Differences in gene expression of enzymes involved in branched-chain amino acid metabolism of abdominal subcutaneous adipose tissue between pregnant women with and without PCOS.

OBJECTIVE: Polycystic ovary syndrome (PCOS) appears to be a common endocrine disorder of women in reproductive age. Adipose tissue (AT) is known as an active tissue in the metabolism of branched-chain amino acids (BCAA; Valine, Leucine, and Isoleucine) that they have associated with blood BCAA levels is a prognostic factor for insulin-resistant. Although the crucial roles of AT in women suffering from PCOS was reported, little information exists on the BCAA metabolism in AT of PCOS women. The aim was to assess and compare the expression of BCAAs metabolism pathway genes in abdominal subcutaneous AT of pregnant women with PCOS and non-PCOS pregnant women. MATERIALS AND METHODS: AT samples from 13 PCOS were compared with samples collected from 6 non-PCOS women, all of whom underwent caesarean. Quantitative real-time PCR technique was used for gene expression of branched chain aminotransferase 2 mitochondrial (BCAT2), branched chain ketoacid dehydrogenase E1-alpha (BCKDHA), branched chain ketoacid dehydrogenase E1-Beta (BCKDHB), dihydrolipoamide branched chain transacylase E2 (DBT), dihydrolipoamide dehydrogenase E3 (DLD), branched chain ketoacid dehydrogenase kinase (BCKDK), Data were analyzed using t-test or U-test. RESULTS: No significant differences were found in age and body mass index (BMI) between non-PCOS and PCOS women. The mRNA level of BCAT2 and DLD in PCOS group was not significantly different from non-PCOS group whereas mRNA level of BCKDHB and DBT was significantly increased in PCOS group (P < 0.0001). In contrast, mRNA level of BCKDHA (P = 0.0001) and BCKDK (P < 0.0001) was significantly decreased in PCOS group. CONCLUSION: The alterations in gene expressions involved BCAA metabolism in age-matched and BMI- matched non-PCOS and PCOS pregnant women at delivery day was shown which warrants further studies regards functional activity. More attention should be given to AT of PCOS mothers that was previously ignored.

Deletion of interleukin 1 receptor-associated kinase 1 (Irak1) improves glucose tolerance primarily by increasing insulin sensitivity in skeletal muscle.

Chronic inflammation may contribute to insulin resistance via molecular cross-talk between pathways for pro-inflammatory and insulin signaling. Interleukin 1 receptor-associated kinase 1 (IRAK-1) mediates pro-inflammatory signaling via IL-1 receptor/Toll-like receptors, which may contribute to insulin resistance, but this hypothesis is untested. Here, we used male Irak1 null (k/o) mice to investigate the metabolic role of IRAK-1. C57BL/6 wild-type (WT) and k/o mice had comparable body weights on low-fat and high-fat diets (LFD and HFD, respectively). After 12 weeks on LFD (but not HFD), k/o mice (versus WT) had substantially improved glucose tolerance (assessed by the intraperitoneal glucose tolerance test (IPGTT)). As assessed with the hyperinsulinemic euglycemic glucose clamp technique, insulin sensitivity was 30% higher in the Irak1 k/o mice on chow diet, but the Irak1 deletion did not affect IPGTT outcomes in mice on HFD, suggesting that the deletion did not overcome the impact of obesity on glucose tolerance. Moreover, insulin-stimulated glucose-disposal rates were higher in the k/o mice, but we detected no significant difference in hepatic glucose production rates (± insulin infusion). Positron emission/computed tomography scans indicated higher insulin-stimulated glucose uptake in muscle, but not liver, in Irak1 k/o mice in vivo Moreover, insulin-stimulated phosphorylation of Akt was higher in muscle, but not in liver, from Irak1 k/o mice ex vivo In conclusion, Irak1 deletion improved muscle insulin sensitivity, with the effect being most apparent in LFD mice.

