Growth arrest-specific 6 modulates adiponectin expression and insulin resistance in adipose tissue

Aims/IntroductionObesity is characterized by disturbed adipocytokine expression and insulin resistance in adipocytes. Growth arrest‐specific 6 (GAS6) is a gene encoding the Gas6 protein, which is expressed in fibroblasts, and its related signaling might be associated with adipose tissue inflammation, glucose intolerance and insulin resistance. The aim of this study was to investigate the associations among Gas6, adipocytokines and insulin resistance in adipocytes.Materials and MethodsMature Simpson Golabi Behmel Syndrome adipocytes were treated with high levels of insulin to mimic insulin resistance, and were examined for the expressions of Gas6, cytokines and adipocytokines from preadipocytes in differentiation. In an animal study, high‐fat diet‐induced obese mice were used to verify the Gas6 expression in vitro.ResultsDuring the differentiation of adipocytes, the expression of Gas6 gradually decreased, and was obviously downregulated with adipocyte inflammation and insulin resistance. Gas6 levels were found to be in proportion to the expression of adiponectin, which has been regarded as closely relevant to improved insulin sensitivity after metformin treatment. Similar results were also confirmed in the animal study.ConclusionsOur results suggest that Gas6 might modulate the expression of adiponectin, and might therefore be associated with insulin resistance in adipose tissues.     

Association of adiponectin and resistin with adipose tissue compartments, insulin resistance and dyslipidaemia

AIM: In this study, we investigated the association of plasma adiponectin and resistin concentrations with adipose tissue compartments in 41 free-living men with a wide range of body mass index (22-35 kg/m(2)). METHODS: Using enzyme immunoassays, plasma adiponectin and resistin were measured. Intraperitoneal, retroperitoneal, subcutaneous abdominal and posterior subcutaneous abdominal adipose tissue masses (IPATM, RPATM, SAATM and PSAATM, respectively) were determined using magnetic resonance imaging. Total adipose tissue mass (TATM) was measured using bioelectrical impedance. Insulin resistance was estimated with the help of homeostasis model assessment (HOMA) score. RESULTS: In univariate regression, plasma adiponectin levels were inversely related to IPATM (r = -0.389, p < 0.05), SAATM (r = -0.500, p < 0.001), PSAATM (r = -0.502, p < 0.001), anterior SAATM (r = -0.422, p < 0.01) and TATM (r = -0.421, p < 0.01). In multiple regression models, adiponectin was chiefly correlated with PSAATM. Plasma adiponectin concentrations were also inversely correlated with HOMA score (r = -0.540, p < 0.001) and triglyceride (r = -0.632, p < 0.001), and positively correlated with high-density lipoprotein cholesterol (r = 0.508, p < 0.001). There were no significant correlations between resistin levels and adipose tissue masses, insulin resistance or dyslipidaemia. CONCLUSIONS: In men, total body fat is significantly correlated with plasma adiponectin, but not with plasma resistin levels. Low plasma adiponectin levels appear to be chiefly determined by the accumulation of posterior subcutaneous abdominal fat mass, as opposed to intra-abdominal fat, and are strongly predictive of insulin resistance and dyslipidaemia.     

