Mice Lacking C1q Are Protected from High Fat Diet-induced Hepatic Insulin Resistance and Impaired Glucose Homeostasis

Complement activation is implicated in the development of obesity and insulin resistance, and loss of signaling by the anaphylatoxin C3a prevents obesity-induced insulin resistance in mice. Here we have identified C1q in the classical pathway as required for activation of complement in response to high fat diets. After 8 weeks of high fat diet, wild-type mice became obese and developed glucose intolerance. This was associated with increased apoptotic cell death and accumulation of complement activation products (C3b/iC3b/C3c) in liver and adipose tissue. Previous studies have shown that high fat diet-induced apoptosis is dependent on Bid; here we report that Bid-mediated apoptosis was required for complement activation in adipose and liver. Although C1qa deficiency had no effect on high fat diet-induced apoptosis, accumulation of complement activation products and the metabolic complications of high fat diet-induced obesity were dependent on C1q. When wild-type mice were fed a high fat diet for only 3 days, hepatic insulin resistance was associated with the accumulation of C3b/iC3b/C3c in the liver. Mice deficient in C3a receptor were protected against this early high fat diet-induced hepatic insulin resistance, whereas mice deficient in the negative complement regulator CD55/DAF were more sensitive to the high fat diet. C1qa^−/− mice were also protected from high fat diet-induced hepatic insulin resistance and complement activation. Evidence of complement activation was also detected in adipose tissue of obese women compared with lean women. Together, these studies reveal an important role for C1q in the classical pathway of complement activation in the development of high fat diet-induced insulin resistance.     

Esculentin-2CHa-Related Peptides Modulate Islet Cell Function and Improve Glucose Tolerance in Mice with Diet-Induced Obesity and Insulin Resistance

The frog skin host-defense peptide esculentin-2CHa (GFSSIFRGVA¹⁰KFASKGLGK D²⁰LAKLGVDLVA³⁰CKISKQC) displays antimicrobial, antitumor, and immunomodulatory properties. This study investigated the antidiabetic actions of the peptide and selected analogues. Esculentin-2CHa stimulated insulin secretion from rat BRIN-BD11 clonal pancreatic β-cells at concentrations greater than 0.3 nM without cytotoxicity by a mechanism involving membrane depolarization and increase of intracellular Ca²⁺. Insulinotropic activity was attenuated by activation of K_ATP channels, inhibition of voltage-dependent Ca²⁺ channels and chelation of extracellular Ca²⁺. The [L21K], [L24K], [D20K, D27K] and [C31S,C37S] analogues were more potent but less effective than esculentin-2CHa whereas the [L28K] and [C31K] analogues were both more potent and produced a significantly (P < 0.001) greater maximum response. Acute administration of [L28K]esculentin-2CHa (75 nmol/kg body weight) to high fat fed mice with obesity and insulin resistance enhanced glucose tolerance and insulin secretion. Twice-daily administration of this dose of [L28K]esculentin-2CHa for 28 days had no significant effect on body weight, food intake, indirect calorimetry or body composition. However, mice exhibited decreased non-fasting plasma glucose (P < 0.05), increased non-fasting plasma insulin (P < 0.05) as well as improved glucose tolerance and insulin secretion (P < 0.01) following both oral and intraperitoneal glucose loads. Impaired responses of isolated islets from high fat fed mice to established insulin secretagogues were restored by [L28K]esculentin-2CHa treatment. Peptide treatment was accompanied by significantly lower plasma and pancreatic glucagon levels and normalization of α-cell mass. Circulating triglyceride concentrations were decreased but plasma cholesterol and LDL concentrations were not significantly affected. The data encourage further investigation of the potential of esculentin-2CHa related peptides for treatment of patients with type 2 diabetes.     

