Role of genetic variation in the cannabinoid type 1 receptor gene (CNR1) in the pathophysiology of human obesity.

AIMS: The endocannabinoid system may contribute to the association of visceral fat accumulation with metabolic diseases. Here we investigated the effects of genetic variation in the cannabinoid type 1 receptor gene (CNR1) on its mRNA expression in adipose tissue from visceral and subcutaneous depots and on the development of obesity. MATERIALS & METHODS: CNR1 was sequenced in 48 nonrelated German Caucasians to detect genetic variation. Five representative variants including HapMap tagging SNPs (rs12720071, rs806368, rs806370, rs1049353 and rs806369) were genotyped for subsequent association studies in two independent cohorts (total n = 2774) with detailed metabolic testing: subjects from the Leipzig Study (n = 1857) and a self-contained population of Sorbs from Germany (n = 917). RESULTS: In a case-control study of lean (BMI <25 kg/m(2)) versus obese (BMI >30 kg/m(2)) subjects, rs806368 was found to be nominally associated with obesity in the Sorbian cohort (adjusted p < 0.05), but not in the Leipzig cohort. Also, several SNPs (rs806368, rs806370 and rs12720071) were nominally associated with serum leptin levels (p < 0.05 after adjusting for age, sex and BMI). However, none of these associations remained significant after accounting for multiple testing. Furthermore, none of the SNPs were related to CNR1 mRNA expression in visceral and subcutaneous fat. CONCLUSION: The data suggest that common genetic variation in the CNR1 gene does not influence mRNA expression in adipose tissue nor does it play a significant role in the pathophysiology of obesity in German and Sorbian populations.

Effect of genetic variation in the human fatty acid synthase gene (FASN) on obesity and fat depot-specific mRNA expression.

Inhibition of fatty acid synthase (FASN) induces a rapid decline in fat stores in mice, suggesting a role for this enzyme in energy homeostasis. To investigate the potential role of FASN in the pathophysiology of human obesity, the FASN gene was sequenced in 48 German whites. Thirty-five single-nucleotide polymorphisms (SNPs) were identified. Eight SNPs representative for their linkage disequilibrium groups and the Val1483Ile (rs2228305) substitution were genotyped for subsequent association analyses in 1,311 adults from Germany. Further, the tagging SNPs were genotyped also in German childhood cohorts (738 schoolchildren, 205 obese children). Effects of genetic variation on FASN mRNA expression in visceral and subcutaneous adipose tissue from a subgroup of 172 subjects were analyzed. Several polymorphisms in the FASN (rs62078748, rs2229422, rs2229425, and rs17848939) were nominally associated with obesity in case-control studies including 446 obese subjects (BMI >or=30 kg/m(2)) and 389 lean controls (BMI

TCF7L2 gene expression in human visceral and subcutaneous adipose tissue is differentially regulated but not associated with type 2 diabetes mellitus.

Variants in the TCF7L2 gene have been associated with type 2 diabetes mellitus (T2DM), but the causal variant(s) is still unknown. We studied the TCF7L2 messenger RNA (mRNA) expression in paired samples of visceral and subcutaneous adipose tissue from 49 subjects using quantitative real-time polymerase chain reaction and its relation to obesity and T2DM. All subjects were genotyped for the previously described TCF7L2 diabetes risk variants. Independent of age, sex, obesity, and diabetes status, we found >3-fold higher TCF7L2 mRNA expression in subcutaneous compared with visceral adipose tissue. There was no correlation between visceral and subcutaneous TCF7L2 expression. No differences in adipose tissue TCF7L2 mRNA expression levels were found between diabetic and nondiabetic subjects, or between lean and obese subjects (all Ps > .05). In addition, there was no association between TCF7L2 genetic variants and mRNA expression. Based on our data, TCF7L2 mRNA expression is fat-depot specific but does not seem to provide the mechanistic link explaining genetic association with T2DM.

Gene expression of adiponectin receptors in human visceral and subcutaneous adipose tissue is related to insulin resistance and metabolic parameters and is altered in response to physical training.