Impact of HDL on adipose tissue metabolism and adiponectin expression

ObjectiveThe objective of the current study was to investigate the hypothesis that high-density lipoprotein (HDL) influences adipocyte metabolism and adiponectin expression. Therefore, HDL was increased in vivo via apolipoprotein (apo) A-I gene transfer and in vitro via supplementation of HDL to partly differentiated adipocytes, in the presence or absence of lipopolysaccharide (LPS), known to decrease HDL cholesterol and adiponectin levels in vivo.Methods and resultsApo A-I transfer resulted in a significant increase of HDL cholesterol in control and LPS-injected C57BL/6 mice, which was paralleled by an increase in plasma adiponectin levels and adiponectin expression in abdominal fat. Triglyceride and free fatty acids levels after LPS administration were 2.2-fold (p < 0.05) and 1.3-fold (p < 0.05) lower, respectively, in Ad.hapoA-I-LPS than in Ad.Null-LPS mice. In parallel, the LPS-induced mRNA expression of hormone sensitive lipase was 3.5-fold (p = 0.05) decreased in the Ad.hapoA-I-LPS group. On the other hand, apo A-I transfer abrogated the LPS-mediated reduction in lipin-1 and CD36 mRNA expression by 8.2-fold (p < 0.05) and 18-fold (p < 0.05), respectively. Concomitantly, the phosphorylation state of Akt was 2.0-fold (p < 0.05) increased in the Ad.hapoA-I-LPS compared to the Ad.Null-LPS group. Pre-incubation of partly differentiated adipocytes with HDL (50 μg protein/ml) increased adiponectin expression by 1.5-fold under basal conditions (p < 0.05) and could abrogate LPS-induced down-regulation of adiponectin, both in a phosphatidylinositol-3-kinase-dependent manner.ConclusionsHDL affects adipocyte metabolism and adiponectin expression.     

Role of insulin receptor substrate-1 serine 307 phosphorylation and adiponectin in adipose tissue insulin resistance in late pregnancy

Insulin resistance is a hallmark of late pregnancy both in human and rat. Adipose tissue is one of the tissues that most actively contributes to this reduced insulin sensitivity. The aim of the present study was to characterize the molecular mechanisms of insulin resistance in adipose tissue at late pregnancy. To this end, we analyzed the insulin signaling cascade in lumbar adipose tissue of nonpregnant and pregnant (d 20) rats both under basal and insulin-stimulated conditions. We found that the levels of relevant signaling proteins, such as insulin receptor (IR), IR substrate-1 (IRS-1), phosphatidylinositol 3-kinase, 3-phosphoinositide-dependent kinase-1, ERK1/2, and phosphatase and tensin homolog (PTEN) did not change at late pregnancy. However, insulin-stimulated tyrosine phosphorylation of both IR and IRS-1 were significantly decreased, coincident with decreased IRS-1/p85 association and impaired phosphorylation of AKR mouse thymoma viral protooncogene (Akt) and ERK1/2. This impaired activation of IRS-1 occurred together with an increase of IRS-1 phosphorylation at serine 307 and a decrease in adiponectin levels. To corroborate the role of IRS-1 in adipose tissue insulin resistance during pregnancy, we treated pregnant rats with the antidiabetic drug englitazone. Englitazone improved glucose tolerance, and this pharmacological reversal of insulin resistance was paralleled by an increase of adiponectin levels in adipose tissue as well as by a reduction of IRS-1 serine phosphorylation. Furthermore, the impaired insulin-stimulated tyrosine phosphorylation of IRS-1 in adipose tissue of pregnant animals could be restored ex vivo by treating isolated adipocytes with adiponectin. Together, our findings support a role for adiponectin and serine phosphorylation of IRS-1 in the modulation of insulin resistance in adipose tissue at late pregnancy.     

Chronic ethanol-induced insulin resistance is associated with macrophage infiltration into adipose tissue and altered expression of adipocytokines