Preserved Pancreatic β-Cell Development and Function in Mice Lacking the Insulin Receptor-Related Receptor

Receptors of the insulin/insulinlike growth factor (IGF) family have been implicated in the regulation of pancreatic beta-cell growth and insulin secretion. The insulin receptor-related receptor (IRR) is an orphan receptor of the insulin receptor gene (Ir) subfamily. It is expressed at considerably higher levels in beta cells than either insulin or IGF-1 receptors, and it has been shown to engage in heterodimer formation with insulin or IGF-1 receptors. To address whether IRR plays a physiologic role in beta-cell development and regulation of insulin secretion, we have characterized mice lacking IRR and generated a combined knockout of Ir and Irr. We report that islet morphology, beta-cell mass, and secretory function are not affected in IRR-deficient mice. Moreover, lack of IRR does not impair compensatory beta-cell hyperplasia in insulin-resistant Ir(+/-) mice, nor does it affect beta-cell development and function in Ir(-/-) mice. We conclude that glucose-stimulated insulin secretion and embryonic beta-cell development occur normally in mice lacking Irr.     

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.     

脓毒症小鼠心肌损伤与中性粒细胞浸润的关系

目的:探讨脓毒症小鼠心肌损伤与中性粒细胞浸润的关系。方法:复制脓毒症动物模型,分对照组、假手术组、造模组,并设不同的时间点(2h、4h、8h、12h)。采用双抗夹心酶联免疫法(ELISA)检测血清肌钙蛋白(IcTnI),并测心肌组织髓过氧化物酶(MPO)活性。结果:正常组与假手术组各时间点血清cTnI、心肌组织MPO活性均无显著性差异。造模组心肌组织MPO活性较正常组和假手术组均有明显升高(P<0.05),且随着时间进展而增加;造模组血清cTnI浓度随着时间进展而增加,2小时与正常组及假手术组无显著性差异(P>0.05),4小时后显著高于正常组和假手术组(P<0.05);血清cTnI浓度与心肌组织MPO活性呈显著正相关(r=0.700,P=0.000)。结论:脓毒症心肌损伤时,心肌组织存在中性粒细胞浸润,中性粒细胞浸润程度与心肌损伤显著正相关。     

Mutations in Mll2, an H3K4 Methyltransferase,Result in Insulin Resistance and Impaired Glucose Tolerance in Mice

We employed a random mutagenesis approach to identify novel monogenic determinants of type 2 diabetes. Here we show that haplo-insufficiency of the histone methyltransferase myeloid-lineage leukemia (Mll2/Wbp7) gene causes type 2 diabetes in the mouse. We have shown that mice heterozygous for two separate mutations in the SET domain of Mll2 or heterozygous Mll2 knockout mice were hyperglycaemic, hyperinsulinaemic and developed non-alcoholic fatty liver disease. Consistent with previous Mll2 knockout studies, mice homozygous for either ENU mutation (or compound heterozygotes) died during embryonic development at 9.5–14.5 days post coitum. Heterozygous deletion of Mll2 induced in the adult mouse results in a normal phenotype suggesting that changes in chromatin methylation during development result in the adult phenotype. Mll2 has been shown to regulate a small subset of genes, a number of which Neurod1, Enpp1, Slc27a2, and Plcxd1 are downregulated in adult mutant mice. Our results demonstrate that histone H3K4 methyltransferase Mll2 is a component of the genetic regulation necessary for glucose homeostasis, resulting in a specific disease pattern linking chromatin modification with causes and progression of type 2 diabetes, providing a basis for its further understanding at the molecular level.     

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.     

Role of NKG2D in Obesity-Induced Adipose Tissue Inflammation and Insulin Resistance

The early events that initiate inflammation in the adipose tissue during obesity are not well defined. It is unclear whether the recruitment of CD8 T cells to the adipose tissue during onset of obesity occurs through antigen-dependent or -independent processes. We have previously shown that interaction between NKG2D (natural-killer group 2, member D) and its ligand Rae-1ε is sufficient to recruit cytotoxic T lymphocytes to the pancreas and induce insulitis. Here, we tested whether NKG2D–NKG2D ligand interaction is also involved in obesity-induced adipose tissue inflammation and insulin resistance. We observed a significant induction of NKG2D ligand expression in the adipose tissue of obese mice, especially during the early stages of obesity. However, mice lacking NKG2D developed similar levels of insulin resistance and adipose tissue inflammation compared to control mice when placed on a high-fat diet. Moreover, overexpression of Rae-1ε in the adipose tissue did not increase immune cell infiltration to the adipose tissue either in the setting of a normal or high-fat diet. These results indicate that, unlike in the pancreas, NKG2D–NKG2D ligand interaction does not play a critical role in obesity-induced inflammation in the adipose tissue.     

