-Associated Inflammation,Insulin Resistance and Hepatic Steatosis

Background

Obesity-associated inflammation is of critical importance in the development of insulin resistance and non-alcoholic fatty liver disease. Since the cannabinoid receptor CB2 regulates innate immunity, the aim of the present study was to investigate its role in obesity-induced inflammation, insulin resistance and fatty liver.

Methodology

Murine obesity models included genetically leptin-deficient ob/ob mice and wild type (WT) mice fed a high fat diet (HFD), that were compared to their lean counterparts. Animals were treated with pharmacological modulators of CB2 receptors. Experiments were also performed in mice knock-out for CB2 receptors (Cnr2 −/−).

Principal Findings

In both HFD-fed WT mice and ob/ob mice, Cnr2 expression underwent a marked induction in the stromal vascular fraction of epididymal adipose tissue that correlated with increased fat inflammation. Treatment with the CB2 agonist JWH-133 potentiated adipose tissue inflammation in HFD-fed WT mice. Moreover, cultured fat pads isolated from ob/ob mice displayed increased Tnf and Ccl2 expression upon exposure to JWH-133. In keeping, genetic or pharmacological inactivation of CB2 receptors decreased adipose tissue macrophage infiltration associated with obesity, and reduced inductions of Tnf and Ccl2 expressions. In the liver of obese mice, Cnr2 mRNA was only weakly induced, and CB2 receptors moderately contributed to liver inflammation. HFD-induced insulin resistance increased in response to JWH-133 and reduced in Cnr2 −/− mice. Finally, HFD-induced hepatic steatosis was enhanced in WT mice treated with JWH-133 and blunted in Cnr2 −/− mice.

Conclusion/Significance

These data unravel a previously unrecognized contribution of CB2 receptors to obesity-associated inflammation, insulin resistance and non-alcoholic fatty liver disease, and suggest that CB2 receptor antagonists may open a new therapeutic approach for the management of obesity-associated metabolic disorders.     

Recombinant Acylation Stimulating Protein Administration to C3?/? Mice Increases Insulin Resistance via Adipocyte Inflammatory Mechanisms

Background

Complement 3 (C3), a key component of the innate immune system, is involved in early inflammatory responses. Acylation stimulating protein (ASP; aka C3adesArg), a C3 cleavage product, is produced in adipose tissue and stimulates lipid storage. We hypothesized that, depending on the diet, chronic ASP administration in C3^−/− mice would affect lipid metabolism and insulin sensitivity via an adaptive adipose tissue inflammatory response.

Methodology/Principal Findings

C3^−/− mice on normal low fat diet (ND) or high fat diet (HFD) were chronically administered recombinant ASP (rASP) for 25 days via an osmotic mini-pump. While there was no effect on food intake, there was a decrease in activity, with a relative increase in adipose tissue weight on ND, and a shift in adipocyte size distribution. While rASP administration to C3^−/− mice on a ND increased insulin sensitivity, on a HFD, rASP administration had the opposite effect. Specifically, rASP administration in C3^−/− HFD mice resulted in decreased gene expression of IRS1, GLUT4, SREBF1 and NFκB in muscle, and decreased C5L2 but increased JNK, CD36, CD11c, CCR2 and NFκB gene expression in adipose tissue as well as increased secretion of proinflammatory cytokines (Rantes, KC, MCP-1, IL-6 and G-CSF). In adipose tissue, although IRS1 and GLUT4 mRNA were unchanged, insulin response was reduced.

Conclusion

The effects of chronic rASP administration are tissue and diet specific, rASP administration enhances the HFD induced inflammatory response leading to an insulin-resistant state. These results suggest that, in humans, the increased plasma ASP associated with obesity and cardiovascular disease could be an additional factor directly contributing to development of metabolic syndrome, insulin resistance and diabetes.     