OBJECTIVE: Adiponectin receptors 1 and 2 (AdipoR1 and AdipoR2, respectively) mediate the effects of adiponectin on glucose and lipid metabolism in vivo. We examined whether AdipoR1 and/or AdipoR2 mRNA expression in human adipose tissue is fat-depot specific. We also studied whether their expression in visceral and subcutaneous fat depots is associated with metabolic parameters and whether their expression is regulated by intensive physical exercise. RESEARCH DESIGN AND METHODS: We determined metabolic parameters and assessed AdipoR1 and AdipoR2 mRNA expression using quantitative real-time PCR in adipose tissue in an observational study of 153 subjects and an interventional study of 60 subjects (20 each with normal glucose tolerance, impaired glucose tolerance, and type 2 diabetes) before and after intensive physical training for 4 weeks. RESULTS: AdipoR1 and AdipoR2 mRNA expression is not significantly different between omental and subcutaneous fat, but their expression is several-fold lower in adipose tissue than in muscle. AdipoR2 mRNA expression in visceral fat is highly correlated with its expression in subcutaneous fat. AdipoR2 mRNA expression in both visceral and subcutaneous fat is positively associated with circulating adiponectin and HDL levels but negatively associated with obesity as well as parameters of insulin resistance, glycemia, and other lipid levels before and after adjustment for fat mass. Physical training for 4 weeks resulted in increased AdipoR1 and AdipoR2 mRNA expression in subcutaneous fat. CONCLUSIONS: AdipoR2 mRNA expression in fat is negatively associated with insulin resistance and metabolic parameters independently of obesity and may mediate the improvement of insulin resistance in response to exercise.

Effects of genetic variation in the human retinol binding protein-4 gene (RBP4) on insulin resistance and fat depot-specific mRNA expression.

OBJECTIVE: Serum retinol binding protein 4 (RBP4) is a new liver- and adipocyte-derived signal that may contribute to insulin resistance. Therefore, the RBP4 gene represents a plausible candidate gene involved in susceptibility to type 2 diabetes. RESEARCH DESIGN AND METHODS: In this study, the RBP4 gene was sequenced in DNA samples from 48 nonrelated Caucasian subjects. Five novel and three known single nucleotide polymorphisms (SNPs) were identified. Furthermore, five recently reported SNPs were genotyped in 90 subjects. Six SNPs, representative of their linkage disequilibrium groups, were then genotyped in 934 diabetic and 716 nondiabetic subjects. RESULTS: A haplotype of six common SNPs (A-G-G-T-G-C) was significantly increased in 934 case subjects with type 2 diabetes compared with 537 healthy control subjects with normal glucose tolerance (P = 0.02; odds ratio 1.37 [95% CI 1.05-1.79]). Furthermore, in the cohort of 716 nondiabetic Caucasian subjects, carriers of the A-G-G-T-G-C haplotype had significantly higher mean fasting plasma insulin and 2-h plasma glucose than subjects without the haplotype. Two single SNPs (rs10882283 and rs10882273) were also associated with BMI, waist-to-hip ratio, and fasting plasma insulin, and several SNPs were associated with circulating free fatty acids (all adjusted P < 0.05). In addition, subjects carrying a previously reported diabetes-associated haplotype had significantly higher mRNA levels in visceral adipose tissue (adjusted P < 0.05) in a subgroup of nondiabetic subjects (n = 170) with measurements of RBP4 mRNA expression in visceral and subcutaneous fat depots. CONCLUSIONS: Our data indicate a role of RBP4 genetic variation in susceptibility to type 2 diabetes and insulin resistance, possibly through an effect on RBP4 expression.

Serum retinol-binding protein is more highly expressed in visceral than in subcutaneous adipose tissue and is a marker of intra-abdominal fat mass.

Intra-abdominal fat is associated with insulin resistance and cardiovascular risk. Levels of serum retinol-binding protein (RBP4), secreted by fat and liver cells, are increased in obesity and type 2 diabetes (T2D). Here we report that, in 196 subjects, RBP4 is preferentially expressed in visceral (Vis) versus subcutaneous (SC) fat. Vis fat RBP4 mRNA was increased approximately 60-fold and 12-fold in Vis and SC obese subjects respectively versus lean subjects, and approximately 2-fold with impaired glucose tolerance/T2D subjects versus normoglycemic subjects. In obese subjects, serum RBP4 was increased 2- to 3-fold, and serum transthyretin, which stabilizes RBP4 in the circulation, was increased 35%. Serum RBP4 correlated positively with adipose RBP4 mRNA and intra-abdominal fat mass and inversely with insulin sensitivity, independently of age, gender, and body mass index. RBP4 mRNA correlated inversely with GLUT4 mRNA in Vis fat and positively with adipocyte size in both depots. RBP4 levels are therefore linked to Vis adiposity, and Vis fat may be a major source of RBP4 in insulin-resistant states.

TCF7L2 gene polymorphisms confer an increased risk for early impairment of glucose metabolism and increased height in obese children.