Influence of betaine and arginine supplementation of reduced protein diets on fatty acid composition and gene expression in the muscle and subcutaneous adipose tissue of cross-bred pigs.

The isolated or combined effects of betaine and arginine supplementation of reduced protein diets (RPD) on fat content, fatty acid composition and mRNA levels of genes controlling lipid metabolism in pig m. longissimus lumborum and subcutaneous adipose tissue (SAT) were assessed. The experiment was performed on forty intact male pigs (Duroc×Large White×Landrace cross-breed) with initial and final live weights of 60 and 93 kg, respectively. Pigs were randomly assigned to one of the following five diets (n 8): 16·0 % of crude protein (control), 13·0 % of crude protein (RPD), RPD supplemented with 0·33 % of betaine, RPD supplemented with 1·5 % of arginine and RPD supplemented with 0·33 % of betaine and 1·5 % of arginine. Data confirmed that RPD increase intramuscular fat (IMF) content and total fat content in SAT. The increased total fat content in SAT was accompanied by higher GLUT type 4, lipoprotein lipase and stearoyl-CoA desaturase mRNA expression levels. In addition, the supplementation of RPD with betaine and/or arginine did not affect either IMF or total fat in SAT. However, dietary betaine supplementation slightly affected fatty acid composition in both muscle and SAT. This effect was associated with an increase of carnitine O-acetyltransferase mRNA levels in SAT but not in muscle, which suggests that betaine might be involved in the differential regulation of some key genes of lipid metabolism in pig muscle and SAT. Although the arginine-supplemented diet decreased the mRNA expression level of PPARG in muscle and SAT, it did not influence fat content or fatty acid composition in any of these pig tissues.

Enhanced insulin signaling in human skeletal muscle and adipose tissue following gastric bypass surgery.

Roux-en-Y gastric bypass (RYGB) leads to increased peripheral insulin sensitivity. The aim of this study was to investigate the effect of RYGB on expression and regulation of proteins involved in regulation of peripheral glucose metabolism. Skeletal muscle and adipose tissue biopsies from glucose-tolerant and type 2 diabetic subjects at fasting and during a hyperinsulinemic-euglycemic clamp before as well as 1 wk and 3 and 12 mo after RYGB were analyzed for relevant insulin effector proteins/signaling components. Improvement in peripheral insulin sensitivity mainly occurred at 12 mo postsurgery when major weight loss was evident and occurred concomitantly with alterations in plasma adiponectin and in protein expression/signaling in peripheral tissues. In skeletal muscle, protein expression of GLUT4, phosphorylated levels of TBC1D4, as well as insulin-induced changes in phosphorylation of Akt and glycogen synthase activity were enhanced 12 mo postsurgery. In adipose tissue, protein expression of GLUT4, Akt2, TBC1D4, and acetyl-CoA carboxylase (ACC), phosphorylated levels of AMP-activated protein kinase and ACC, as well as insulin-induced changes in phosphorylation of Akt and TBC1D4, were enhanced 12 mo postsurgery. Adipose tissue from glucose-tolerant subjects was the most responsive to RYGB compared with type 2 diabetic patients, whereas changes in skeletal muscle were largely similar in these two groups. In conclusion, an improved molecular insulin-sensitive phenotype of skeletal muscle and adipose tissue appears to contribute to the improved whole body insulin action following a substantial weight loss after RYGB.

17ß-Estradiol and estradiol fatty acyl esters and estrogen-converting enzyme expression in adipose tissue in obese men and women.