CD40L deficiency attenuates diet-induced adipose tissue inflammation by impairing immune cell accumulation and production of pathogenic IgG-antibodies

Background

Adipose tissue inflammation fuels the metabolic syndrome. We recently reported that CD40L – an established marker and mediator of cardiovascular disease – induces inflammatory cytokine production in adipose cells in vitro. Here, we tested the hypothesis that CD40L deficiency modulates adipose tissue inflammation in vivo.

Methodology/Principal Findings

WT or CD40L^−/− mice consumed a high fat diet (HFD) for 20 weeks. Inflammatory cell recruitment was impaired in mice lacking CD40L as shown by a decrease of adipose tissue macrophages, B-cells, and an increase in protective T-regulatory cells. Mechanistically, CD40L-deficient mice expressed significantly lower levels of the pro-inflammatory chemokine MCP-1 both, locally in adipose tissue and systemically in plasma. Moreover, levels of pro-inflammatory IgG-antibodies against oxidized lipids were reduced in CD40L^−/− mice. Also, circulating low-density lipoproteins and insulin levels were lower in CD40L^−/− mice. However, CD40L^−/− mice consuming HFD were not protected from the onset of diet-induced obesity (DIO), insulin resistance, and hepatic steatosis, suggesting that CD40L selectively limits the inflammatory features of diet-induced obesity rather than its metabolic phenotype. Interestingly, CD40L^−/− mice consuming a low fat diet (LFD) showed both, a favorable inflammatory and metabolic phenotype characterized by diminished weight gain, improved insulin tolerance, and attenuated plasma adipokine levels.

Conclusion

We present the novel finding that CD40L deficiency limits adipose tissue inflammation in vivo. These findings identify CD40L as a potential mediator at the interface of cardiovascular and metabolic disease.     

Deficiency of Oncostatin M Receptor β (OSMRβ) Exacerbates High-fat Diet-induced Obesity and Related Metabolic Disorders in Mice

Oncostatin M (OSM) belongs to the IL-6 family of cytokines and has diverse biological effects, including the modulation of inflammatory responses. In the present study we analyzed the roles of OSM signaling in obesity and related metabolic disorders. Under a high-fat diet condition, OSM receptor β subunit-deficient (OSMRβ^−/−) mice exhibited increases in body weight and food intake compared with those observed in WT mice. In addition, adipose tissue inflammation, insulin resistance, and hepatic steatosis were more severe in OSMRβ^−/− mice than in wild-type (WT) mice. These metabolic phenotypes did not improve when OSMRβ^−/− mice were pair-fed with WT mice, suggesting that the effects of OSM signaling on these phenotypes are independent of the increases in the body weight and food intake. In the liver of OSMRβ^−/− mice, the insulin-induced phosphorylation of p70 S6 kinase remained intact, whereas insulin-induced FOXO1 phosphorylation was impaired. In addition, OSMRβ^−/− mice displayed a higher expression of genes related to de novo lipogenesis in the liver than WT mice. Furthermore, treatment of genetically obese ob/ob mice with OSM improved insulin resistance, adipose tissue inflammation, and hepatic steatosis. Intraportal administration of OSM into ob/ob mice activated STAT3 and increased the expression of long-chain acyl-CoA synthetase (ACSL) 3 and ACSL5 with decreased expression of fatty acid synthase in the liver, suggesting that OSM directly induces lipolysis and suppresses lipogenesis in the liver of obese mice. These findings suggest that defects in OSM signaling promote the deterioration of high-fat diet-induced obesity and related metabolic disorders.     

Liraglutide Increases FGF-21 Activity and Insulin Sensitivity in High Fat Diet and Adiponectin Knockdown Induced Insulin Resistance

Background

Liraglutide is a glucagon-like peptide-1 analogue that stimulates insulin secretion and improves β-cell function. However, it is not clear whether liraglutide achieves its glucose lowering effect only by its known effects or whether other as yet unknown mechanisms are involved. The aim of this study was to examine the effects of liraglutide on Fibroblast growth factor-21 (FGF-21) activity in High-fat diet (HFD) fed ApoE^−/− mice with adiponectin (Acrp30) knockdown.

