Beneficial effects of green tea polyphenols (GTP) against obesity have been reported, however, the mechanism of this protection is not clear. Therefore, the objective of this study was to identify GTP-targeted genes in obesity using the high-fat-diet-induced obese rat model. A total of three groups (n = 12/group) of Sprague Dawley (SD) female rats were tested, including the control group (rats fed with low-fat diet), the HF group (rats fed with high-fat diet), and the HF+GTP group (rats fed with high-fat diet and GTP in drinking water). The HF group increased body weight as compared to the control group. Supplementation of GTP in the drinking water in the HF+GTP group reduced body weight as compared to the HF group. RNA from liver samples was extracted for gene expression analysis. A total of eighty-four genes related to obesity were analyzed using PCR array. Compared to the rats in the control group, the rats in the HF group had the expression levels of 12 genes with significant changes, including 3 orexigenic genes (Agrp, Ghrl, and Nr3c1); 7 anorectic genes (Apoa4, Cntf, Ghr, IL-1β, Ins1, Lepr, and Sort); and 2 genes that relate to energy expenditure (Adcyap1r1 and Adrb1). Intriguingly, the HF+GTP group restored the expression levels of these genes in the high-fat-induced obese rats. The protein expression levels of IL-1β and IL-6 in the serum samples from the control, HF, and HF+GTP groups confirmed the results of gene expression. Furthermore, the protein expression levels of superoxide dismutase-1 (SOD1) and catechol-O-methyltransferase (COMT) also showed GTP-regulated protective changes in this obese rat model. Collectively, this study revealed the beneficial effects of GTP on body weight via regulating obesity-related genes, anti-inflammation, anti-oxidant capacity, and estrogen-related actions in high-fat-induced obese rats.

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Green tea polyphenols reduce body weight in rats by modulating obesity-related genes.

Background: Candidate genes of psychological importance include 5HT2A, 5HT2C, and COMT, implicated in the serotonin, noradrenaline and dopamine pathways, which also may be involved in regulation of energy balance. We investigated the associations of single nucleotide polymorphisms (SNPs) of these genes with obesity and metabolic traits. Methodology/Principal Findings: In a population of 166 200 young men examined at the draft boards, obese men (n=726, BMI$31.0 kg/m 2 ) and a randomly selected group (n=831) were re-examined at two surveys at mean ages 46 and 49 years (S-46, S-49). Anthropometric, physiological and biochemical measures were available. Logistic regression analyses were used to assess age-adjusted odds ratios. No significant associations were observed of 5HT2A rs6311, 5HT2C rs3813929 and COMT rs4680 with obesity, except that COMT rs4680 GG-genotype was associated with fat-BMI (OR=1.08, CI=1.01–1.16). The SNPs were associated with a number of physiological variables; most importantly 5HT2C rs3813929 T-allele was associated with glucose (OR=4.56, CI=1.13–18.4) and acute insulin response (OR=0.65, CI=0.44–0.94) in S-49. COMT rs4680 GG-genotype was associated with glucose (OR=1.04, CI=1.00–1.09). Except for an association between 5HT2A rs6311 and totalcholesterol at both surveys, significant in S-46 (OR=2.66, CI=1.11–6.40), no significant associations were observed for the other phenotypes. Significant associations were obtained when combined genotype of 5HT2C rs3813929 and COMT rs4680 were examined in relation to BMI (OR=1.12, CI=1.03–1.21), fat-BMI (OR=1.22, CI=1.08–1.38), waist (OR=1.13, CI=1.04– 1.22), and cholesterol (OR=5.60, CI=0.99–31.4). Analyses of impaired glucose tolerance (IGT) and type 2 diabetes (T2D) revealed, a 12.3% increased frequency of 5HT2C rs3813929 T-allele and an 11.6% increased frequency of COMT rs4680 GGgenotype in individuals with IGT or T2D (x 2 , p=0.05 and p=0.06, respectively). Examination of the combined genotypes of 5HT2C and COMT showed a 34.0% increased frequency of IGT or T2D (x 2 , p=0.01). Conclusions: The findings lend further support to the involvement of serotonin, noradrenaline and dopamine pathways on obesity and glucose homeostasis, in particular when combined genotype associations are explored.

