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Central (visceral) obesity is more closely associated with insulin resistance, type 2 diabetes, and cardiovascular disease than is peripheral [subcutaneous (sc)] obesity, but the underlying mechanism for this pathophysiological difference is largely unknown. To understand the molecular basis of this difference, we sequenced 10,437 expressed sequence tags (ESTs) from a human omental fat cDNA library and discovered a novel visceral fat depot-specific secretory protein, which we have named omentin. Omentin ESTs were more abundant than many known adipose genes, such as perilipin, adiponectin, and leptin in the cDNA library. Protein sequence analysis indicated that omentin mRNA encodes a peptide of 313 amino acids, containing a secretory signal sequence and a fibrinogen-related domain. Northern analysis demonstrated that omentin mRNA was predominantly expressed in visceral adipose tissue and was barely detectable in sc fat depots in humans and rhesus monkeys. Quantative real-time PCR showed that omentin mRNA was expressed in stromal vascular cells, but not fat cells, isolated from omental adipose tissue, with >150-fold less in sc cell fractions. Accordingly, omentin protein was secreted into the culture medium of omental, but not sc, fat explants. Omentin was detectable in human serum by Western blot analysis. Addition of recombinant omentin in vitro did not affect basal but enhanced insulin-stimulated glucose uptake in both sc (47%, n = 9, P = 0.003) and omental (~30%, n = 3, P < 0.05) human adipocytes. Omentin increased Akt phosphorylation in the absence and presence of insulin. In conclusion, omentin is a new adipokine that is expressed in omental adipose tissue in humans and may regulate insulin action. obesity; diabetes; insulin resistance; visceral fat; metabolic syndrome

In its role as an endothelial cell proliferation and migration factor, vascular endothelial growth factor (VEGF) can affect peripheral circulation and therefore impact maximal oxygen consumption (V[O.sub.2 max]). Because of the role of VEGF, and because variation in the VEGF gene has the ability to alter VEGF gene expression and VEGF protein level, we hypothesized that VEGF gene polylnorphisms are related to VEGF gene expression in human myoblasts and V[O.sub.2 max] before and after aerobic exercise training. We analyzed the effects of the VEGF -2578/ -1154/-634 protnoter region haplotype on VEGF gene expression by using a luciferase reporter assay in cultured human myoblasts and found that the AAG and CGC haplotypes resulted m significantly higher hypoxia-stimulated VEGF gene expression than the AGG and CGG haplotypes. Consistent with these results, we found that individuals with at least one copy of the AAG or CGC haplotype had higher V[O.sub.2 max] before and after aerobic exercise training than did subjects with only the AGG and/or CGG haplotype. In conclusion, we found that VEGF-2578/-1154/-634 haplotype impacts VEGF gene expression in human myoblasts and is associated with V[O.sub.2 max]. These results have potential implications for aerobic exercise training and may prove relevant in the study of pathological conditions that can be affected by angiogenesis, such as coronary artery disease and peripheral artery disease. angiogenesis; exercise; genetics; polymorphism

The A4 complementary DNA (cDNA) expressed by the HT-29 human colon cancer cell line was analyzed to determine the biochemical function of the A4 protein. Analysis of the deduced amino acid of A4 cDNA indicated the presence of a hydropathy profile that was similar to transmembrane proteins. The hydrophobic sequence of the transmembrane domains of A4 also contained single, acidic amino acid residues that are similar to integral membrane proteins or proteolipids.