Method

HFD-fed ApoE^−/− mice were treated with adenovirus vectors expressing shAcrp30 to produce insulin resistance. Hyperinsulinemic-euglycemic clamp studies were performed to evaluate insulin sensitivity of the mouse model. QRT-PCR and Western blot were used to measure the mRNA and protein expression of the target genes.

Results

The combination of HFD, ApoE deficiency, and hypoadiponectinemia resulted in an additive effect on insulin resistance. FGF-21 mRNA expressions in both liver and adipose tissues were significantly increased while FGF-21 receptor 1 (FGFR-1) and β-Klotho mRNA levels in adipose tissue, as well as FGFR-1-3 and β-Klotho mRNA levels in liver were significantly decreased in this model. Liraglutide treatment markedly improved insulin resistance and increased FGF-21 expression in liver and FGFR-3 in adipose tissue, restored β-Klotho mRNA expression in adipose tissue as well as FGFR-1-3, β-Klotho levels and phosphorylation of FGFR1 up to the levels observed in control mice in liver. Liraglutide treatment also further increased FGF-21 proteins in liver and plasma. In addition, as shown by hyperinsulinemic-euglycemic clamp, liraglutide treatment also markedly improved glucose metabolism and insulin sensitivity in these animals.

Conclusion

These findings demonstrate an additive effect of HFD, ApoE deficiency, and adiponectin knockdown on insulin resistance and unveil that the regulation of glucose metabolism and insulin sensitivity by liraglutide may be partly mediated via increased FGF-21 and its receptors action.     

Partial Deletion of eNOS Gene Causes Hyperinsulinemic State,Unbalance of Cardiac Insulin Signaling Pathways and Coronary Dysfunction Independently of High Fat Diet

Abnormalities in eNOS gene, possibly interacting with high fat diet (HFD), affect peripheral vascular function and glucose metabolism. The relative role of eNOS gene, HFD and metabolic derangement on coronary function has not been fully elucidated. We test whether eNOS gene deficiency per se or in association with HFD modulates coronary function through mechanisms involving molecular pathways related to insulin signaling. Wild type (WT), eNOS^−/− and eNOS^+/− mice were studied. WT and eNOS^+/− mice were fed with either standard or HF diet for 16 weeks and compared with standard diet fed eNOS^−/−. Glucose and insulin tolerance tests were performed during the last week of diet. Coronary resistance (CR) was measured at baseline and during infusions of acetylcholine (Ach) or sodium-nitroprusside (SNP) to evaluate endothelium-dependent or independent vasodilation, in the Langendorff isolated hearts. Cardiac expression of Akt and ERK genes as evaluation of two major insulin-regulated signaling pathways involved in the control of vascular tone were assessed by western blot. HFD-fed mice developed an overt diabetic state. Conversely, chow-fed genetically modified mice (in particular eNOS^−/−) showed a metabolic pattern characterized by normoglycemia and hyperinsulinemia with a limited degree of insulin resistance. CR was significantly higher in animals with eNOS gene deletions than in WT, independently of diet. Percent decrease in CR, during Ach infusion, was significantly lower in both eNOS^−/− and eNOS^+/− mice than in WT, independently of diet. SNP reduced CR in all groups except eNOS^−/−. The cardiac ERK1-2/Akt ratio, increased in animals with eNOS gene deletions compared with WT, independently of diet. These results suggest that the eNOS genetic deficiency, associated or not with HFD, has a relevant effect on coronary vascular function, possibly mediated by increase in blood insulin levels and unbalance in insulin-dependent signaling in coronary vessels, consistent with a shift towards a vasoconstrictive pattern.     