BACKGROUND: Chronic ethanol consumption disrupts glucose homeostasis and is associated with the development of insulin resistance. While adipose tissue and skeletal muscle are the two major organs utilizing glucose in response to insulin, the relative contribution of these two tissues to impaired glucose homeostasis during chronic ethanol feeding is not known. As other models of insulin resistance, such as obesity, are characterized by an infiltration of macrophages into adipose tissue, as well as changes in the expression of adipocytokines that play a central role in the regulation of insulin sensitivity, we hypothesized that chronic ethanol-induced insulin resistance would be associated with increased macrophage infiltration into adipose tissue and changes in the expression of adipocytokines by adipose tissue. METHODS: Male Wistar rats were fed a liquid diet containing ethanol as 36% of calories or pair-fed a control diet for 4 weeks. The effects of chronic ethanol feeding on insulin-stimulated glucose utilization were studied using the hyperinsulinemic-euglycemic clamp technique, coupled with the use of isotopic tracers. Further, macrophage infiltration into adipose tissue and expression of adipocytokines were also assessed after chronic ethanol feeding. RESULTS: Hyperinsulinemic-euglycemic clamp studies revealed that chronic ethanol feeding to rats decreased whole-body glucose utilization and decreased insulin-mediated suppression of hepatic glucose production. Chronic ethanol feeding decreased glucose uptake in epididymal, subcutaneous, and omental adipose tissue during the hyperinsulinemic-euglycemic clamp, but had no effect on glucose disposal in skeletal muscle. Chronic ethanol feeding increased the infiltration of macrophages into epididymal adipose tissue and changed the expression of mRNA for adipocytokines: expression of mRNA for monocyte chemoattractant protein 1, tumor necrosis factor alpha, and interleukin-6 were increased, while expression of mRNA for retinol binding protein 4 and adiponectin were decreased in epididymal adipose tissue. CONCLUSIONS: These data demonstrate that chronic ethanol feeding results in the development of insulin resistance, associated with impaired insulin-mediated suppression of hepatic glucose production and decreased insulin-stimulated glucose uptake into adipose tissue. Chronic ethanol-induced insulin resistance was associated with increased macrophage infiltration into adipose tissue, as well as changes in the expression of adipocytokines by adipose tissue.     

肥胖小鼠脂肪组织缺氧对脂联素表达的转录抑制作用

目的观察缺氧对脂肪细胞脂联素mRNA和蛋白表达的影响,探讨肥胖小鼠脂肪组织缺氧导致脂肪组织脂联素表达下降的机制。方法采用实时定量聚合酶链反应（qRT—PCR）和蛋白免疫印迹法（Western blotting）检测遗传型肥胖小鼠（ob／ob,12周）和高脂饮食肥胖小鼠（HFD,53周）的附睾旁脂肪中脂联素mRNA和蛋白的表达;用小鼠3T3-L1脂肪细胞系为模型,采用RT—PCR和荧光素酶报告基因方法检测缺氧处理后脂联素和过氧化物酶体增殖物激活受体（PPAR）-mRNA的表达和稳定性、脂联素启动子的活性;用Western blotting和荧光素酶报告基因检测缺氧对PPAR-γ在核蛋白中集聚以及PPAR-γ转录因子活性的影响。组间数据比较采用t检验。结果（1）缺氧时两种肥胖小鼠的脂肪组织中脂联素mRNA和蛋白的表达均显著下降（P〈0．01）;333-L1脂肪细胞系在缺氧8h和24h后,脂联素mRNA表达量分别下降至0．65±0．05和0．29±0．05,较对照组（1．00±0．04）明显降低,差异有统计学意义（t=11．548、24．893,均P〈0．01）,但缺氧对脂联素mRNA的稳定性并没有影响;荧光素酶报告基因方法表明,脂联素启动子的活性受到缺氧的抑制。（2）在两种肥胖小鼠的脂肪组织中,PPAR-γmRNA和蛋白的表达均明显下降（P〈0．01）;小鼠333-L1脂肪细胞系在缺氧8h和24h后,PPAR- γmRNA的表达量分别下降至0．72±0．09和0．54±0．07,与对照组（1．00±0．09）相比,差异有统计学意义（t：5．134、9．876,均P〈0．01）;PPAR一1蛋白的核转位以及PPAR一^y转录因子活性也受到缺氧的抑制。结论肥胖小鼠脂肪组织缺氧抑制了脂联素的表达,抑制作用可能发生在转录水平;其机制可能是通过抑制PPAR-γmRNA的表达和PPAR-γ转录因子的活性而实现的。     

Renal cortical cyclooxygenase 2 expression is differentially regulated by angiotensin II AT(1) and AT(2) receptors

