Glucocorticoids regulate the secretion of a 21kDa-IGF-binding protein by sheep adipose tissue explants

Using a solution phase assay we have demonstrated that sheep adipose tissue explants secrete insulin-like growth factor binding proteins (IGFBPs) when cultured in serum-free medium over a 24 h period. Further, we demonstrate that secretion of IGFBP(s) is inhibited (up to 50%) by incubation of the cultures in the presence of 10^–8M dexamethasone. This inhibitory effects is overcome when insulin (10 ng/ml) and ovine growth hormone (100 ng/ml) are incubated together (but not separately) with glucocorticoid. Further characterisation of this IGF binding activity by high performance size exclusion chromatography and Western ligand blot analysis indicated that under our culture conditions sheep adipose tissue explants secrete one predominant 21 kDa IGFBP and it is this BP which is hormonally regulated as described above. We discuss our results in the context of endocrine/paracrine/autocrine control of adipose tissue metabolism and differentiation.Abbreviations IGF insulin-like growth factor - IGF-BP insulin-like growth factor binding proteins - DX dexamethasone - GH ovine growth hormone - CM conditioned medium     

Adipose tissue as a secretory organ: from adipogenesis to the metabolic syndrome

Adipose tissue contains various types of cells that include preadipocytes and adipocytes. Studies have emphasized that (i) preadipocytes secrete factors involved in their own differentiation and (ii) adipocytes acquire the ability to communicate systemically with other organs (brain, liver, skeletal muscle) and locally with other cells (preadipocytes, endothelial cells and monocytes/macrophages). Adipocytes secrete proteins exhibiting either beneficial (leptin, adiponectin) or deleterious effects (angiotensinogen). Associated to the effect of secretory products from macrophages (cytokines), a disturbance in the balance between these various secreted factors leads to the development of a metabolic syndrome.     

Dietary fat modifies lipid metabolism in the adipose tissue of metabolic syndrome patients

Adipose tissue (AT) is a key organ in the regulation of total body lipid homeostasis, which is responsible for the storage and release of fatty acids according to metabolic needs. We aimed to investigate the effect of the quantity and quality of dietary fat on the lipogenesis and lipolysis processes in the AT of metabolic syndrome (MetS) patients. A randomized, controlled trial conducted within the LIPGENE study assigned MetS patients to one of four diets: (a) high-saturated fatty acid (HSFA) (b) high-monounsaturated fatty acid, and (c, d) two low-fat, high-complex carbohydrate diets supplemented with long chain (LC) n-3 (LFHCC n-3) polyunsaturated fatty acids (PUFA) or placebo (LFHCC), for 12 weeks each. A fat challenge reflecting the same fatty acid composition as the original diets was conducted post-intervention. Long-term consumption of the LFHCC diet induced an increase in the fasting expression levels of the sterol regulatory element binding protein-1 and stearoyl-CoA desaturase D9-desaturase genes, whereas the supplementation of this diet with n-3 PUFA reversed this effect (p = 0.007). In contrast, long-term consumption of the HSFA diet increased the expression of the adipose triglyceride lipase (ATGL) gene, at both fasting and postprandial states (both, p < 0.001). Our results showed the anti-lipogenic effect exerted by LC n-3 PUFA when administered together with a LFHCC diet. Conversely, a diet high in saturated fat increased the expression of the lipolytic gene ATGL relative to the other diets.

Electronic supplementary material

The online version of this article (doi:10.1007/s12263-014-0409-3) contains supplementary material, which is available to authorized users.     

Different metabolic responses of human brown adipose tissue to activation by cold and insulin

We investigated the metabolism of human brown adipose tissue (BAT) in healthy subjects by determining its cold-induced and insulin-stimulated glucose uptake and blood flow (perfusion) using positron emission tomography (PET) combined with computed tomography (CT). Second, we assessed gene expression in human BAT and white adipose tissue (WAT). Glucose uptake was induced 12-fold in BAT by cold, accompanied by doubling of perfusion. We found a positive association between whole-body energy expenditure and BAT perfusion. Insulin enhanced glucose uptake 5-fold in BAT independently of its perfusion, while the effect on WAT was weaker. The gene expression level of insulin-sensitive glucose transporter GLUT4 was also higher in BAT as compared to WAT. In conclusion, BAT appears to be differently activated by insulin and cold; in response to insulin, BAT displays high glucose uptake without increased perfusion, but when activated by cold, it dissipates energy in a perfusion-dependent manner.     