Kdm2a deficiency in macrophages enhances thermogenesis to protect mice against HFD-induced obesity by enhancing H3K36me2 at the Pparg locus

Kdm2a catalyzes H3K36me2 demethylation to play an intriguing epigenetic regulatory role in cell proliferation, differentiation, and apoptosis. Herein we found that myeloid-specific knockout of Kdm2a (LysM-Cre-Kdm2a^f/f, Kdm2a^−/−) promoted macrophage M2 program by reprograming metabolic homeostasis through enhancing fatty acid uptake and lipolysis. Kdm2a^−/− increased H3K36me2 levels at the Pparg locus along with augmented chromatin accessibility and Stat6 recruitment, which rendered macrophages with preferential M2 polarization. Therefore, the Kdm2a^−/− mice were highly protected from high-fat diet (HFD)-induced obesity, insulin resistance, and hepatic steatosis, and featured by the reduced accumulation of adipose tissue macrophages and repressed chronic inflammation following HFD challenge. Particularly, Kdm2a^−/− macrophages provided a microenvironment in favor of thermogenesis. Upon HFD or cold challenge, the Kdm2a^−/− mice manifested higher capacity for inducing adipose browning and beiging to promote energy expenditure. Collectively, our findings demonstrate the importance of Kdm2a-mediated H3K36 demethylation in orchestrating macrophage polarization, providing novel insight that targeting Kdm2a in macrophages could be a viable therapeutic approach against obesity and insulin resistance.Subject terms: Chronic inflammation, Histone post-translational modifications, Epigenetics, Endocrine system and metabolic diseases     

Inhibition of intestinal cholesterol absorption decreases atherosclerosis but not adipose tissue inflammation

Adipose tissue inflammation is associated with insulin resistance and increased cardiovascular disease risk in obesity. We previously showed that addition of cholesterol to a diet rich in saturated fat and refined carbohydrate significantly worsens dyslipidemia, insulin resistance, adipose tissue macrophage accumulation, systemic inflammation, and atherosclerosis in LDL receptor-deficient (Ldlr^−/−) mice. To test whether inhibition of intestinal cholesterol absorption would improve metabolic abnormalities and adipose tissue inflammation in obesity, we administered ezetimibe, a dietary and endogenous cholesterol absorption inhibitor, to Ldlr^−/− mice fed chow or high-fat, high-sucrose (HFHS) diets without or with 0.15% cholesterol (HFHS+C). Ezetimibe blunted weight gain and markedly reduced plasma lipids in the HFHS+C group. Ezetimibe had no effect on glucose homeostasis or visceral adipose tissue macrophage gene expression in the HFHS+C fed mice, although circulating inflammatory markers serum amyloid A (SSA) and serum amyloid P (SSP) levels decreased. Nevertheless, ezetimibe treatment led to a striking (>85%) reduction in atherosclerotic lesion area with reduced lesion lipid and macrophage content in the HFHS+C group. Thus, in the presence of dietary cholesterol, ezetimibe did not improve adipose tissue inflammation in obese Ldlr^−/− mice, but it led to a major reduction in atherosclerotic lesions associated with improved plasma lipids and lipoproteins.     

Kinin B1 Receptor in Adipocytes Regulates Glucose Tolerance and Predisposition to Obesity

Background

Kinins participate in the pathophysiology of obesity and type 2 diabetes by mechanisms which are not fully understood. Kinin B₁ receptor knockout mice (B₁ ^−/−) are leaner and exhibit improved insulin sensitivity.

