Impaired insulin action in subcutaneous adipocytes from women with visceral obesity

Visceral obesity is associated with resistance to the antilipolytic effect of insulin in vivo. We investigated whether subcutaneous abdominal and gluteal adipocytes from viscerally obese women exhibit insulin resistance in vitro. Subjects were obese black and white premenopausal nondiabetic women matched for visceral adipose tissue (VAT), total adiposity, and age. Independently of race and adipocyte size, increased VAT was associated with decreased sensitivity to insulin's antilipolytic effect in subcutaneous abdominal and gluteal adipocytes. Absolute lipolytic rates at physiologically relevant concentrations of insulin or the adenosine receptor agonist N(6)-(phenylisopropyl)adenosine were higher in subjects with the highest vs. lowest VAT area. Independently of cell size, abdominal adipocytes were less sensitive to the antilipolytic effect of insulin than gluteal adipocytes, which may partly explain increased nonesterified fatty acid fluxes in upper vs. lower body obese women. Moreover, increased VAT was associated with decreased responsiveness, but not decreased sensitivity, to insulin's stimulatory effect on glucose transport in abdominal adipocytes. These data suggest that insulin resistance of subcutaneous abdominal and, to a lesser extent, gluteal adipocytes may contribute to increased systemic lipolysis in both black and white viscerally obese women.     

Insulin sensitive and resistant obesity in humans: AMPK activity, oxidative stress, and depot-specific changes in gene expression in adipose tissue

We previously reported that adenosine monophosphate-activated protein kinase (AMPK) activity is lower in adipose tissue of morbidly obese individuals who are insulin resistant than in comparably obese people who are insulin sensitive. However, the number of patients and parameters studied were small. Here, we compared abdominal subcutaneous, epiploic, and omental fat from 16 morbidly obese individuals classified as insulin sensitive or insulin resistant based on the homeostatic model assessment of insulin resistance. We confirmed that AMPK activity is diminished in the insulin resistant group. A custom PCR array revealed increases in mRNA levels of a wide variety of genes associated with inflammation and decreases in PGC-1α and Nampt in omental fat of the insulin resistant group. In contrast, subcutaneous abdominal fat of the same patients showed increases in PTP-1b, VEGFa, IFNγ, PAI-1, and NOS-2 not observed in omental fat. Only angiotensinogen and CD4(+) mRNA levels were increased in both depots. Surprisingly, TNFα was only increased in epiploic fat, which otherwise showed very few changes. Protein carbonyl levels, a measure of oxidative stress, were increased in all depots. Thus, adipose tissues of markedly obese insulin resistant individuals uniformly show decreased AMPK activity and increased oxidative stress compared with insulin sensitive patients. However, most changes in gene expression appear to be depot-specific.     

Vaspin gene in rat adipose tissue: relation to obesity-induced insulin resistance

Visceral adipose fat has been claimed to be the link between obesity and insulin resistance through the released adipokines. This study aimed to assess the expression of vaspin as one of the recent adipokines in rats abdominal subcutaneous and visceral fat in diet-induced obese (DIO) and in DIO performing 3 weeks swimming exercise (DIO + EXE) compared to control and control + exercise (C + EXE) groups. Vaspin mRNA and protein expression assessed using RT-PCR and Western blotting analysis revealed vaspin expression in DIO and DIO + EXE but not in controls groups. In DIO group, visceral vaspin expression was higher than in that of subcutaneous fat and was positively correlated with body weight. Upregulation of visceral vaspin expression in DIO was concomitant with the development of insulin resistance (increase in fasting serum insulin and HOMA-IR) and rise in serum leptin level. Unchanged visceral vaspin mRNA in DIO + EXE rats, with significant improvements of insulin resistance parameters and serum leptin compared to DIO group was found. In conclusion, increased visceral vaspin expression in obesity was associated with insulin resistance. Further investigations into the molecular links between vaspin and obesity may unravel innovative therapeutic strategies in people affected by obesity-linked insulin resistance, metabolic syndrome, and type 2 diabetes.     