Hepatic Circadian-Clock System Altered by Insulin Resistance,Diabetes and Insulin Sensitizer in Mice

Circadian rhythms are intrinsic rhythms that are coordinated with the rotation of the Earth and are also generated by a set of circadian-clock genes at the intracellular level. Growing evidence suggests a strong link between circadian rhythms and energy metabolism; however, the fundamental mechanisms remain unclear. In the present study, neonatal streptozotocin (STZ)-treated mice were used to model the molecular and physiological progress from insulin resistance to diabetes. Two-day-old male C57BL/6 mice received a single injection of STZ and were tested for non-obese, hyperglycemic and hyperinsulinemic conditions in the early stage, insulin resistance in the middle stage, and diabetes in the late stage. Gene expression levels of the hepatic circadian-clock system were examined by real-time quantitative PCR. Most of the components of the hepatic circadian-clock gene expression system, such as the mRNAs of Bmal1 (brain and muscle Arnt-like protein-1), Per2 (period 2) and Cry1 (cryptochrome 1), were elevated, and circadian patterns were retained in the early and middle stages of insulin-resistant conditions. The insulin sensitizer, rosiglitazone, returns the physiological and molecular changes associated with the diabetic phenotype to normal levels through peroxisome proliferator-activated receptor γ (PPARγ) rather than PPARα. Early and chronic treatment with rosiglitazone has been shown to be effective to counter the diabetic condition. Over time, this effect acts to attenuate the increased gene expression levels of the hepatic circadian-clock system and delay the severity of diabetic conditions. Together, these results support an essential role for the hepatic circadian-clock system in the coordinated regulation and/or response of metabolic pathways.     

Crif1 Deficiency Reduces Adipose OXPHOS Capacity and Triggers Inflammation and Insulin Resistance in Mice

Impaired mitochondrial oxidative phosphorylation (OXPHOS) has been proposed as an etiological mechanism underlying insulin resistance. However, the initiating organ of OXPHOS dysfunction during the development of systemic insulin resistance has yet to be identified. To determine whether adipose OXPHOS deficiency plays an etiological role in systemic insulin resistance, the metabolic phenotype of mice with OXPHOS–deficient adipose tissue was examined. Crif1 is a protein required for the intramitochondrial production of mtDNA–encoded OXPHOS subunits; therefore, Crif1 haploinsufficient deficiency in mice results in a mild, but specific, failure of OXPHOS capacity in vivo. Although adipose-specific Crif1-haploinsufficient mice showed normal growth and development, they became insulin-resistant. Crif1-silenced adipocytes showed higher expression of chemokines, the expression of which is dependent upon stress kinases and antioxidant. Accordingly, examination of adipose tissue from Crif1-haploinsufficient mice revealed increased secretion of MCP1 and TNFα, as well as marked infiltration by macrophages. These findings indicate that the OXPHOS status of adipose tissue determines its metabolic and inflammatory responses, and may cause systemic inflammation and insulin resistance.     

Myocardial Mitochondrial and Contractile Function Are Preserved in Mice Lacking Adiponectin