Antioxidant system response is modified by dietary fat in adipose tissue of metabolic syndrome patients

Metabolic syndrome (MetS) is associated with high oxidative stress, which is caused by an increased expression of NADPH-oxidase and a decreased expression of antioxidant enzymes in the adipose tissue. Our aim was to evaluate whether the quality and quantity of dietary fat can modify that process. A randomized, controlled trial conducted within the LIPGENE study assigned MetS patients to one of four diets for 12 wk each: (i) high-saturated fatty acid (HSFA), (ii) high-monounsaturated fatty acid (HMUFA), (iii) and (iv) two low-fat, high-complex carbohydrate diet supplemented with n-3 polyunsaturated fatty acids (LFHCC n3), or placebo (LFHCC). A fat challenge reflecting the same fatty acid composition as the original diets was conducted post-intervention. The intake of an HSFA meal induced a higher postprandial increase in gp91phox and p67phox mRNA levels than after the intake of HMUFA, LFHCC and LFHCC n-3 meals (all p-values < 0.05). The postprandial decrease in CAT, GPXs and TXNRD1 mRNA levels after the HSFA meal intake was higher than after the intake of HMUFA, LFHCC and LFHCC n-3 meals (all p-values < 0.05). The intake of an HSFA meal induced a higher postprandial increase in KEAP1 mRNA levels than after the consumption of the HMUFA (P = .007) and LFHCC n-3 (P = .001) meals. Our study demonstrated that monounsaturated fat consumption reduces oxidative stress as compared to saturated fat by inducing higher postprandial antioxidant response in adipose tissue, and thus, replacing SFA for MUFA may be an effective dietary strategy to reduce the oxidative stress in MetS patients and its pathophysiological consequences.     

Cardiotrophin-1 is expressed in adipose tissue and upregulated in the metabolic syndrome

Adipose tissue is a target for cardiotrophin-1 (CT-1), a cytokine member of the IL-6 family of cytokines that is involved in cardiac growth and dysfunction. However, it is unknown whether adipocytes are a source of CT-1 and whether CT-1 is overexpressed in diseases characterized by increased fat depots [i.e., the metabolic syndrome (MS)]. Thus this work aimed 1) to test whether adipose tissue expresses CT-1 and whether CT-1 expression can be modulated and 2) to compare serum CT-1 levels in subjects with and without MS diagnosed by National Cholesterol Education Program Adult Treatment Panel III criteria. Gene and protein expression of CT-1 was determined by real-time RT-PCR, ELISA, and Western blotting. CT-1 expression progressively increased, along with differentiation time from preadipocyte to mature adipocyte in 3T3-L1 cells. CT-1 expression was enhanced by glucose in a dose-dependent manner in these cells. mRNA and protein CT-1 expression was also demonstrated in human adipose biopsies. Immunostaining showed positive staining in adipocytes. Finally, increased CT-1 serum levels were observed in patients with MS compared with control subjects (127 +/- 9 vs. 106 +/- 4 ng/ml, P < 0.05). Circulating levels of CT-1 were associated with glucose levels (r = 0.2, P < 0.05). Taken together, our data suggest that adipose tissue can be recognized as a source of CT-1, which could account for the high circulating levels of CT-1 in patients with MS.     

Regulation of adipose tissue energy availability through blood flow control in the metabolic syndrome

Maintenance of blood flow rate is a critical factor for tissue oxygen and substrate supply. The potentially large mass of adipose tissue deeply influences the body distribution of blood flow. This is due to increased peripheral resistance in obesity and the role of this tissue as the ultimate destination of unused excess of dietary energy. However, adipose tissue cannot grow indefinitely, and the tissue must defend itself against the avalanche of nutrients provoking inordinate growth and inflammation. In the obese, large adipose tissue masses show lower blood flow, limiting the access of excess circulating substrates. Blood flow restriction is achieved by vasoconstriction, despite increased production of nitric oxide, the vasodilatation effects of which are overridden by catecholamines (and probably also by angiotensin II and endothelin). Decreased blood flow reduces the availability of oxygen, provoking massive glycolysis (hyperglycemic conditions), which results in the production of lactate, exported to the liver for processing. However, this produces local acidosis, which elicits the rapid dissociation of oxyhemoglobin, freeing bursts of oxygen in localized zones of the tissue. The excess of oxygen (and of nitric oxide) induces the production of reactive oxygen species, which deeply affect the endothelial, blood, and adipose cells, inducing oxidative and nitrosative damage and eliciting an increased immune response, which translates into inflammation. The result of the defense mechanism for adipose tissue, localized vasoconstriction, may thus help develop a more generalized pathologic response within the metabolic syndrome parameters, extending its effects to the whole body.     