Methodology/Principal Findings

Here we show that kinin B₁ receptors in adipocytes play a role in controlling whole body insulin action and glucose homeostasis. Adipocytes isolated from mouse white adipose tissue (WAT) constitutively express kinin B₁ receptors. In these cells, treatment with the B₁ receptor agonist des-Arg⁹-bradykinin improved insulin signaling, GLUT4 translocation, and glucose uptake. Adipocytes from B₁ ^−/− mice showed reduced GLUT4 expression and impaired glucose uptake at both basal and insulin-stimulated states. To investigate the consequences of these phenomena to whole body metabolism, we generated mice where the expression of the kinin B₁ receptor was limited to cells of the adipose tissue (aP2-B₁/B₁ ^−/−). Similarly to B₁ ^−/− mice, aP2-B₁/B₁ ^−/− mice were leaner than wild type controls. However, exclusive expression of the kinin B₁ receptor in adipose tissue completely rescued the improved systemic insulin sensitivity phenotype of B₁ ^−/− mice. Adipose tissue gene expression analysis also revealed that genes involved in insulin signaling were significantly affected by the presence of the kinin B₁ receptor in adipose tissue. In agreement, GLUT4 expression and glucose uptake were increased in fat tissue of aP2-B₁/B₁ ^−/− when compared to B₁ ^−/− mice. When subjected to high fat diet, aP2-B₁/B₁ ^−/− mice gained more weight than B₁ ^−/− littermates, becoming as obese as the wild types.

Conclusions/Significance

Thus, kinin B₁ receptor participates in the modulation of insulin action in adipocytes, contributing to systemic insulin sensitivity and predisposition to obesity.     

Osteopontin is required for the early onset of high fat diet-induced insulin resistance in mice

Background

Insulin resistance is manifested in muscle, adipose tissue, and liver and is associated with adipose tissue inflammation. The cellular components and mechanisms that regulate the onset of diet-induced insulin resistance are not clearly defined.

Methodology and Principal Findings

We initially observed osteopontin (OPN) mRNA over-expression in adipose tissue of obese, insulin resistant humans and rats which was normalized by thiazolidinedione (TZD) treatment in both species. OPN regulates inflammation and is implicated in pathogenic maladies resulting from chronic obesity. Thus, we tested the hypothesis that OPN is involved in the early development of insulin resistance using a 2–4 week high fat diet (HFD) model. OPN KO mice fed HFD for 2 weeks were completely protected from the severe skeletal muscle, liver and adipose tissue insulin resistance that developed in wild type (WT) controls, as determined by hyperinsulinemic euglycemic clamp and acute insulin-stimulation studies. Although two-week HFD did not alter body weight or plasma free fatty acids and cytokines in either strain, HFD-induced hyperleptinemia, increased adipose tissue inflammation (macrophages and cytokines), and adipocyte hypertrophy were significant in WT mice and blunted or absent in OPN KO mice. Adipose tissue OPN protein isoform expression was significantly altered in 2- and 4-week HFD-fed WT mice but total OPN protein was unchanged. OPN KO bone marrow stromal cells were more osteogenic and less adipogenic than WT cells in vitro. Interestingly, the two differentiation pathways were inversely affected by HFD in WT cells in vitro.

Conclusions

The OPN KO phenotypes we report reflect protection from insulin resistance that is associated with changes in adipocyte biology and adipose tissue inflammatory status. OPN is a key component in the development of HFD-induced insulin resistance.     

Enhanced Lipid Oxidation and Maintenance of Muscle Insulin Sensitivity Despite Glucose Intolerance in a Diet-Induced Obesity Mouse Model

Background

Diet-induced obesity is a rising health concern which can lead to the development of glucose intolerance and muscle insulin resistance and, ultimately, type II diabetes mellitus. This research investigates the associations between glucose intolerance or muscle insulin resistance and tissue specific changes during the progression of diet-induced obesity.

Methodology

C57BL/6J mice were fed a normal or high-fat diet (HFD; 60% kcal fat) for 3 or 8 weeks. Disease progression was monitored by measurements of body/tissue mass changes, glucose and insulin tolerance tests, and ex vivo glucose uptake in intact muscles. Lipid metabolism was analyzed using metabolic chambers and ex vivo palmitate assays in intact muscles. Skeletal muscle, liver and adipose tissues were analyzed for changes in inflammatory gene expression. Plasma was analyzed for insulin levels and inflammatory proteins. Histological techniques were used on muscle and liver cryosections to assess metabolic and morphological changes.