Thromboxane A2, an arachidonic acid-derived eicosanoid generated by thromboxane synthase (TBXAS), plays critical roles in hemostasis and inflammation. However, the contribution of thromboxane A2 to obesity-linked metabolic dysfunction remains incompletely understood. Here, we used in vitro and mouse models to better define the role of TBXAS in metabolic homeostasis. We found that adipose expression of Tbxas and thromboxane A2 receptor (Tbxa2r) was significantly upregulated in genetic and dietary mouse models of obesity and diabetes. Expression of Tbxas and Tbxa2r was detected in adipose stromal cells, including macrophages. Furthermore, stimulation of macrophages with interferon-7 or resistin factors known to be upregulated in obesity induced Tbxas and Tbxa2r expression. Mice lacking Tbxas had similar weight gain, food intake, and energy expenditure. However, loss of Tbxas markedly enhanced insulin sensitivity in mice fed a low-fat diet. Improvement in glucose homeostasis was correlated with the upregulated expression of multiple secreted metabolic regulators (Ctrp3, Ctrp9, and Ctrp12) in the visceral fat depot. Following a challenge with a high-fat diet, Tbxas deficiency led to attenuated adipose tissue fibrosis and reduced circulating IL-6 levels without adipose tissue macrophages being affected; however, these changes were not sufficient to improve whole body insulin action. Together, our results highlight a novel, diet-dependent role for thromboxane A2 in modulating peripheral tissue insulin sensitivity and adipose tissue fibrosis. [ABSTRACT FROM AUTHOR]

EIF2AK3 is a type I transmembrane protein that functions as an endoplasmic reticulum (ER) stress sensor to regulate global protein synthesis. Rare mutations in EIF2AK3 cause Wolcott-Rallison syndrome (OMIM 226980), an autosomal recessive disorder characterized by diabetes, epiphyseal dysplasia, osteoporosis, and growth retardation. To investigate the role of common genetic variation in EIF2AK3 as a determinant of bone mineral density (BMD) and osteoporosis, we sequenced all exons and flanking regions, then genotyped six potentially functional single nucleotide polymorphisms (SNPs) in this gene in 997 Amish subjects for association analysis, and attempted replication in 887 Mexican Americans. We found that the minor allele of a nonsynonymous SNP rs13045 had borderline associations with decreased forearm BMD in both discovery and replication cohorts (unadjusted p = 0.036 and β = − 0.007 for the Amish; unadjusted p = 0.031 and β = − 0.008 for Mexican Americans). A meta-analysis indicated this association achieved statistical significance in the combined sample (unadjusted p = 0.003; Bonferroni corrected p = 0.009). Rs13045 and three other potentially functional SNPs, a promoter SNP (rs6547787) and two nonsynonymous SNPs (rs867529 and rs1805165), formed two haplotypes: a low-BMD associated haplotype, denoted haplotype B [minor allele frequency (MAF) = 0.311] and a common haplotype A (MAF = 0.676). There were no differences in mRNA expression in lymphoblastoid cell lines between the two haplotypes. However, after treating lymphoblastoid cell lines with thapsigargin to induce ER stress, cell lines with haplotype B showed increased sensitivity to ER stress ( p = 0.014) compared with cell lines with haplotype A. Taken together, our results suggest that common nonsynonymous sequence variants in EIF2AK3 have a modest effect on ER stress response and may contribute to the risk for low BMD through this mechanism. © 2012 American Society for Bone and Mineral Research [ABSTRACT FROM AUTHOR]

Postprandial triglyceridemia is an emerging risk factor for cardiovascular disease. However, most of the genes that influence postprandial triglyceridemia are not known. We evaluated whether a common nonsynonymous SNP rs1260326/P446L in the glucokinase regulatory protein ( GCKR) gene influenced variation in the postprandial lipid response after a high-fat challenge in seven hundred and seventy participants in the Amish HAPI Heart Study who underwent an oral high-fat challenge and had blood samples taken in the fasting state and during the postprandial phase at 1, 2, 3, 4, and 6 h. We found that the minor T allele at rs1260326 was associated with significantly higher fasting TG levels after adjusting for age, sex, and family structure ( Pa = 0.06 for additive model, and Pr = 0.0003 for recessive model). During the fat challenge, the T allele was associated with significantly higher maximum TG level ( Pa = 0.006), incremental maximum TG level ( Pa = 0.006), TG area under the curve ( Pa = 0.02) and incremental TG area under the curve ( Pa = 0.03). Our data indicate that the rs1260326 T allele of GCKR is associated with both higher fasting levels of TG as well as the postprandial TG response, which may result in higher atherogenic risk. [ABSTRACT FROM AUTHOR]