Macula densa cyclooxygenase 2 (COX-2)-derived prostaglandins serve as important modulators of the renin-angiotensin system, and cross-talk exists between these two systems. Cortical COX-2 induction by angiotensin-converting enzyme (ACE) inhibitors or AT(1) receptor blockers (ARBs) suggests that angiotensin II may inhibit cortical COX-2 by stimulating the AT(1) receptor pathway. In the present studies we determined that chronic infusion of either hypertensive or nonhypertensive concentrations of angiotensin II attenuated cortical COX-2. Angiotensin II infusion reversed cortical COX-2 elevation induced by ACE inhibitors. However, we found that angiotensin II infusion further stimulated cortical COX-2 elevation induced by ARBs, suggesting a potential role for an AT(2) receptor-mediated pathway when the AT(1) receptor was inhibited. Both WT and AT(2) receptor knockout mice were treated for 7 days with either ACE inhibitors or ARBs. Cortical COX-2 increased to similar levels in response to ACE inhibition in both knockout and WT mice. In WT mice ARBs increased cortical COX-2 more than ACE inhibitors, and this stimulation was attenuated by the AT(2) receptor antagonist PD123319. In the knockout mice ARBs led to significantly less cortical COX-2 elevation, which was not attenuated by PD123319. PCR confirmed AT(1a) and AT(2) receptor expression in the cultured macula densa cell line MMDD1. Angiotensin II inhibited MMDD1 COX-2, and CGP42112A, an AT(2) receptor agonist, stimulated MMDD1 COX-2. In summary, these results demonstrate that macula densa COX-2 expression is oppositely regulated by AT(1) and AT(2) receptors and suggest that AT(2) receptor-mediated cortical COX-2 elevation may mediate physiologic effects that modulate AT(1)-mediated responses.     

FKBP5 expression in human adipose tissue increases following dexamethasone exposure and is associated with insulin resistance

Objective

To study effects of dexamethasone on gene expression in human adipose tissue aiming to identify potential novel mechanisms for glucocorticoid-induced insulin resistance.

Materials/methods

Subcutaneous and omental adipose tissue, obtained from non-diabetic donors (10 M/15 F; age: 28–60 years; BMI: 20.7–30.6 kg/m²), was incubated with or without dexamethasone (0.003–3 μmol/L) for 24 h. Gene expression was assessed by microarray and real time-PCR and protein expression by immunoblotting.

Results

FKBP5 (FK506-binding protein 5) and CNR1 (cannabinoid receptor 1) were the most responsive genes to dexamethasone in both subcutaneous and omental adipose tissue (~ 7-fold). Dexamethasone increased FKBP5 gene and protein expression in a dose-dependent manner in both depots. The gene product, FKBP51 protein, was 10-fold higher in the omental than in the subcutaneous depot, whereas the mRNA levels were similar. Higher FKBP5 gene expression in omental adipose tissue was associated with reduced insulin effects on glucose uptake in both depots. Furthermore, FKBP5 gene expression in subcutaneous adipose tissue was positively correlated with serum insulin, HOMA-IR and subcutaneous adipocyte diameter and negatively with plasma HDL-cholesterol. FKBP5 SNPs were found to be associated with type 2 diabetes and diabetes-related phenotypes in large population-based samples.

Conclusions

Dexamethasone exposure promotes expression of FKBP5 in adipose tissue, a gene that may be implicated in glucocorticoid-induced insulin resistance.     

Activation of the hexosamine signaling pathway in adipose tissue results in decreased serum adiponectin and skeletal muscle insulin resistance