Adiponectin deficiency leads to increased myocardial infarct size following ischemia reperfusion and to exaggerated cardiac hypertrophy following pressure overload, entities that are causally linked to mitochondrial dysfunction. In skeletal muscle, lack of adiponectin results in impaired mitochondrial function. Thus, it was our objective to investigate whether adiponectin deficiency impairs mitochondrial energetics in the heart. At 8 weeks of age, heart weight-to-body weight ratios were not different between adiponectin knockout (ADQ^-/-) mice and wildtypes (WT). In isolated working hearts, cardiac output, aortic developed pressure and cardiac power were preserved in ADQ^-/- mice. Rates of fatty acid oxidation, glucose oxidation and glycolysis were unchanged between groups. While myocardial oxygen consumption was slightly reduced (-24%) in ADQ^-/- mice in isolated working hearts, rates of maximal ADP-stimulated mitochondrial oxygen consumption and ATP synthesis in saponin-permeabilized cardiac fibers were preserved in ADQ^-/- mice with glutamate, pyruvate or palmitoyl-carnitine as a substrate. In addition, enzymatic activity of respiratory complexes I and II was unchanged between groups. Phosphorylation of AMP-activated protein kinase and SIRT1 activity were not decreased, expression and acetylation of PGC-1α were unchanged, and mitochondrial content of OXPHOS subunits was not decreased in ADQ^-/- mice. Finally, increasing energy demands due to prolonged subcutaneous infusion of isoproterenol did not differentially affect cardiac contractility or mitochondrial function in ADQ^-/- mice compared to WT. Thus, mitochondrial and contractile function are preserved in hearts of mice lacking adiponectin, suggesting that adiponectin may be expendable in the regulation of mitochondrial energetics and contractile function in the heart under non-pathological conditions.     

Increased Adiposity,Dysregulated Glucose Metabolism and Systemic Inflammation in Galectin-3 KO Mice

Obesity and type 2 diabetes are associated with increased production of Galectin-3 (Gal-3), a protein that modulates inflammation and clearance of glucose adducts. We used Lean and Diet-induced Obese (DIO) WT and Gal-3 KO mice to investigate the role of Gal-3 in modulation of adiposity, glucose metabolism and inflammation. Deficiency of Gal-3 lead to age-dependent development of excess adiposity and systemic inflammation, as indicated by elevated production of acute-phase proteins, number of circulating pro-inflammatory Ly6C^high monocytes and development of neutrophilia, microcytic anemia and thrombocytosis in 20-week-old Lean and DIO male Gal-3 KO mice. This was associated with impaired fasting glucose, heightened response to a glucose tolerance test and reduced adipose tissue expression of adiponectin, Gal-12, ATGL and PPARγ, in the presence of maintained insulin sensitivity and hepatic expression of gluconeogenic enzymes in 20-week-old Gal-3 KO mice compared to their diet-matched WT controls. Expression of PGC-1α and FGF-21 in the liver of Lean Gal-3 KO mice was comparable to that observed in DIO animals. Impaired fasting glucose and altered responsiveness to a glucose load preceded development of excess adiposity and systemic inflammation, as demonstrated in 12-week-old Gal-3 KO mice. Finally, a role for the microflora in mediating the fasting hyperglycemia, but not the excessive response to a glucose load, of 12-week-old Gal-3 KO mice was demonstrated by administration of antibiotics. In conclusion, Gal-3 is an important modulator of glucose metabolism, adiposity and inflammation.     

Relationships Among Obesity,Inflammation, and Insulin Resistance in African Americans and West Africans

Several research studies in different populations indicate that inflammation may be the link between obesity and insulin resistance (IR). However, this relationship has not been adequately explored among African Americans, an ethnic group with disproportionately high rates of obesity and IR. In this study, we conducted a comparative study of the relationship among adiposity, inflammation, and IR in African Americans and West Africans, the ancestral source population for African Americans. The associations between obesity markers (BMI and waist‐to‐hip ratio (WHR)), inflammatory markers (high‐sensitivity C‐reactive protein (hsCRP), haptoglobin, interleukin (IL)‐6, and tumor necrosis factor (TNF)‐α), and IR (homeostasis model assessment of insulin resistance (HOMA_IR)) were evaluated in 247 West Africans and 315 African Americans. In average, African Americans were heavier than the West Africans (by an average of 1.6 BMI units for women and 3 BMI units for men). Plasma hsCRP, haptoglobin, and IL‐6 (but not TNF‐α level) were higher in African Americans than in West Africans. In both populations, BMI was associated with markers of inflammation and with HOMA_IR, and these associations remained significant after adjusting for sex and age. However, the pattern of associations between measured inflammatory markers and IR was different between the two groups. In West Africans, hsCRP was the only inflammatory marker associated with IR. In contrast, hsCRP, haptoglobin, and IL‐6 were all associated with IR in African Americans. Interestingly, none of the associations between markers of inflammation and IR remained significant after adjusting for BMI. This finding suggests that in African Americans, the relationship between inflammatory markers and IR is mediated by adiposity.     