Hemiparetic stroke leads to major skeletal muscle abnormalities, as illustrated by paretic leg atrophy, weakness, and spasticity. Furthermore, the hemiparetic limb muscle shifts to a fast-twitch muscle fiber phenotype with anaerobic metabolism. This study investigated whether skeletal muscle genes were altered in chronic hemiparetic stroke. The nonparetic leg muscle served as an internal control. We used Affymetrix microarray analysis to survey gene expression differences between paretic and nonparetic vastus lateralis muscle punch biopsies from 10 subjects with chronic hemiparetic stroke. Stroke latency was greater than 6 months. We found that 116 genes were significantly altered between the paretic and nonparetic vastus lateralis muscles. These gene differences were consistent with reported differences after stroke in areas such as injury and inflammation markers, the myosin heavy chain profile, and high prevalence of impaired glucose tolerance and type 2 diabetes. Furthermore, while many other families of genes were altered, the gene families with the most genes altered included inflammation, cell cycle regulation, signal transduction, metabolism, and muscle contractile protein genes. This study is an early step toward identification of specific gene regulatory pathways that might lead to these differences, propagate disability, and increase vascular disease risk. [ABSTRACT FROM AUTHOR]

Central (visceral) obesity is more closely associated with insulin resistance, type 2 diabetes, and cardiovascular disease than is peripheral [subcutaneous (sc)] obesity, but the underlying mechanism for this pathophysiological difference is largely unknown. To understand the molecular basis of this difference, we sequenced 10,437 expressed sequence tags (ESTs) from a human omental fat cDNA library and discovered a novel visceral fat depot-specific secretory protein, which we have named omentin. Omentin ESTs were more abundant than many known adipose genes, such as perilipin, adiponectin, and leptin in the cDNA library. Protein sequence analysis indicated that omentin mRNA encodes a peptide of 313 amino acids, containing a secretory signal sequence and a fibrinogen-related domain. Northern analysis demonstrated that omentin mRNA was predominantly expressed in visceral adipose tissue and was barely detectable in sc fat depots in humans and rhesus monkeys. Quantative real-time PCR showed that omentin mRNA was expressed in stromal vascular cells, but not fat cells, isolated from omental adipose tissue, with > 150-fold less in sc cell fractions. Accordingly, omentin protein was secreted into the culture medium of omental, but not sc, fat explants. Omentin was detectable in human serum by Western blot analysis. Addition of recombinant omentin in vitro did not affect basal but enhanced insulin-stimulated glucose uptake in both sc (47%, n = 9, P = 0.003) and omental (∼30%, n = 3, P < 0.05) human adipocytes. Omentin increased Akt phosphorylation in the absence and presence of insulin. In conclusion, omentin is a new adipokine that is expressed in omental adipose tissue in humans and may regulate insulin action. [ABSTRACT FROM AUTHOR]