Recombinant human alpha-fetoprotein as a regulator of adipose tissue stromal cell activity

Recombinant human alpha-fetoprotein (rhAFP), expressed in the yeast system as a glycoprotein, has been isolated and purified to 98% by the multistep method. The testing of rhAFP in the culture of adipose tissue stromal cells (hASC) has revealed its ability to enhance hASC proliferation and migration, as well as the vascular endothelial growth factor production, with no reliable influence on cell invasion and secretion of matrix metalloproteinases-2 and -9. It has also been established that rhAFP is internalized in hASC via a clathrin-dependent mechanism. The study in the murine experimental model of hindlimb ischemia has shown the ability of rhAFP to enhance blood flow recovery. These data suggest that rhAFP is a promising agent for enhancement of the hASC regenerative ability.     

Xenotransplantation of human fetal adipose tissue: a model of in vivo adipose tissue expansion and adipogenesis

Obesity during childhood and beyond may have its origins during fetal or early postnatal life. At present, there are no suitable in vivo experimental models to study factors that modulate or perturb human fetal white adipose tissue (WAT) expansion, remodeling, development, adipogenesis, angiogenesis, or epigenetics. We have developed such a model. It involves the xenotransplantation of midgestation human WAT into the renal subcapsular space of immunocompromised SCID-beige mice. After an initial latency period of approximately 2 weeks, the tissue begins expanding. The xenografts are healthy and show robust expansion and angiogenesis for at least 2 months following transplantation. Data and cell size and gene expression are consistent with active angiogenesis. The xenografts maintain the expression of genes associated with differentiated adipocyte function. In contrast to the fetal tissue, adult human WAT does not engraft. The long-term viability and phenotypic maintenance of fetal adipose tissue following xenotransplantation may be a function of its autonomous high rates of adipogenesis and angiogenesis. Through the manipulation of the host mice, this model system offers the opportunity to study the mechanisms by which nutrients and other environmental factors affect human adipose tissue development and biology.     

cAMP-positive regulation of angiotensinogen gene expression and protein secretion in rat adipose tissue

The adipose renin-angiotensin system (RAS) has been assigned to participate in the control of adipose tissue development and in the pathogenesis of obesity-related hypertension. In adipose cells, the biological responses to beta-adrenergic stimulation are mediated by an increase in intracellular cAMP. Because cAMP is known to promote adipogenesis and because an association exists between body fat mass, hypertension, and increased sympathetic stimulation, we examined the influence of cAMP on angiotensinogen (ATG) expression and secretion in rat adipose tissue. Exposure of primary cultured differentiated preadipocytes to the cAMP analog 8-bromoadenosine 3',5'-cyclic monophosphate (8-BrcAMP) or cAMP-stimulating agents (forskolin and IBMX) results in a significant increase in ATG mRNA levels. In adipose tissue fragments, 8-BrcAMP also increases ATG mRNA levels and protein secretion, but not in the presence of the protein kinase A inhibitor H89. The addition of isoproterenol, known to stimulate the synthesis of intracellular cAMP via beta-adrenoreceptors, had the same stimulatory effect on ATG expression and secretion. These results indicate that cAMP in vitro upregulates ATG expression and secretion in rat adipose tissue via the protein kinase A-dependent pathway. Further studies are required to determine whether this regulatory pathway is activated in human obesity, where increased sympathetic tone is frequently observed, and to elucidate the importance of adipose ATG to the elevated blood pressure observed in this pathological state.     

Dying for a cause: invertebrate genetics takes on human neurodegeneration

If invertebrate neurons are injured by hostile environments or aberrant proteins they die much like human neurons, indicating that the powerful advantages of invertebrate molecular genetics might be successfully used for testing specific hypotheses about human neurological diseases, for drug discovery and for non-biased screens for suppressors and enhancers of neurodegeneration. Recent molecular dissection of the genetic requirements for hypoxia, excitotoxicity and death in models of Alzheimer disease, polyglutamine-expansion disorders, Parkinson disease and more, is providing mechanistic insights into neurotoxicity and suggesting new therapeutic interventions. An emerging theme is that neuronal crises of distinct origins might converge to disrupt common cellular functions, such as protein folding and turnover.     