Principal Findings/Conclusions

A rapid shift in whole body metabolism towards lipids was observed with HFD. Following 3 weeks of HFD, elevated total lipid oxidation and an oxidative fiber type shift had occurred in the skeletal muscle, which we propose was responsible for delaying intramyocellular lipid accumulation and maintaining muscle’s insulin sensitivity. Glucose intolerance was present after three weeks of HFD and was associated with an enlarged adipose tissue depot, adipose tissue inflammation and excess hepatic lipids, but not hepatic inflammation. Furthermore, HFD did not significantly increase systemic or muscle inflammation after 3 or 8 weeks of HFD suggesting that early diet-induced obesity does not cause inflammation throughout the whole body. Overall these findings indicate skeletal muscle did not contribute to the development of HFD-induced impairments in whole-body glucose tolerance following 3 weeks of HFD.     

No Evidence for a Role of Adipose Tissue-Derived Serum Amyloid A in the Development of Insulin Resistance or Obesity-Related Inflammation in hSAA1+/? Transgenic Mice

Obesity is associated with a low-grade inflammation including moderately increased serum levels of the acute phase protein serum amyloid A (SAA). In obesity, SAA is mainly produced from adipose tissue and serum levels of SAA are associated with insulin resistance. SAA has been described as a chemoattractant for inflammatory cells and adipose tissue from obese individuals contains increased numbers of macrophages. However, whether adipose tissue-derived SAA can have a direct impact on macrophage infiltration in adipose tissue or the development of insulin resistance is unknown. The aim of this study was to investigate the effects of adipose tissue-derived SAA1 on the development of insulin resistance and obesity-related inflammation. We have previously established a transgenic mouse model expressing human SAA1 in the adipose tissue. For this report, hSAA1^+/− transgenic mice and wild type mice were fed with a high fat diet or normal chow. Effects of hSAA1 on glucose metabolism were assessed using an oral glucose tolerance test. Real-time PCR was used to measure the mRNA levels of macrophage markers and genes related to insulin sensitivity in adipose tissue. Cytokines during inflammation were analyzed using a Proinflammatory 7-plex Assay. We found similar insulin and glucose levels in hSAA1 mice and wt controls during an oral glucose tolerance test and no decrease in mRNA levels of genes related to insulin sensitivity in adipose tissue in neither male nor female hSAA1 animals. Furthermore, serum levels of proinflammatory cytokines and mRNA levels of macrophage markers in adipose tissue were not increased in hSAA1 mice. Hence, in this model we find no evidence that adipose tissue-derived hSAA1 influences the development of insulin resistance or obesity-related inflammation.     

Increased Adiposity in Annexin A1-Deficient Mice

Production of Annexin A1 (ANXA1), a protein that mediates the anti-inflammatory action of glucocorticoids, is altered in obesity, but its role in modulation of adiposity has not yet been investigated. The objective of this study was to investigate modulation of ANXA1 in adipose tissue in murine models of obesity and to study the involvement of ANXA1 in diet-induced obesity in mice. Significant induction of ANXA1 mRNA was observed in adipose tissue of both C57BL6 and Balb/c mice with high fat diet (HFD)-induced obesity versus mice on chow diet. Upregulation of ANXA1 mRNA was independent of leptin or IL-6, as demonstrated by use of leptin-deficient ob/ob mice and IL-6 KO mice. Compared to WT mice, female Balb/c ANXA1 KO mice on HFD had increased adiposity, as indicated by significantly elevated body weight, fat mass, leptin levels, and adipocyte size. Whereas Balb/c WT mice upregulated expression of enzymes involved in the lipolytic pathway in response to HFD, this response was absent in ANXA1 KO mice. A significant increase in fasting glucose and insulin levels as well as development of insulin resistance was observed in ANXA1 KO mice on HFD compared to WT mice. Elevated plasma corticosterone levels and blunted downregulation of 11-beta hydroxysteroid dehydrogenase type 1 in adipose tissue was observed in ANXA1 KO mice compared to diet-matched WT mice. However, no differences between WT and KO mice on either chow or HFD were observed in expression of markers of adipose tissue inflammation.These data indicate that ANXA1 is an important modulator of adiposity in mice, with female ANXA1 KO mice on Balb/c background being more susceptible to weight gain and diet-induced insulin resistance compared to WT mice, without significant changes in inflammation.     