The Roles of Adipokines,Proinflammatory Cytokines,and Adipose Tissue Macrophages in Obesity-Associated Insulin Resistance in Modest Obesity and Early Metabolic Dysfunction

The roles of adipokines, proinflammatory cytokines, and adipose tissue macrophages in obesity-associated insulin resistance have been explored in both animal and human studies. However, our current understanding of obesity-associated insulin resistance relies on studies of artificial metabolic extremes. The purpose of this study was to explore the roles of adipokines, proinflammatory cytokines, and adipose tissue macrophages in human patients with modest obesity and early metabolic dysfunction. We obtained omental adipose tissue and fasting blood samples from 51 females undergoing gynecologic surgery. We investigated serum concentrations of proinflammatory cytokines and adipokines as well as the mRNA expression of proinflammatory and macrophage phenotype markers in visceral adipose tissue using ELISA and quantitative RT-PCR. We measured adipose tissue inflammation and macrophage infiltration using immunohistochemical analysis. Serum levels of adiponectin and leptin were significantly correlated with HOMA-IR and body mass index. The levels of expression of MCP-1 and TNF-α in visceral adipose tissue were also higher in the obese group (body mass index ≥ 25). The expression of mRNA MCP-1 in visceral adipose tissue was positively correlated with body mass index (r = 0.428, p = 0.037) but not with HOMA-IR, whereas TNF-α in visceral adipose tissue was correlated with HOMA-IR (r = 0.462, p = 0.035) but not with body mass index. There was no obvious change in macrophage phenotype or macrophage infiltration in patients with modest obesity or early metabolic dysfunction. Expression of mRNA CD163/CD68 was significantly related to mitochondrial-associated genes and serum inflammatory cytokine levels of resistin and leptin. These results suggest that changes in the production of inflammatory biomolecules precede increased immune cell infiltration and induction of a macrophage phenotype switch in visceral adipose tissue. Furthermore, serum resistin and leptin have specific roles in the regulation of adipose tissue macrophages in patients with modest obesity or early metabolic dysfunction.     

Amyloid precursor protein expression is upregulated in adipocytes in obesity

The aim of this study was to determine whether amyloid precursor protein (APP) is expressed in human adipose tissue, dysregulated in obesity, and related to insulin resistance and inflammation. APP expression was examined by microarray expression profiling of subcutaneous abdominal adipocytes (SAC) and cultured preadipocytes from obese and nonobese subjects. Quantitative real-time PCR (QPCR) was performed to confirm differences in APP expression in SAC and to compare APP expression levels in adipose tissue, adipocytes, and stromal vascular cells (SVCs) from subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT) specimens. Adipose tissue samples were also examined by western blot and immunofluorescence confocal microscopy. Microarray studies demonstrated that APP mRNA expression levels were higher in SAC (approximately 2.5-fold) and preadipocytes (approximately 1.4) from obese subjects. Real-time PCR confirmed increased APP expression in SAC in a separate group of obese compared with nonobese subjects (P=0.02). APP expression correlated to in vivo indices of insulin resistance independently of BMI and with the expression of proinflammatory genes, such as monocyte chemoattractant protein-1 (MCP-1) (R=0.62, P=0.004), macrophage inflammatory protein-1alpha (MIP-1alpha) (R=0.60, P=0.005), and interleukin-6 (IL-6) (R=0.71, P=0.0005). Full-length APP protein was detected in adipocytes by western blotting and APP and its cleavage peptides, Abeta40 and Abeta42, were observed in SAT and VAT by immunofluorescence confocal microscopy. In summary, APP is highly expressed in adipose tissue, upregulated in obesity, and expression levels correlate with insulin resistance and adipocyte cytokine expression levels. These data suggest a possible role for APP and/or Abeta in the development of obesity-related insulin resistance and adipose tissue inflammation.     

Inflammation Correlates With Markers of T‐Cell Subsets Including Regulatory T Cells in Adipose Tissue From Obese Patients

Accumulation of cytotoxic and T‐helper (T_h)1 cells together with a loss of regulatory T cells in gonadal adipose tissue was recently shown to contribute to obesity‐induced adipose tissue inflammation and insulin resistance in mice. Human data on T‐cell populations in obese adipose tissue and their potential functional relevance are very limited. We aimed to investigate abundance and proportion of T‐lymphocyte sub‐populations in human adipose tissue in obesity and potential correlations with anthropometric data, insulin resistance, and systemic and adipose tissue inflammation. Therefore, we analyzed expression of marker genes specific for pan‐T cells and T‐cell subsets in visceral and subcutaneous adipose tissue from highly obese patients (BMI >40 kg/m², n = 20) and lean to overweight control subjects matched for age and sex (BMI <30 kg/m²; n = 20). All T‐cell markers were significantly upregulated in obese adipose tissue and correlated with adipose tissue inflammation. Proportions of cytotoxic T cells and T_h1 cells were unchanged, whereas those of regulatory T cells and T_h2 were increased in visceral adipose tissue from obese compared to control subjects. Systemic and adipose tissue inflammation positively correlated with the visceral adipose abundance of cytotoxic T cells and T_h1 cells but also regulatory T cells within the obese group. Therefore, this study confirms a potential role of T cells in human obesity‐driven inflammation but does not support a loss of protective regulatory T cells to contribute to adipose tissue inflammation in obese patients as suggested by recent animal studies.     