Inhibition by potential metabolic inhibitors of in vitro adipose tissue lipogenesis

To study the pathway of lactate utilization as a carbon source for fatty acid synthesis, the effect of (-)-hydroxycitrate, agaric acid, sodium oxamate, 2-n-butyl malonate and alpha-cyano-4-hydroxycinnamate on the rate of in vitro conversion of lactate, acetate and glucose to fatty acids was measured in bovine and rat adipose tissues. Sodium oxamate and hydroxycitrate caused less fatty acid to be synthesized from lactate in bovine adipose tissue. Hydroxycitrate depressed fatty acid synthesis from glucose in rat adipose tissue. alpha-Cyano-4-hydroxycinnamate was an effective inhibitor of lipogenesis from all substrates and may act as a specific inhibitor in adipose tissue. Although the inhibitors were absorbed poorly into adipocytes, the results indicate that conversion of lactate to fatty acids probably occurs by way of the citrate cleavage pathway.     

Nijmegen breakage syndrome 1 protein hyperacetylation as a molecular mechanism underlying metabolic syndrome

Metabolic Syndrome (MetS) is characterized by visceral obesity, insulin resistance, hypertension, atherogenic dyslipidemia, and increased atherosclerotic plaque development and cardiovascular disease. It arises from a high-calorie “Western diet” and physical inactivity. MetS confers an elevated risk for type II diabetes, cancer, and cardiovascular disease, significantly shortening the affected individual's life. While many gene products affect the course of MetS, SirT1 and ataxia–telangiectasia mutated (ATM) protein activities ameliorate the pathophysiological effects of MetS in rodent models. SirT1 activity protects mice from the deleterious effects of a high-fat diet, promoting insulin sensitivity, fat mobilization, lowered blood pressure, and cell survival and genomic stability. ATM activation attenuates hypertension, diet-induced atherosclerotic plaque development, and glucose resistance in mice. ATM activity partially depends on Nijmegen breakage syndrome 1 protein (NBS1) activity. NBS1 can be acetylated, which inhibits its interactions with ATM, attenuating ATM function. Restoration of ATM activity requires NBS1 deacetylation by Sirt1. Interestingly, ATM activation increases SirT1 expression. Several studies show that a high-fat–sugar/high-calorie diet suppresses SirT1 expression in many tissues. Here we hypothesize that SirT1 suppression increases NBS1 acetylation, suppressing ATM activity, and finally attenuating ATM-mediated SirT1 expression. The resulting viscous cycle would promote MetS.     

Mitochondria of adult human brown adipose tissue contain a 32 000-Mr uncoupling protein

Using both immunohistological study and photoaffinity labelling with radioactive azido-ATP, evidence is presented that the mitochondrial membranes of the brown adipose tissue of the human adult contain a 32 000-Mr uncoupling protein, which is probably similar to the uncoupling protein of BAT of rodents.     

A specific l-tri-iodothyronine-binding protein in the cytosol fraction of human breast adipose tissue

1. Binding of l-tri-[(125)I]iodothyronine to the cytosol fraction of normal human female breast adipose tissue was investigated by the charcoal adsorption method. Equilibrium of binding was reached after 120s at 25 degrees C. 2. The l-tri-[(125)I]iodothyronine-binding component is a protein; this was confirmed by experiments in which binding was totally lost after heating the cytosol fraction for 10min at 100 degrees C and in which binding was diminished after treatment with proteolytic enzymes and with thiol-group-blocking reagents. The binding protein was stable at -38 degrees C for several months. 3. It displayed saturability, high affinity (apparent K(d) 3.28nm) and a single class of binding sites. 4. High specificity for l-tri-iodothyronine and l-3,5-di-iodo-3'-isopropylthyronine was observed, whereas other iodothyronines were less effective in displacing l-tri-[(125)I]-iodothyronine from its binding site. 5. The binding of the hormone by the cytosol fraction did not show a pH optimum. 6. When cytosol fractions of adipose tissue from different females were subjected to radioimmunoassay for the determination of thyroxine-binding globulin a value of 0.304+/-0.11mug/mg of cytosol protein (mean+/-s.d., n=4) was obtained; the mean concentration in plasma was 0.309+/-0.07mug/mg of plasma protein (mean+/-s.d., n=3). 7. The K(a) value of 6.3x10(8)m(-1) of l-tri-[(125)I]iodothyronine for binding to plasma, the similar thermalinactivation profiles of binding and the reactivity to thiol-group-blocking reagents were some properties common between the binding components from the cytosol fraction and plasma. 8. These results suggest that the cytosol fraction of human female breast adipose tissue contains thyroxine-binding globulin; the protein that binds l-tri-[(125)I]iodothyronine with high affinity and specificity appears to be similar to thyroxine-binding globulin.