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Polymorphisms of Serotonin Receptor 2A and 2C Genes and COMT in Relation to Obesity and Type 2 Diabetes

Catechol-O-methyltransferase (COMT) metabolizes 2-hydroxyestradiol into 2-methoxyestradiol (2-ME); COMT deficiency has shown to be associated with hypertension in men and preeclampsia, the disease associated with hypersensitivity of pressor response against angiotensin II (Ang II). Here, we found that COMT deficiency could explain the hypersensitivity of pressor response against Ang II in mice because of the lack of 2-ME-dependent suppression of angiotensin II receptor type 1 (AT1R). Male C57BL/6 mice were subjected to COMT inhibitor (COMTi: 25 mg/kg per day) or oil (control) for 4 weeks, with or without low-dose Ang II infusion (ANGII: 70 ng/kg per minute) for the last 3 weeks. The Ang II-infused mice were treated with 2-ME (10 ng/d) or vehicle for the last 1 week. We obtained the following experimental groups: control, ANGII, COMTi, COMTi+ANGII, and COMTi+ANGII+2-ME. We performed similar experiments using the in vivo administration of small interfering RNA of COMT instead of COMTi. Neither ANGII nor COMTi exhibited significant alterations in systolic blood pressure. Compared with ANGII or COMTi, COMTi+ANGII displayed significantly higher systolic blood pressure, albuminuria, and glomerular endotheliosis; 2-ME normalized such alterations. Similar phenotypes were observed in COMT small interfering RNA-treated mice. In the aorta of COMT-deficient mice, AT1R expression was increased; 2-ME suppressed AT1R expression. The 2-ME exhibited peroxisome proliferator-activated receptor γ agonistic activity in vitro and ex vivo plasma from pregnant female mice as well. In vitro, 2-ME suppressed both basal and Ang II-induced AT1R levels in a peroxisome proliferator-activated receptor γ-dependent manner. The 2-ME is relevant to combat COMT deficiency-associated hypertensive disorders via suppression of AT1R by its peroxisome proliferator-activated receptor γ activity.

Catechol-O-Methyltransferase Deficiency Leads to Hypersensitivity of the Pressor Response Against Angiotensin II.

In estrogen metabolic pathways, the COMT enzyme is related to detoxification. COMT gene polymorphisms have been shown to effect enzyme function. We hypothesized that these polymorphisms may be risk factors for endometrial cancer (EC). DNA samples from 150 cases of EC and healthy controls (n = 165) were analyzed by PCR-RFLP to determine the genotypic frequency of four different polymorphic loci on COMT [codon 62 (rs4633), 102 (rs5031015), 136 (rs4818), 158 (rs4680)]. Genotyping was confirmed by direct DNA sequencing. We also conducted haplotype analysis of COMT and investigated the relationship between COMT expression and COMT SNPs in EC tissues by immunohistochemistry. A significant increase in the T/T genotype of codon 62 (C/T) was observed in patients compared to controls (OR = 2.39, 95% CI: 1.31-4.37, P = 0.004). The frequency of the C-G haplotype of codon 62 C/T and codon 158 G/A was significantly higher in controls (P < 0.0001) than in patients. The expression level of COMT protein in EC tissues was significantly lower in COMT codon 62 variant TT and codon 158 variant AA genotype carriers. Therefore, the EC samples with polymorphic variants of COMT lead to lower expression of COMT protein whereas EC samples with wild-type codon 62 C/C and codon 158 G/G have higher expression of COMT protein. This is the first study demonstrating that polymorphisms in COMT codon 62 and codon 158 altered protein expression levels in EC, suggesting that they may be risk factors for EC in Caucasians.

COMT polymorphisms affecting protein expression are risk factors for endometrial cancer.