Abdominal adiposity and insulin resistance in obese men

We examined the independent relationships among various visceral and abdominal subcutaneous adipose tissue (AT) depots, glucose tolerance, and insulin sensitivity in 89 obese men. Measurements included an oral glucose tolerance test (OGTT), glucose disposal by euglycemic clamp, and abdominal and nonabdominal (e.g., peripheral) AT by magnetic resonance imaging (MRI). OGTT glucose and glucose disposal rates were related (P < 0.05) to visceral AT (r = 0.50 and -0.41, respectively). These observations remained significant (P < 0.05) after control for nonabdominal and abdominal subcutaneous AT, and maximal O(2) consumption (VO(2 max)). Abdominal subcutaneous AT was not a significant correlate (P > 0.05) of any metabolic variable after control for nonabdominal and visceral AT and VO(2 max). Division of abdominal subcutaneous AT into deep and superficial depots and visceral AT into intra- and extraperitoneal AT depots did not alter the observed relationships. Further analysis matched two groups of men for abdominal subcutaneous AT but also for low and high visceral AT. Men with high visceral AT had higher OGTT glucose values and lower glucose disposal rates compared with those with low visceral AT values (P < 0.05). A similar analysis performed on two groups of men matched for visceral AT but also for high and low abdominal subcutaneous AT revealed no statistically different values for any metabolic variable (P > 0.10). In conclusion, visceral AT alone is a strong correlate of insulin resistance independent of nonabdominal and abdominal subcutaneous AT and cardiovascular fitness. Subdivision of visceral and abdominal subcutaneous AT by MRI did not provide additional insight into the relationship between abdominal obesity and metabolic risk in obese men.     

Tissue leptin and plasma insulin are associated with lipoprotein lipase activity in severely obese patients

The development of metabolic complications of obesity has been associated with the existence of depot-specific differences in the biochemical properties of adipocytes. The aim of this study was to investigate, in severely obese men and women, both gender- and depot-related differences in lipoprotein lipase (LPL) expression and activity, as well as the involvement of endocrine and biometric factors and their dependence on gender and/or fat depot. Morbidly obese, nondiabetic, subjects (9 men and 22 women) aged 41.1+/-1.9 years, with a body mass index (BMI) of 54.7+/-1.7 kg/m(2) who had undergone abdominal surgery were studied. Both expression and activity of LPL and leptin expression were determined in adipose samples from subcutaneous and visceral fat depots. In both men and women, visceral fat showed higher LPL mRNA levels as well as lower ob mRNA levels and tissue leptin content than the subcutaneous one. In both subcutaneous and visceral adipose depots, women exhibited higher protein content, decreased fat cell size and lower LPL activity than men. The gender-related differences found in abdominal fat LPL activity could contribute to the increased risk for developing obesity-associated diseases shown by men, even in morbid obesity, in which the massive fat accumulation could mask these differences. Furthermore, the leptin content of fat depots as well as plasma insulin concentrations appear in our population as the main determinants of adipose tissue LPL activity, adjusted by gender, depot and BMI.     

Clodronate liposomes improve metabolic profile and reduce visceral adipose macrophage content in diet-induced obese mice

Background

Obesity-related adipose inflammation has been thought to be a causal factor for the development of insulin resistance and type 2 diabetes. Infiltrated macrophages in adipose tissue of obese animals and humans are an important source for inflammatory cytokines. Clodronate liposomes can ablate macrophages by inducing apoptosis. In this study, we aim to determine whether peritoneal injection of clodronate liposomes has any beneficial effect on systemic glucose homeostasis/insulin sensitivity and whether macrophage content in visceral adipose tissue will be reduced in diet-induced obese (DIO) mice.