Overexpression of the rate-limiting enzyme for hexosamine synthesis (glutamine:fructose-6-phosphate amidotransferase) in muscle and adipose tissue of transgenic mice was previously shown to result in insulin resistance and hyperleptinemia. Explanted muscle from transgenic mice was not insulin resistant in vitro, suggesting that muscle insulin resistance could be mediated by soluble factors from fat tissue. To dissect the relative contributions of muscle and fat to hexosamine-induced insulin resistance, we overexpressed glutamine:fructose-6-phosphate amidotransferase 2.5-fold, specifically in fat under control of the aP2 promoter. Fasting glucose, insulin, and triglycerides were unchanged in the transgenic mice; leptin and beta-hydroxybutyrate levels were 91% and 29% higher, respectively. Fasted transgenic mice have mild glucose intolerance and skeletal muscle insulin resistance in vivo. In fasting transgenic mice, glucose disposal rates with hyperinsulinemia were decreased 27% in females and 10% in males. Uptake of 2-deoxy-D-glucose into muscle was diminished by 45% in female and 21% in male transgenics. Serum adiponectin was also lower in the fasted transgenics, by 37% in females and 22% in males. TNF alpha and resistin mRNA levels in adipose tissue were not altered in the fasted transgenics; levels of mRNA for leptin were increased and peroxisome proliferator-activated receptor gamma decreased. To further explore the relationship between adiponectin and insulin sensitivity, we examined mice that have been refed for 6 h after a 24-h fast. Refeeding wild-type mice resulted in decreased serum adiponectin and increased leptin. In transgenic mice, however, the regulation of these hormones by refeeding was lost for adiponectin and diminished for leptin. Refed transgenic female and male mice no longer exhibited decreased serum adiponectin in the refed state, and they were no longer insulin resistant as by lower or unchanged insulin and glucose levels. We conclude that increased hexosamine levels in fat, mimicking excess nutrient delivery, are sufficient to cause insulin resistance in skeletal muscle. Changes in serum adiponectin correlate with the insulin resistance of the transgenic animals.     

In humans the adiponectin receptor R2 is expressed predominantly in adipose tissue and linked to the adipose tissue expression of MMIF‐1

In this study, the regional adipose tissue‐adiponectin (AT‐ADN) and adiponectin receptor (R1 and R2) expression and their relation with metabolic parameters, circulating and AT‐derived cytokine expressions were compared. Paired subcutaneous adipose tissue (SCAT) and visceral adipose tissue (VAT) were taken from 18 lean and 39 obese humans, AT‐mRNA expression of adipokines analysed by RT‐PCR and corresponding serum levels by enzyme‐linked immunosorbent assay (ELISA). R1 and R2 adipocyte expression was compared with 17 other human tissues. ADN‐gene expression was lower in VAT than SCAT [mean (SD) 1.54 (1.1) vs. 2.84 (0.87); p < 0.001], and lower in obese subjects (VAT : p = 0.01;SCAT : p < 0.001). SCAT‐ADN correlated positively with serum ADN (r = 0.33;p = 0.036) but not VAT‐ADN. AT expressions of ADN and macrophage migration inhibiting factor (MMIF), IL18 and cluster of differentiation factor 14 (CD14) in both depots showed inverse correlations. R1 and R2 were expressed ubiquitously and R2 highest in SCAT, and this is much higher (×100) than R1 (×100). R expression was similar in lean and obese subjects and unrelated to the metabolic syndrome, however, receptors correlated with VAT‐MMIF (R 1: r = 0.4;p = 0.008;R 2: r = 0.35,p = 0.02) and SCAT‐MMIF expression (R 2: r = 0.43;p = 0.004). Unlike ADN, its receptors are expressed in many human tissues. Human R2 expression is not highest in the liver but in AT where it is associated with MMIF expression. The adiponectin‐dependent insulin‐sensitizing action of thiazolidinediones is thus probably to differ amongst species with weaker effects on the human liver.     

The polygenetically inherited metabolic syndrome of WOKW rats is associated with insulin resistance and altered gene expression in adipose tissue

BACKGROUND: Wistar Ottawa Karlsburg W (RT1u) rats (WOKW) develop a complete metabolic syndrome closely resembling the human disease. The aim of this study was to characterize the phenotype of adipose tissue in WOKW rats with regard to adipocyte metabolism, insulin resistance, and gene expression and thus to define the phenotype more precisely. METHODS: Glucose metabolism, insulin sensitivity, and gene expression of key adipocyte genes, including adiponectin, interleukin 6 (Il6), 11 beta-hydroxysteroid dehydrogenase (11beta Hsd), peroxisome proliferator-activated receptor gamma (Ppar gamma), forkhead box O1 (Foxo1), glucose transporter 4 (Glut4), CCAAT/enhancer binding protein (C/ebp alpha), and fatty acid synthase (Fasn) were characterized in adipocytes from epididymal and subcutaneous fat depots of 28-week-old male WOKW rats and Dark Agouti (DA) controls. RESULTS: WOKW rats display decreased insulin-stimulated glucose uptake and decreased insulin sensitivity during lipogenesis and lipolysis in isolated adipocytes. The severe insulin resistance predominantly in epididymal adipose tissue of WOKW rats is associated with a 10-fold decrease in adipocyte adiponectin gene expression, decreased Ppar gamma, but increased Foxo1 gene expression compared to DA rats. CONCLUSIONS: Insulin resistance in adipose tissue is associated with altered adipocyte gene expression in WOKW rats, additionally completing the picture of the metabolic syndrome in this animal model. This fact not only qualifies the WOKW rat for further detailed analysis of genetic determinants of metabolic syndrome but also highlights its suitability for pharmacological research.     