Phosphorylation of Akt Substrate 160 (AS 160) by Akt and other kinases is the most proximal step in the regulation of GLUT4 translocation and glucose uptake. Akt and AS 160 phosphorylation increases after acute aerobic exercise and is associated with increased glucose transport in rodent models. We hypothesized that chronic weight loss and aerobic training (WL+AEX) would increase human skeletal muscle Akt and AS 160 phosphorylation with associated improvements in glucose utilization. To test this hypothesis, we measured in vivo glucose utilization (M: 3-hr 80 mU/m²/min euglycemic-hyperinsulinemic clamp) and basal and insulin-stimulated phosphorylation of Akt and AS 160 in vastus lateralis muscle biopsies (taken 120 min into the clamp) before and after 6-month WL+AEX (clamp and biopsies 36-48 hrs after exercise) in 10 non-diabetic men (age = 66 ± 6 yrs). Men were overweight/obese (BMI = 30 ± 4 kg/m²; % fat = 32 ±10 %; X ± SD) and sedentary (VO[sub 2]max = 20 ± 4 ml/kg/min). Aerobic capacity increased by 23% (p < 0.01), and body weight and fat mass (DXA) decreased by 7% and 21%, respectively (p < 0.05) after WL+AEX. M increased by 29% (39 ± 13 vs. 51 ± 15 umol/kg[sub BW]/min) and M/I increased by 38% (0.037 ± .016 to 0.051 ± 0.019 umol/kg[sub BW]/min/pmol; an index of insulin sensitivity). Both fasting glucose (98 ± 8 vs. 91 ± 6 mg/dl; p < 0.01), and insulin (93 ± 38 vs. 67 ± 22 pmol/ml; p < 0.05) decreased with WL+AEX. Hyperinsulinemia increased phosphorylation of Akt (6-fold; p < 0.01 and 7-fold; p < 0.01)) and AS160 (l.9-fold; p < 0.05 and 1.5-fold; p = 0.09) before and after the intervention respectively. The insulin stimulated increase in Akt and AS 160 phosphorylation did not change with WL+AEX. Neither the change in M nor M/I after WL+AEX was associated with the changes in Akt and AS 160 phosphorylation. In contrast to acute exercise in rodents, our results suggest that the improvement in glucose utilization after chronic WL+AEX in older men may not be mediated by increased activation of Akt and AS160 measured 36-48 hours after exercise training. [ABSTRACT FROM AUTHOR]

Thromboxane A2, an arachidonic acid-derived eicosanoid generated by thromboxane synthase (TBXAS), plays critical roles in hemostasis and inflammation. However, the contribution of thromboxane A2 to obesity-linked metabolic dysfunction remains incompletely understood. Here, we used in vitro and mouse models to better define the role of TBXAS in metabolic homeostasis. We found that adipose expression of Tbxas and thromboxane A2 receptor (Tbxa2r) was significantly upregulated in genetic and dietary mouse models of obesity and diabetes. Expression of Tbxas and Tbxa2r was detected in adipose stromal cells, including macrophages. Furthermore, stimulation of macrophages with interferon-γ or resistin factors known to be upregulated in obesity induced Tbxas and Tbxa2r expression. Mice lacking Tbxas had similar weight gain, food intake, and energy expenditure. However, loss of Tbxas markedly enhanced insulin sensitivity in mice fed a low-fat diet. Improvement in glucose homeostasis was correlated with the upregulated expression of multiple secreted metabolic regulators (Ctrp3, Ctrp9, and Ctrp12) in the visceral fat depot. Following a challenge with a high-fat diet, Tbxas deficiency led to attenuated adipose tissue fibrosis and reduced circulating IL-6 levels without adipose tissue macrophages being affected; however, these changes were not sufficient to improve whole body insulin action. Together, our results highlight a novel, diet-dependent role for thromboxane A2 in modulating peripheral tissue insulin sensitivity and adipose tissue fibrosis., (Copyright © 2015 the American Physiological Society.)

Hemiparetic stroke leads to major skeletal muscle abnormalities, as illustrated by paretic leg atrophy, weakness, and spasticity. Furthermore, the hemiparetic limb muscle shifts to a fast-twitch muscle fiber phenotype with anaerobic metabolism. This study investigated whether skeletal muscle genes were altered in chronic hemiparetic stroke. The nonparetic leg muscle served as an internal control. We used Affymetrix microarray analysis to survey gene expression differences between paretic and nonparetic vastus lateralis muscle punch biopsies from 10 subjects with chronic hemiparetic stroke. Stroke latency was greater than 6 months. We found that 116 genes were significantly altered between the paretic and non paretic vastus lateralis muscles. These gene differences were consistent with reported differences after stroke in areas such as injury and inflammation markers, the myosin heavy chain profile, and high prevalence of impaired glucose tolerance and type 2 diabetes. Furthermore, while many other families of genes were altered, the gene families with the most genes altered included inflammation, cell cycle regulation, signal transduction, metabolism, and muscle contractile protein genes. This study is an early step toward identification of specific gene regulatory pathways that might lead to these differences, propagate disability, and increase vascular disease risk.