CONTEXT: Variants in the transcription factor 7-like2 (TCF7L2) gene have been associated with an increased risk for type 2 diabetes in adults. To evaluate whether the five reported risk variants confer a higher risk for obesity and early impairment of glucose metabolism in children, we genotyped these risk variants of the TCF7L2 gene in a representative cohort of 1029 Caucasian children and an independent cohort of 283 obese children. RESULTS: Applying a case control design, we observed a significantly lower prevalence of the rs11196205 and rs7895340 risk alleles in the obese (n = 283) compared with lean (n = 672) children (0.40 vs. 0.45; P = 0.02). There was, however, no statistically significant relationship between these genotypes and quantitative traits of obesity in either a normal representative cohort (n = 1029) or an obesity cohort. Obese children were significantly taller than lean children. This increase in height was independently associated with risk variants of the TCF7L2 gene, whereas in the normal representative cohort height appeared to be decreased in carriers of the minor alleles. In the obese cohort, three risk alleles (rs7901695, rs7903146, and rs1225572) were significantly associated with higher fasting and 120-min blood glucose levels independent of sex, age, pubertal stage, and body mass index. Fasting and peak insulin levels and HOMA-IR appeared with a similar tendency but were not statistically significant. CONCLUSIONS: Our data indicate for the first time that TCF7L2 gene variants confer an increased risk for early impairment of glucose metabolism in obese children, which is consistent with adult studies identifying TCF7L2 as a major diabetes susceptibility gene.

Dysregulation of the peripheral and adipose tissue endocannabinoid system in human abdominal obesity.

The endocannabinoid system has been suspected to contribute to the association of visceral fat accumulation with metabolic diseases. We determined whether circulating endocannabinoids are related to visceral adipose tissue mass in lean, subcutaneous obese, and visceral obese subjects (10 men and 10 women in each group). We further measured expression of the cannabinoid type 1 (CB(1)) receptor and fatty acid amide hydrolase (FAAH) genes in paired samples of subcutaneous and visceral adipose tissue in all 60 subjects. Circulating 2-arachidonoyl glycerol (2-AG) was significantly correlated with body fat (r = 0.45, P = 0.03), visceral fat mass (r = 0.44, P = 0.003), and fasting plasma insulin concentrations (r = 0.41, P = 0.001) but negatively correlated to glucose infusion rate during clamp (r = 0.39, P = 0.009). In visceral adipose tissue, CB(1) mRNA expression was negatively correlated with visceral fat mass (r = 0.32, P = 0.01), fasting insulin (r = 0.48, P < 0.001), and circulating 2-AG (r = 0.5, P < 0.001), whereas FAAH gene expression was negatively correlated with visceral fat mass (r = 0.39, P = 0.01) and circulating 2-AG (r = 0.77, P < 0.001). Our findings suggest that abdominal fat accumulation is a critical correlate of the dysregulation of the peripheral endocannabinoid system in human obesity. Thus, the endocannabinoid system may represent a primary target for the treatment of abdominal obesity and associated metabolic changes.

Evidence for a role of developmental genes in the origin of obesity and body fat distribution.

Obesity, especially central obesity, is a hereditable trait associated with a high risk for development of diabetes and metabolic disorders. Combined gene expression analysis of adipocyte- and preadipocyte-containing fractions from intraabdominal and subcutaneous adipose tissue of mice revealed coordinated depot-specific differences in expression of multiple genes involved in embryonic development and pattern specification. These differences were intrinsic and persisted during in vitro culture and differentiation. Similar depot-specific differences in expression of developmental genes were observed in human subcutaneous versus visceral adipose tissue. Furthermore, in humans, several genes exhibited changes in expression that correlated closely with body mass index and/or waist/hip ratio. Together, these data suggest that genetically programmed developmental differences in adipocytes and their precursors in different regions of the body play an important role in obesity, body fat distribution, and potential functional differences between internal and subcutaneous adipose tissue.

Genetic variation in the visfatin gene (PBEF1) and its relation to glucose metabolism and fat-depot-specific messenger ribonucleic acid expression in humans.

