Thymic stromal lymphopoietin: an immune cytokine gene associated with the metabolic syndrome and blood pressure in severe obesity.

A previous expression profiling of VAT (visceral adipose tissue) revealed that the TSLP (thymic stromal lymphopoietin) gene was less expressed in severely obese men with (n=7) compared with without (n=7) the MetS (metabolic syndrome). We hypothesized that TSLP SNPs (single nucleotide polymorphisms) are associated with TSLP gene expression in VAT and with MetS phenotypes. Following validation of lower TSLP expression (P=0.003) in VAT of severely obese men and women with (n=70) compared with without (n=60) the MetS, a detailed genetic investigation was performed at the TSLP locus by sequencing its promoter, exons and intron-exon splicing boundaries using DNA of 25 severely obese subjects. Five tagging SNPs were genotyped in the 130 subjects from the expression analysis to test whether these SNPs contributed to TSLP expression variability (ANOVAs) and then genotyped in two independent samples of severely obese men (total, n=389) and women (total, n=894). In a sex-stratified multistage experimental design, ANOVAs were performed to test whether tagging SNPs were associated with MetS components treated as continuous variables. We observed that the non-coding SNP rs2289277 was associated with TSLP mRNA abundance (P=0.04), as well as with SBP [systolic BP (blood pressure)] (P=0.004) and DBP (diastolic BP) (P=0.0003) in men when adjusting for age, waist circumference, smoking and medication treating hypertension. These novel observations suggest that TSLP expression in VAT may partly explain the inter-individual variability for metabolic impairments in the presence of obesity and that specific SNPs (rs2289277 and/or correlating SNPs) may influence TSLP gene expression as well as BP in obese men.

Genetic contribution to C-reactive protein levels in severe obesity.

Obese individuals are characterized by a chronic, low-grade inflammatory state. Increased levels of C-reactive protein (CRP), a marker of inflammation, have been observed in subjects with the metabolic syndrome. We have previously reported that genes encoding proteins involved in the anti-inflammatory and immune response are differentially expressed in visceral adipose tissue of obese men with or without the metabolic syndrome. Among these genes, the interferon-gamma-inducible protein 30 (IFI30), CD163 molecule (CD163), chemokine (C-X-C motif) ligand 9 (CXCL9) and thymic stromal lymphopoietin (TSLP), were selected for further genetic analyses. The aim of the study was to verify whether IFI30, CD163, CXCL9 and TSLP gene polymorphisms contribute to explain the inter-individual variability of the inflammatory profile of obesity assessed by plasma high-sensitivity CRP concentrations. A total of 1185 severely obese individuals were genotyped for single nucleotide polymorphisms (SNPs) covering most of the sequence-derived genetic variability at the IFI30, CD163, CXCL9 and TSLP gene loci (total of 27 SNPs). Following measurement of plasma CRP levels, subjects were divided into two groups, low vs. high using the median value of plasma CRP levels (8.31 mg/L) as a cutoff point. Genotype frequencies were compared between groups. Associations between genotypes and plasma CRP levels (continuous variable) were also tested after adjustments for age, sex, smoking and BMI. The rs11554159 and rs7125 IFI30 SNPs showed a significant difference in genotype frequencies (p<0.05) between subgroups of low vs. high plasma CRP levels (wild type homozygotes: rs11554159=47% vs. 55%, rs7125=31% vs. 24%, for low vs. high CRP groups, respectively). The association between rs11554159 and CRP levels as a continuous variable remained significant (p=0.004). Both carriers of the GA and AA genotypes demonstrated, on average, a 13% lower CRP levels in comparison with GG homozygotes. No association was observed between SNPs in the CD163, CXCL9 and TSLP genes and CRP levels. The IFI30 rs11554159 polymorphism could partially explain the inter-individual variability observed in the inflammatory profile associated with obesity.

DPP4 gene DNA methylation in the omentum is associated with its gene expression and plasma lipid profile in severe obesity.