CONTEXT: Obesity is associated with increased circulating 17ß-estradiol (E2), but less is known about E2 concentrations in adipose tissue. In addition to E2, adipose tissue synthesizes E2 fatty acyl esters (E2-FAE). OBJECTIVE: The aim was to compare estrogen concentrations and expression of estrogen-converting enzymes in adipose tissue between severely obese men and women. DESIGN AND SETTING: Tissue samples were obtained during elective surgery in University Central Hospital in the years 2008 through 2011. PATIENTS: We studied 14 men and 22 premenopausal women undergoing bariatric surgery and 10 control women operated for nonmalignant reasons. INTERVENTIONS: Paired samples were taken from abdominal sc and visceral adipose tissue and serum and analyzed for E2 and E2-FAE by fluoroimmunoassay and liquid chromatography-tandem mass spectrometry. mRNA expression of genes was analyzed by quantitative PCR. RESULTS: Compared with men, E2 levels in sc adipose tissue in obese women were higher, along with higher relative mRNA expression of steroid sulfatase and 17ß-hydroxysteroid dehydrogenases 1, 7, and 12. In men, E2-FAE concentrations in adipose tissue were similar to E2 but in women significantly lower compared with E2. Adipose tissue E2-FAE and serum E2-FAE levels correlated positively in obese subjects. Serum E2 did not significantly correlate with E2 concentration or mRNA expression of genes in adipose tissue in obese men or women. CONCLUSIONS: The production of E2 by the large adipose mass was not reflected by increased circulating E2 concentrations in severely obese men or women. However, adipose tissue may contribute to concentrations of serum E2-FAE.

Population pharmacokinetic/pharmacodynamic model of subcutaneous adipose 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) activity after oral administration of AMG 221, a selective 11ß-HSD1 inhibitor.

Inhibition of 11ß-HSD1 is hypothesized to improve measures of insulin sensitivity and hepatic glucose output in patients with type II diabetes. AMG 221 is a potent, small molecule inhibitor of 11ß-HSD1. The objective of this analysis is to describe the pharmacokinetic/pharmacodynamic (PK/PD) relationship between AMG 221 and 11ß-HSD1 inhibition in ex vivo adipose tissue samples. Healthy, obese subjects were administered a single dose of 3, 30, or 100 mg of oral AMG 221 (n = 44) or placebo (n = 11). Serial blood samples were collected over 24 hours. Subcutaneous adipose tissue samples were collected by open biopsy. Population PK/PD analysis was conducted using NONMEM. The inhibitory effects (mean ± standard error of the estimate) of AMG 221 on 11ß-HSD1 activity were directly related to adipose concentrations with I(max) (the maximal inhibition of 11ß-HSD1 activity) and IC50 (the plasma AMG 221 concentration associated with 50% inhibition of enzyme activity) of 0.975 ± 0.003 and 1.19 ± 0.12 ng/mL, respectively. The estimated baseline 11ß-HSD1 enzyme activity was 755 ± 61 pmol/mg. An equilibration rate constant (k(eo)) of 0.220 ± 0.021 h⁻¹ described the delay between plasma and adipose tissue AMG 221 concentrations. AMG 221 potently blocked 11ß-HSD1 activity, producing sustained inhibition for the 24-hour study duration as measured in ex vivo adipose samples. Early characterization of concentration-response relationships can support rational selection of dose and regimen for future studies.

Obesity is accompanied by disturbances in peripheral glucocorticoid metabolism and changes in FA recycling.

The glucocorticoid activating enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11betaHSD1) is of major interest in obesity-related morbidity. Alterations in tissue-specific cortisol levels may influence lipogenetic and gluco/glyceroneogenetic pathways in fat and liver. We analyzed the expression and activity of 11betaHSD1 as well as the expression of phosphoenolpyruvate carboxykinase (PEPCK), sterol regulatory element binding protein (SREBP), and fatty acid synthase (FAS) in adipose and liver and investigated putative associations between 11betaHSD1 and energy metabolism genes. A total of 33 obese women (mean BMI 44.6) undergoing gastric bypass surgery were enrolled. Subcutaneous adipose tissue (SAT), omental fat (omental adipose tissue (OmAT)), and liver biopsies were collected during the surgery. 11betaHSD1 gene expression was higher in SAT vs. OmAT (P = 0.013), whereas the activity was higher in OmAT (P = 0.009). The SAT 11betaHSD1 correlated with waist circumference (P = 0.045) and was an independent predictor for the OmAT area in a linear regression model. Energy metabolism genes had AT depot-specific expression; higher leptin and SREBP in SAT than OmAT, but higher PEPCK in OmAT than SAT. The expression of 11betaHSD1 correlated with PEPCK in both AT depots (P = 0.05 for SAT and P = 0.0001 for OmAT). Hepatic 11betaHSD1 activity correlated negatively with abdominal adipose area (P = 0.002) and expression positively with PEPCK (P = 0.003). In human obesity, glucocorticoid regeneration in the SAT is associated with central fat accumulation indicating that the importance of this specific fat depot is underestimated. Central fat accumulation is negatively associated with hepatic 11betaHSD1 activity. A disturbance in peripheral glucocorticoid metabolism is associated with changes in genes involved in fatty acid (FA) recycling in adipose tissue (AT).