CONTEXT AND OBJECTIVE: Visfatin is a peptide suggested to play a role in glucose homeostasis. In the present study, we investigated the role of genetic variation in the visfatin gene in the pathophysiology of obesity/type 2 diabetes mellitus (T2DM). DESIGN: The visfatin gene (PBEF1) was sequenced in DNA samples from 24 nonrelated Caucasian subjects. Identified genetic variants were used for association analyses of T2DM in a case-control study (503 diabetic subjects and 476 healthy controls) and T2DM-related traits in 626 nondiabetic subjects. The effect of genetic variation in the visfatin gene on its mRNA expression in a subgroup of 157 nondiabetic subjects with measurements of visfatin mRNA expression in visceral and sc fat depots was also analyzed. RESULTS: Seven single-nucleotide polymorphisms (SNPs) and one insertion/deletion were identified. Three SNPs (rs9770242, -948G-->T, rs4730153) that were representatives of their linkage disequilibrium groups were genotyped in Caucasians from Germany with a wide range of body fat distribution and insulin sensitivity for association analyses. No association of T2DM with any of the genotyped SNPs or their haplotypes was found. However, the ratio of visceral/sc visfatin mRNA expression was associated with all three genetic polymorphisms (P < 0.05). Moreover, the -948G-->T variant was associated with 2-h plasma glucose and fasting insulin concentrations (P < 0.05) in nondiabetic subjects. CONCLUSIONS: In conclusion, our data suggest that genetic variation in the visfatin gene may have a minor effect on visceral and sc visfatin mRNA expression profiles but does not play a major role in the development of obesity or T2DM.

Vaspin gene expression in human adipose tissue: association with obesity and type 2 diabetes.

Recently, vaspin was identified as an adipokine with insulin-sensitizing effects, which is predominantly secreted from visceral adipose tissue in a rat model of type 2 diabetes. In this study, we examined whether vaspin mRNA expression is a marker of visceral obesity and correlates with anthropometric and metabolic parameters in paired samples of visceral and subcutaneous adipose tissue from 196 subjects with a wide range of obesity, body fat distribution, insulin sensitivity, and glucose tolerance. Vaspin mRNA expression was only detectable in 23% of the visceral and in 15% of the subcutaneous (SC) adipose tissue samples. Vaspin mRNA expression was not detectable in lean subjects (BMI<25) and was more frequently detected in patients with type 2 diabetes. No significant correlations were found between visceral vaspin gene expression and visceral fat area or SC vaspin expression. However, visceral vaspin expression significantly correlates with BMI, % body fat, and 2 h OGTT plasma glucose. Subcutaneous vaspin mRNA expression is significantly correlated with WHR, fasting plasma insulin concentration, and glucose infusion rate during steady state of an euglycemic-hyperinsulinemic clamp. Multivariate linear regression analysis revealed % body fat as strongest predictor of visceral vaspin and insulin sensitivity as strongest determinant of SC vaspin mRNA expression. In conclusion, our data indicate that induction of human vaspin mRNA expression in adipose tissue is regulated in a fat depot-specific manner and could be associated with parameters of obesity, insulin resistance, and glucose metabolism.

Plasma visfatin concentrations and fat depot-specific mRNA expression in humans.

Visceral and subcutaneous adipose tissue display important metabolic differences that underlie the association of visceral obesity with obesity-related cardiovascular and metabolic alterations. Recently, visfatin was identified as an adipokine, which is predominantly secreted from visceral adipose tissue both in humans and mice. In this study, we examined whether visfatin plasma concentrations (using enzyme immunosorbent assay) and mRNA expression (using RT-PCR) in visceral and subcutaneous fat correlates with anthropometric and metabolic parameters in 189 subjects with a wide range of obesity, body fat distribution, insulin sensitivity, and glucose tolerance. Visfatin plasma concentration correlates positively with the visceral visfatin mRNA expression (r(2) = 0.17, P < 0.0001), BMI (r(2) = 0.062, P = 0.004), percent body fat (r(2) = 0.048, P = 0.01), and negatively with subcutaneous visfatin mRNA expression (r(2) = 0.18, P < 0.0001). However, in a subgroup of 73 individuals, in which visceral fat mass was calculated from computed tomography scans, there was no correlation between plasma visfatin concentrations and visceral fat mass. We found no significant correlation between visfatin plasma concentrations and parameters of insulin sensitivity, including fasting insulin, fasting plasma glucose concentrations, and the glucose infusion rate during the steady state of an euglycemic-hyperinsulinemic clamp independent of percent body fat. Visfatin gene expression was not different between visceral and subcutaneous adipose tissue in the entire study group nor in selected subgroups. We found a significant correlation between visceral visfatin gene expression and BMI (r(2) = 0.06, P = 0.001) and percent body fat (measured using dual-energy X-ray absorptiometry) (r(2) = 0.044, P = 0.004), whereas no significant association between BMI or percent body fat and subcutaneous visfatin mRNA expression existed (both P >0.5). In conclusion, visfatin plasma concentrations and visceral visfatin mRNA expression correlated with measures of obesity but not with visceral fat mass or waist-to-hip ratio. In addition, we did not find differences in visfatin mRNA expression between visceral and subcutaneous adipose tissue in humans.