Methodology/Principal Findings

Clodronate liposomes were used to deplete macrophages in lean and DIO mice. Macrophage content in visceral adipose tissue, metabolic parameters, glucose and insulin tolerance, adipose and liver histology, adipokine and cytokine production were examined. Hyperinsulinemic-euglycemic clamp study was also performed to assess systemic insulin sensitivity. Peritoneal injection of clodronate liposomes significantly reduced blood glucose and insulin levels in DIO mice. Systemic glucose tolerance and insulin sensitivity were mildly improved in both lean and DIO mice treated with clodronate liposomes by intraperitoneal (ip) injection. Hepatosteatosis was dramatically alleviated and suppression of hepatic glucose output was markedly increased in DIO mice treated with clodronate liposomes. Macrophage content in visceral adipose tissue of DIO mice was effectively decreased without affecting subcutaneous adipose tissue. Interestingly, levels of insulin sensitizing hormone adiponectin, including the high molecular weight form, were significantly elevated in circulation.

Conclusions/Significance

Intraperitoneal injection of clodronate liposomes reduces visceral adipose tissue macrophages, improves systemic glucose homeostasis and insulin sensitivity in DIO mice, which can be partially attributable to increased adiponectin levels.     

Regulation of Adipose Tissue Inflammation and Insulin Resistance by MAPK Phosphatase 5

Obesity and metabolic disorders such as insulin resistance and type 2 diabetes have become a major threat to public health globally. The mechanisms that lead to insulin resistance in type 2 diabetes have not been well understood. In this study, we show that mice deficient in MAPK phosphatase 5 (MKP5) develop insulin resistance spontaneously at an early stage of life and glucose intolerance at a later age. Increased macrophage infiltration in white adipose tissue of young MKP5-deficient mice correlates with the development of insulin resistance. Glucose intolerance in MKP5-deficient mice is accompanied by significantly increased visceral adipose weight, reduced AKT activation, enhanced p38 activity, and increased inflammation in visceral adipose tissue when compared with wild-type (WT) mice. Deficiency of MKP5 resulted in increased inflammatory activation in macrophages. These findings thus demonstrate that MKP5 critically controls inflammation in white adipose tissue and the development of metabolic disorders.     

PPARγ2 Pro12Ala polymorphism is associated with improved lipoprotein lipase functioning in adipose tissue of insulin resistant obese women

Lipoprotein lipase (LPL) plays a pivotal role in lipid metabolism, contributes to metabolic disorders related to insulin action and body weight regulation, and is influenced by inflammation. The Pro12Ala polymorphism of the peroxisome proliferator-activated receptor (PPAR)γ2 gene seems to influence LPL functioning, but its role in obesity and insulin resistance status, which usually coexist in the clinical setting, has not been explored. Our aim was to analyze the association of obesity and insulin resistance with adipose LPL activity and expression, and the influence of the PPARγ2 Pro12Ala polymorphism. A cross-sectional study was conducted in 58 reproductive-age women who underwent elective abdominal surgery. Free-fatty acids, glucose, insulin, and selected adipokines were measured in fasting blood samples. DNA was isolated and the polymorphism genotyped. Biopsies of abdominal subcutaneous adipose tissue obtained during surgery were used to determine enzymatic LPL activity and expression; and expression of selected cytokines. Overweight/obese women presented lower LPL activity (P = 0.022) and higher circulating TNF-α (P = 0.020) than controls. Insulin resistant women also showed borderline lower LPL activity than non-resistant (P = 0.052), but adiposity and inflammatory molecules were comparable. Nevertheless, LPL activity was higher in Pro12Ala carriers than in non-carriers after adjusting for obesity, insulin resistance and inflammation. Likewise, adipose LPL expression was increased in carriers while expression of cytokines was decreased. Our data suggest that insulin resistance is associated with low adipose LPL activity independently of obesity, but the PPARγ2 Pro12Ala polymorphism seems to protect the LPL functioning of obese insulin resistant women, likely through regulating inflammation in adipose tissue.     