Circulating IL-6 concentrations and abdominal adipocyte isoproterenol-stimulated lipolysis in women.

OBJECTIVE: To examine the association of plasma interleukin-6 (IL-6) concentrations with adiposity and fat cell metabolism in women. METHODS AND PROCEDURES: Omental (OM) and subcutaneous (SC) adipose tissue samples were obtained from 48 healthy women (age: 47+/-5 years, BMI: 26.9+/-5.3 kg/m(2)) undergoing gynecological surgeries. Total and visceral adiposity were assessed by dual-energy X-ray absorptiometry and computed tomography, respectively. Measures of adipocyte lipolysis (basal, isoproterenol-, forskolin-, and cyclic dibutyryl-adenosine monophosphate (AMP)-stimulated) and adipose tissue lipoprotein lipase (LPL) activity were obtained. Plasma IL-6 was measured by radioimmunoassay. RESULTS: Plasma IL-6 was positively correlated with total body fat mass (r=0.32, P<0.05), SC adipose tissue area (r=0.35, P<0.05), SC adipocyte diameter (r=0.30, P<0.05), and a trend was observed with visceral adipose tissue area (r=0.20, P<0.07). Plasma IL-6 was positively correlated with glycerol released in response to isoproterenol (10(-5) to 10(-8) mol/l) by isolated SC (0.31

Intra-adipose sex steroid metabolism and body fat distribution in idiopathic human obesity.

OBJECTIVE: Causes of visceral fat accumulation include glucocorticoid excess or decreased oestrogen/androgen ratio either in plasma or within adipose tissue. In obese subjects, the intra-adipose cortisol-generating enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) is increased, but information on sex steroid signalling is sparse. We aimed to test associations between body fat or fat distribution and mRNA transcript levels for androgen and oestrogen receptors and for enzymes metabolizing sex steroids in adipose tissue. DESIGN: A cross-sectional study. PATIENTS: Forty-five healthy men and women with body mass index (BMI) 21-36 kg/m(2). MEASUREMENTS: In subcutaneous adipose biopsies we measured mRNAs for enzymes metabolizing local oestrogens (aromatase) and androgens [5alpha-reductase type 1; AKR1C2 (3alpha-HSD3); AKR1C3 (17beta-HSD5, 3alpha-HSD2)] and for sex steroid receptors [oestrogen receptor (ER)-alpha and androgen receptor (AR)]. We related these to body fat mass and distribution. RESULTS: Generalized obesity (BMI) was associated with increased aromatase mRNA (r = 0.35, P < 0.05). Central obesity (waist : hip ratio) was associated with mRNA for AKR1C2 (r = 0.28, P < 0.05) and AKR1C3 (r = 0.38, P < 0.01) but not aromatase (r = 0.06). 5alpha-Reductase type 1, ER and AR mRNA levels did not predict fat amount or its distribution. CONCLUSION: These data on transcript levels suggest that, in idiopathic obesity, increased intra-adipose oestrogen generation by aromatase predicts peripheral fat distribution, while androgen metabolism by AKR1C isoforms predicts central fat distribution, supporting the hypothesis that intra-adipose sex steroid metabolism is a determinant of gynoid vs. android patterns of body fat.