Several anthropometric measures have been associated with hormone-related cancers. However, it is unknown whether estrogen metabolism plays an important role in these relationships. We examined whether measured current body mass index (BMI), waist-to-hip ratio (WHR), height, and self-reported BMI at age 18 years were associated with serum estrogens/estrogen metabolites using baseline, cross-sectional data from 1835 postmenopausal women enrolled in the Women’s Health Initiative Observational Study. Fifteen estrogens/estrogen metabolites were quantified using liquid chromatography-tandem mass spectrometry. Geometric means (GMs) of estrogens/estrogen metabolites (in picomoles per liter) were estimated using inverse probability weighted linear regression, adjusting for potential confounders and stratified on menopausal hormone therapy (MHT) use. Among never or former MHT users, current BMI (≥30 vs. <25 kg/m2) was positively associated with parent estrogens (multivariable adjusted GM 432 vs. 239 pmol/L for estrone, 74 vs. 46 pmol/L for estradiol; p-trend < 0.001 for both) and all of the 2-, 4-, and 16-pathway estrogen metabolites evaluated (all p-trend ≤ 0.02). After additional adjustment for estradiol, unconjugated methylated 2-catechols were inversely associated (e.g., 2-methoxyestrone multivariable GM 9.3 vs. 12.0 pmol/L; p-trend < 0.001). Among current MHT users, current BMI was not associated with parent estrogens but was inversely associated with methylated catechols (e.g., 2-methoxyestrone multivariable GM 216 vs. 280 pmol/L; p-trend = 0.008). Similar patterns of association were found with WHR; however, the associations were not independent of BMI. Height and BMI at age 18 years were not associated with postmenopausal estrogens/estrogen metabolite levels. Our data suggest that postmenopausal BMI is associated with increased circulating levels of parent estrogens and reduced methylation of catechol estrogen metabolites, the estrogen metabolism patterns that have previously been associated with higher breast cancer risk.

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Anthropometric measures and serum estrogen metabolism in postmenopausal women: the Women’s Health Initiative Observational Study

Catechol-O-methyltransferase (COMT) is an important modulator in the catabolism of extraneural dopamine, which plays an important role in drug reward mechanisms. It is hypothesized that genetic variations in the COMT gene, which can result in a three to fourfold difference in COMT enzyme activity, may be associated with several reward-motivated behaviors. The aim of our study was to examine the relationship between COMT polymorphisms with smoking, obesity and alcohol. Three single nucleotide polymorphisms (SNPs) in COMT were genotyped in 2,371 participants selected randomly from the screening arm of the PLCO Cancer Screening Trial after stratifying by sex, age, and smoking status. Smoking, obesity, and alcohol consumption were assessed by questionnaire. SNP and haplotype associations were estimated using odds ratios (ORs) and 95% confidence intervals (CIs) derived from conditional logistic regression models, adjusted for race/ethnicity. The COMT Ex4-76C > G (Leu136Leu) polymorphism was statistically significantly associated with individuals who had >30% increases in BMI from ages 20 to 50 years, compared to those with 0-5% increase in BMI (0-5%) over the same age period: (CC is referent; OR(CG )= 1.42, OR(GG )= 1.46, P (trend )= 0.06). By sex, the increased risk was further pronounced among females (OR(CG )= 1.50, OR(GG )= 2.10, P (trend )= 0.03). Consistent with our analyses of single polymorphisms, individuals whose BMI increased >30% from ages 20 to 50 years were more likely than individuals with 0-5% increases in BMI to possess COMT haplotypes [COMT Ex3-104C > T-COMT Ex4-76 C > G-COMT Ex4-12 A > G] that included the variant allele for COMT Ex4-76 C > G: C-G-G (T-C-A is referent: OR(C-G-G )= 1.33, 95% CI 1.01-1.77) and C-G-A (OR(C-G-A )= 1.79, 95% CI 0.72-4.49). We observed no association between any of the COMT polymorphisms with smoking behavior or alcohol intake. The COMT Ex4-76C > G (Leu136Leu) polymorphism appears to play a role in large increases in BMI. The null association with smoking and alcohol and the pronounced association with increasing BMI among women further implicates COMT's role in estrogen metabolism as a potentially culpable pathway. Our results support a need for comprehensive evaluation of COMT variations and their functional relevance as COMT may be an important molecular target to evaluate for new treatments regarding obesity.

Elizabeth Z. Lan

Papers

Genetic variation in catechol-O-methyltransferase (COMT) and obesity in the prostate, lung, colorectal, and ovarian (PLCO) cancer screening trial.