Severely obese subjects with the metabolic syndrome (MS) have higher dipeptidyl peptidase-4 (DPP4) expression in their visceral adipose tissue (VAT) compared to obese individuals without MS. We tested the hypothesis that methylation level of CpG sites in the DPP4 promoter CpG island in VAT was genotype-dependent and associated with DPP4 mRNA abundance and MS-related phenotypes. The VAT DNA was extracted in 92 severely obese premenopausal women undergoing biliopancreatic derivation for the treatment of obesity. Women were nondiabetic and none of them used medication to treat MS features. Cytosine methylation rates (%) of 102 CpG sites in the DPP4 CpG island were assessed by pyrosequencing of sodium bisulfite-treated DNA. Methylation rates were >10% for CpG sites 94-102. Their mean methylation rate (%Meth(94-102)) was different between genotypes for DPP4 polymorphisms rs13015258 (P = 0.001), rs17848915 (P = 0.0004), and c.1926 G>A (P = 0.001). The %Meth(94-102) correlated negatively with DPP4 mRNA abundance (r = -0.25, P < 0.05) and positively with plasma high-density lipoprotein (HDL) cholesterol concentrations (r = 0.22, P < 0.05), whereas DPP4 mRNA abundance correlated positively with plasma total-/HDL-cholesterol ratio (r = 0.25; P < 0.05). In the VAT of nondiabetic severely obese women, genotype-dependent methylation levels of specific CpG sites in the DPP4 promoter CpG island were associated with DPP4 gene expression and variability in the plasma lipid profile. Higher DPP4 gene expression in VAT and its relationship with the plasma lipid profile may be explained by actually unknown DPP4 biological effect or, to another extent, may also be a marker of VAT inflammation known to be associated with metabolic disturbances.

Association of OSBPL11 gene polymorphisms with cardiovascular disease risk factors in obesity.

The prevalence of morbid obesity and its associated metabolic complications has risen rapidly in the past decade. Recently, we have established the transcriptome of the visceral adipose tissue of nondiabetic severely obese men with and without metabolic syndrome (MetS) that provided new candidate genes for cardiovascular disease (CVD) risk factors. The oxysterol-binding protein-like protein 11 (OSBPL11) that belongs to the OSBP family of intracellular receptors was one of the genes found to be significantly overexpressed in the MetS group. To determine whether OSBPL11 gene polymorphisms are associated with CVD risk factors and diabetes, OSBPL11 gene promoter and coding regions were sequenced in 25 individuals and six tagging single-nucleotide polymorphisms (SNPs) capturing 85% of gene sequence-derived common genetic variability (minor allele frequency (MAF) > 5%) were genotyped in two samples for a total of 962 obese individuals. Using a multistage experimental design, chi(2)-tests and logistic regressions were applied to compare genotype frequencies and to compute odds ratios (ORs) for low and high CVD risk groups. Significant associations between rs1055419 and diastolic blood pressure (OR = 0.53; P = 0.01) were found whereas IVS12+95 T>C, a newly discovered SNP, was associated with low-density lipoprotein-cholesterol levels (OR = 1.63; P < 0.001), hyperglycemia/diabetes (OR = 1.48; P < 0.004) as well as with MetS per se (OR = 1.56; P < 0.01). These results suggest that the OSBPL11 gene is involved in cholesterol and glucose metabolism in obese individuals.

Comprehensive genetic analysis of the dipeptidyl peptidase-4 gene and cardiovascular disease risk factors in obese individuals.

The incretin system has been shown to stimulate insulin secretion in a glucose dependent manner and currently fosters considerable hope for the treatment of diabetes. Recently, we have shown that the dipeptidylpeptidase-4 (DPP4) gene, which is responsible for incretin inactivation, was overexpressed in omental adipose tissue of obese men with the metabolic syndrome, compared to men not characterized by this condition. Since the cardiovascular disease (CVD) risk profile shows substantial inter-individual variability in obesity, this study aimed at verifying whether DPP4 polymorphisms contribute to explain such a difference. In the first step of this multi-stage study, seven tagging SNPs were genotyped in a sample of 576 obese (BMI>40 kg/m(2)) individuals and tested for their association with blood pressure and lipids, as well as diabetes-related phenotypes. Then, in an additional sample of 572 obese individuals (stage 2), SNPs showing trends (P<0.10) for an association in the first sample were genotyped and reanalyzed. Logistic regressions were used to compute odds ratio for obesity-related metabolic complications. In sample 1, homozygotes for rs17848915 and rs7608798 minor alleles were at lower risk of hyperglycemia/diabetes (P=0.002) and elevated plasma triglyceride levels (P=0.030) respectively, whereas rs1558957 heterozygotes were at higher risk to have high plasma triglyceride (P=0.040), HDL- (P=0.021), LDL- (P=0.001) and total-cholesterol (P=0.003) levels. However, none of these associations was consistently replicated in stage 2. This first comprehensive genetic analysis does not support the notion that DPP4 polymorphisms could modulate the CVD risk profile among obese patients.