Exercise Protects against Diet-Induced Insulin Resistance through Downregulation of Protein Kinase Cβ in Mice

Physical exercise is an important and effective therapy for diabetes. However, its underlying mechanism is not fully understood. Protein kinase Cβ (PKCβ) has been suggested to be involved in the pathogenesis of obesity and insulin resistance, but the role of PKCβ in exercise-induced improvements in insulin resistance is completely unknown. In this study, we evaluated the involvement of PKCβ in exercise-attenuated insulin resistance in high-fat diet (HFD)-fed mice. PKCβ^-/- and wild-type mice were fed a HFD with or without exercise training. PKC protein expression, body and tissue weight change, glucose and insulin tolerance, metabolic rate, mitochondria size and number, adipose inflammation, and AKT activation were determined to evaluate insulin sensitivity and metabolic changes after intervention. PKCβ expression decreased in both skeletal muscle and liver tissue after exercise. Exercise and PKCβ deficiency can alleviate HFD-induced insulin resistance, as evidenced by improved insulin tolerance. In addition, fat accumulation and mitochondrial dysfunction induced by HFD were also ameliorated by both exercise and PKCβ deficiency. On the other hand, exercise had little effect on PKCβ^-/- mice. Further, our data indicated improved activation of AKT, the downstream signal molecule of insulin, in skeletal muscle and liver of exercised mice, whereas PKCβ deficiency blunted the difference between sedentary and exercised mice. These results suggest that downregulation of PKCβ contributes to exercise-induced improvement of insulin resistance in HFD-fed mice.     

A high-fat diet catalyzes progression to hyperglycemia in mice with selective impairment of insulin action in Glut4-expressing tissues

Insulin resistance impairs postprandial glucose uptake through glucose transporter type 4 (GLUT4) and is the primary defect preceding type 2 diabetes. We previously generated an insulin-resistant mouse model with human GLUT4 promoter-driven insulin receptor knockout (GIRKO) in the muscle, adipose, and neuronal subpopulations. However, the rate of diabetes in GIRKO mice remained low prior to 6 months of age on normal chow diet (NCD), suggesting that additional factors/mechanisms are responsible for adverse metabolic effects driving the ultimate progression of overt diabetes. In this study, we characterized the metabolic phenotypes of the adult GIRKO mice acutely switched to high-fat diet (HFD) feeding in order to identify additional metabolic challenges required for disease progression. Distinct from other diet-induced obesity (DIO) and genetic models (e.g., db/db mice), GIRKO mice remained leaner on HFD feeding, but developed other cardinal features of insulin resistance syndrome. GIRKO mice rapidly developed hyperglycemia despite compensatory increases in β-cell mass and hyperinsulinemia. Furthermore, GIRKO mice also had impaired oral glucose tolerance and a limited glucose-lowering benefit from exendin-4, suggesting that the blunted incretin effect contributed to hyperglycemia. Secondly, GIRKO mice manifested severe dyslipidemia while on HFD due to elevated hepatic lipid secretion, serum triglyceride concentration, and lipid droplet accumulation in hepatocytes. Thirdly, GIRKO mice on HFD had increased inflammatory cues in the gut, which were associated with the HFD-induced microbiome alterations and increased serum lipopolysaccharide (LPS). In conclusion, our studies identified important gene/diet interactions contributing to diabetes progression, which might be leveraged to develop more efficacious therapies.