运动对胰岛素抵抗大鼠脂联素表达的影响

目的:观察运动干预对高脂高糖喂养诱导胰岛素抵抗大鼠的循环和脂肪组织中脂联素表达的影响。方法:29只雄性SD大鼠被随机分为对照组9只和造模组20只,分别给予基础饲料与高脂高糖饲料喂养,6周后,将造模成功的18只大鼠再随机分为模型组9只,运动组9只。运动干预6周后,采用酶联免疫吸附法测定血清脂联素,逆转录聚合酶联反应(RT-PCR)法检测脂肪组织中脂联素mRNA的表达。结果:与对照组比较,模型组出现明显的胰岛素抵抗[胰岛素敏感指数(ISI)(-4.11±0.33)∶(-5.32±0.21),P0.01];血清脂联素(ng/ml)水平显著降低[(0.86±0.08)∶(0.77±0.09),P0.05]脂肪组织中脂联素mRNA表达减弱(P0.05)。运动6周后,与模型组比较,运动组胰岛素抵抗明显改善[ISI(-5.51±0.16)∶(-5.10±0.31),P0.01],血清脂联素(ng/ml)水平显著升高[(0.77±0.09)∶(0.86±0.10),P0.05];脂肪组织中脂联素mRNA表达增强(P0.05)。结论:运动干预能够明显改善胰岛素抵抗,其机理可能与调节脂肪组织中脂联素的表达有关。     

NF-kappaB-inducing kinase (NIK) mediates skeletal muscle insulin resistance: blockade by adiponectin

Enhanced levels of nuclear factor (NF)-κB-inducing kinase (NIK), an upstream kinase in the NF-κB pathway, have been implicated in the pathogenesis of chronic inflammation in diabetes. We investigated whether increased levels of NIK could induce skeletal muscle insulin resistance. Six obese subjects with metabolic syndrome underwent skeletal muscle biopsies before and six months after gastric bypass surgery to quantitate NIK protein levels. L6 skeletal myotubes, transfected with NIK wild-type or NIK kinase-dead dominant negative plasmids, were treated with insulin alone or with adiponectin and insulin. Effects of NIK overexpression on insulin-stimulated glucose uptake were estimated using tritiated 2-deoxyglucose uptake. NF-κB activation (EMSA), phosphatidylinositol 3 (PI3) kinase activity, and phosphorylation of inhibitor κB kinase β and serine-threonine kinase (Akt) were measured. After weight loss, skeletal muscle NIK protein was significantly reduced in association with increased plasma adiponectin and enhanced AMP kinase phosphorylation and insulin sensitivity in obese subjects. Enhanced NIK expression in cultured L6 myotubes induced a dose-dependent decrease in insulin-stimulated glucose uptake. The decrease in insulin-stimulated glucose uptake was associated with a significant decrease in PI3 kinase activity and protein kinase B/Akt phosphorylation. Overexpression of NIK kinase-dead dominant negative did not affect insulin-stimulated glucose uptake. Adiponectin treatment inhibited NIK-induced NF-κB activation and restored insulin sensitivity by restoring PI3 kinase activation and subsequent Akt phosphorylation. These results indicate that NIK induces insulin resistance and further indicate that adiponectin exerts its insulin-sensitizing effect by suppressing NIK-induced skeletal muscle inflammation. These observations suggest that NIK could be an important therapeutic target for the treatment of insulin resistance associated with inflammation in obesity and type 2 diabetes.