Subdivisions of subcutaneous abdominal adipose tissue and insulin resistance

Whereas truncal (central) adiposity is strongly associated with the insulin resistant metabolic syndrome, it is uncertain whether this is accounted for principally by visceral adiposity (VAT). Several recent studies find as strong or stronger association between subcutaneous abdominal adiposity (SAT) and insulin resistance. To reexamine the issue of truncal adipose tissue depots, we performed cross-sectional abdominal computed tomography, and we undertook the novel approach of partitioning SAT into the plane superficial to the fascia within subcutaneous adipose tissue (superficial SAT) and that below this fascia (deep SAT), as well as measurement of VAT. Among 47 lean and obese glucose-tolerant men and women, insulin-stimulated glucose utilization, measured by euglycemic clamp, was strongly correlated with both VAT and deep SAT (r = -0.61 and -0.64, respectively; both P < 0.001), but not with superficial SAT (r = -0.29, not significant). Also, VAT and deep SAT followed a highly congruent pattern of associations with glucose and insulin area under the curve (75-g oral glucose tolerance test), mean arterial blood pressure, apoprotein-B, high-density lipoprotein cholesterol, and triglyceride. Superficial SAT had markedly weaker association with all these parameters and instead followed the pattern observed for thigh subcutaneous adiposity. We conclude that there are two functionally distinct compartments of adipose tissue within abdominal subcutaneous fat and that the deep SAT has a strong relation to insulin resistance.     

Toll-like receptor 3 ablation prevented high-fat diet-induced obesity and metabolic disorder

Inflammation in insulin-sensitive tissues (e.g., liver, visceral adipose tissue [VAT]) plays a major role in obesity and insulin resistance. Recruitment of innate immune cells drives the dysregulation of glucose and lipid metabolism. We aimed to seek the role of Toll like receptor 3 (TLR3), a pattern recognition receptor involved in innate immunity, obesity and the metabolic disorder. TLR3 expression in liver and VAT from diet induced obese mice and in VAT from overweight women was examined. Body weight, glucose homeostasis and insulin sensitivity were evaluated in TLR3 wild-type and knockout (KO) mice on a chow diet (CD) or high-fat diet for 15 weeks. At euthanasia, blood was collected, and plasma biochemical parameters and adipokines were determined with commercial kits. Flow cytometry was used to measure macrophage infiltration and activation in VAT. Standard western blot, immunohistochemistry and quantative PCR were used to assess molecules in pathways about lipid and glucose metabolism, insulin and inflammation in tissues of liver and VAT. Utilizing human and animal samples, we found that expression of TLR3 was upregulated in the liver and VAT in obese mice as well as VAT in overweight women. TLR3-deficiency protected against high-fat diet induced obesity, glucose intolerance, insulin resistance and lipid accumulation. Lipolysis was enhanced in VAT and hepatic lipogenesis was inhibited in TLR3 KO animals. Macrophages infiltration into adipose tissue was attenuated in TLR3 KO mice, accompanied with inhibition of NF-κB-dependent AMPK/Akt signaling pathway. These findings demonstrated that TLR3 ablation prevented obesity and metabolic disorders, thereby providing new mechanistic links between inflammation and obesity and associated metabolic abnormalities in lipid/glucose metabolism.     

Differential patterns of serum concentration and adipose tissue expression of chemerin in obesity: Adipose depot specificity and gender dimorphism

Chemerin, a recognized chemoattractant, is expressed in adipose tissue and plays a role in adipocytes differentiation and metabolism. Gender- and adipose tissue-specific differences in human chemerin expression have not been well characterized. Therefore, these differences were assessed in the present study. The body mass index (BMI) and the circulating levels of chemerin and other inflammatory, adiposity and insulin resistance markers were assessed in female and male adults of varying degree of obesity. Chemerin mRNA expression was also measured in paired subcutaneous and visceral adipose tissue samples obtained from a subset of the study subjects. Serum chemerin concentrations correlated positively with BMI and serum leptin levels and negatively with high density lipoprotein (HDL)-cholesterol levels. No correlation was found between serum chemerin concentrations and fasting glucose, total cholesterol, low density lipoprotein (LDL)-cholesterol, triglycerides, insulin, C-reactive protein or adiponectin. Similarly, no relation was observed with the homeostasis model assessment for insulin resistance (HOMA-IR) values. Gender- and adipose tissue-specific differences were observed in chemerin mRNA expression levels, with expression significantly higher in women than men and in subcutaneous than visceral adipose tissue. Interestingly, we found a significant negative correlation between circulating chemerin levels and chemerin mRNA expression in subcutaneous fat. Among the subjects studied, circulating chemerin levels were associated with obesity markers but not with markers of insulin resistance. At the tissue level, fat depot-specific differential regulation of chemerin mRNA expression might contribute to the distinctive roles of subcutaneous vs. visceral adipose tissue in human obesity.