Postprandial Hypertriglyceridemia Predicts Development of Insulin Resistance Glucose Intolerance and Type 2 Diabetes

Insulin resistance (IR) and type 2 diabetes mellitus (T2DM) have been found to be associated with postprandial hypertriglyceridemia (PPHTg). However, whether PPHTg can cause IR and diabetes is not clear. We therefore investigated the role of PPHTg in development of T2DM in rat model of T2DM. 96 male Wistar rats were randomized into four groups (24 rats each). Control Group A, high sucrose diet (HSD) Group B, HSD+Pioglitazone (10mg/kg/day) Group C and HSD+Atorvastatin (20mg/kg/day) Group D. Fat and glucose tolerance tests were done at regular intervals in all groups besides insulin and body weight measurement. At 26 weeks, low dose streptozotocin (15mg/kg,i.p.) was given to half of the rats. All rats were followed up till 48 weeks. PPHTg developed as early as week 2 in Group B and stabilized by week 14. Group B displayed highest PPHTg compared to other groups. Atorvastatin treatment (Group D) abolished PPHTg which became comparable to controls, pioglitazone treatment partially blunted PPHTg resulting in intermediate PPHTg. Group B with highest PPHTg showed highest subsequent IR, glucose intolerance (GI) and highest incidence of prediabetes at week 26 and diabetes at week 34 and 46 compared to other groups. Group D rats displayed lower IR, GI, low incidence of prediabetes and diabetes at these time points compared to Groups B and C. ROC analysis showed that triglyceride area under the curve of each time point significantly predicts the risk of diabetes. Present study provides the evidence that PPHTg predicts the development of IR, GI and T2DM in rat model of diet induced T2DM.     

Insulin Resistance in Relation to Lipids and Inflammation in Type-2 Diabetic Patients and Non-Diabetic People

BackgroundWe demonstrated in experimental studies that hypercholesterolaemia enhances the proliferation of haematopoietic stem cells and the subsequent differentiation to neutrophils, whereas HDL-cholesterol inhibits these processes. To translate our experimental findings to clinical practice, we investigated in Chinese type-2 diabetic patients and in Flemish non-diabetic people the independent and joint associations of insulin resistance with markers of dyslipidaemia and inflammation, while looking for consistency between ethnicities and across the spectrum of insulin resistance.MethodsWe studied 798 Chinese patients with type-2 diabetes (53.6% women; mean age, 60.6 years) admitted to a tertiary referral centre and 1060 white Flemish (50.5%; 51.1 years) randomly recruited in Northern Belgium. Fasting insulin resistance (HOMA-IR) was derived from C-peptide in Chinese and from insulin in Flemish using the Homeostasis Model of Assessment algorithm. In multivariable-adjusted analyses, HOMA-IR was regressed on triglycerides, HDL-cholesterol and neutrophil count.ResultsIn Chinese patients, the percentage changes in HOMA-IR associated with triglycerides, HDL-cholesterol and neutrophils (per 1-SD increment) amounted to 8.1 (95% confidence interval, 3.0 to 13.4; p = 0.0015), -8.7 (-13.0 to -4.2; p = 0.0002) and 5.6 (1.0 to 10.4; p = 0.017). In non-diabetic Flemish, the corresponding estimates were 11.7 (8.3 to 15.1; p<0.0001), -1.7 (-4.6 to 1.4; p = 0.28) and 3.3% (0.5 to 6.3; p = 0.022), respectively. None of the interaction terms between the three explanatory variables reached significance in Chinese or Flemish (p≥0.10).ConclusionsInsulin resistance increases with the serum level of triglycerides and the blood neutrophil count, but decreases with serum HDL-cholesterol concentration. These associations were consistent in Chinese type-2 diabetic patients and non-diabetic Flemish people and were independent from one another. The clinical implications are that future studies should focus on intervening with serum triglyceride and HDL-cholesterol levels or controlling inflammation as a way to prevent or treat insulin resistance.