Omental and subcutaneous adipose tissue steroid levels in obese men

We examined plasma and fat tissue sex steroid levels in a sample of 28 men aged 24.8-62.2 years (average BMI value of 46.3 +/- 12.7 kg/m(2)). Abdominal adipose tissue biopsies were obtained during general or obesity surgery. Omental and subcutaneous adipose tissue steroid levels were measured by gas chromatography and chemical ionization mass spectrometry after appropriate extraction procedures. BMI and waist circumference were negatively correlated with plasma testosterone (r = -0.49 and -0.50, respectively, p < 0.01) and dihydrotestosterone (r = -0.58 and -0.56, respectively, p < 0.01), and positively associated with estrone levels (r = 0.64 and 0.62, respectively, p < 0.001). Regional differences in adipose tissue steroid levels were observed for dihydrotestosterone (p < 0.005), androstenedione (p < 0.0001) and dehydroepiandrosterone levels (p < 0.05), which were all significantly more concentrated in omental versus subcutaneous fat. Positive significant associations were found between circulating level of a steroid and its concentration in omental and subcutaneous adipose tissue, for estrone (r = 0.72 and 0.57, respectively, p < 0.01), testosterone (r = 0.66 and 0.58, respectively, p < 0.01) and dihydrotestosterone (r = 0.58 and 0.45, respectively, p < 0.05). Positive correlations were observed between plasma dehydroepiandrosterone-sulfate and omental (r = 0.56, p < 0.01) as well as subcutaneous adipose tissue dehydroepiandrosterone level (r = 0.38, p = 0.05). Positive significant associations were found between omental adipocyte responsiveness to positive lipolytic stimuli (isoproterenol, dibutyryl cyclic AMP and forskolin) and plasma or omental fat tissue androgen levels. In conclusion, although plasma androgen or estrogen levels are strong correlates of adipose tissue steroid content both in the omental and subcutaneous fat depots, regional differences may be observed. Androgen concentration differences in omental versus subcutaneous adipose tissue suggest a depot-specific impact of these hormones on adipocyte function and metabolism.     

Vaspin gene expression in human adipose tissue: association with obesity and type 2 diabetes

Recently, vaspin was identified as an adipokine with insulin-sensitizing effects, which is predominantly secreted from visceral adipose tissue in a rat model of type 2 diabetes. In this study, we examined whether vaspin mRNA expression is a marker of visceral obesity and correlates with anthropometric and metabolic parameters in paired samples of visceral and subcutaneous adipose tissue from 196 subjects with a wide range of obesity, body fat distribution, insulin sensitivity, and glucose tolerance. Vaspin mRNA expression was only detectable in 23% of the visceral and in 15% of the subcutaneous (SC) adipose tissue samples. Vaspin mRNA expression was not detectable in lean subjects (BMI<25) and was more frequently detected in patients with type 2 diabetes. No significant correlations were found between visceral vaspin gene expression and visceral fat area or SC vaspin expression. However, visceral vaspin expression significantly correlates with BMI, % body fat, and 2 h OGTT plasma glucose. Subcutaneous vaspin mRNA expression is significantly correlated with WHR, fasting plasma insulin concentration, and glucose infusion rate during steady state of an euglycemic-hyperinsulinemic clamp. Multivariate linear regression analysis revealed % body fat as strongest predictor of visceral vaspin and insulin sensitivity as strongest determinant of SC vaspin mRNA expression. In conclusion, our data indicate that induction of human vaspin mRNA expression in adipose tissue is regulated in a fat depot-specific manner and could be associated with parameters of obesity, insulin resistance, and glucose metabolism.     

Melanoma and subcutaneous adipose tissue: Role of peritumoral adipokines in disease characterization and prognosis

In the last decades, the concept of adipose organ has emerged, giving adipose tissue an active endocrine and immunologic function through the secretion of multiple cytokines and chemokines that seem to be implicated in the development and progression of several cancer, including cutaneous melanoma. In this pilot experimental study, we analyzed the expression in the peritumor subcutaneous adipose tissue of the most significant adipokines involved in the processes of carcinogenesis and metastasis in a population of melanoma patients and in two control groups composed of melanocytic nevi and epidermoid cysts, respectively. We correlated the results obtained with the main disease prognostic factors observing a statistically significant increase in the expression of PAI1, LEP, CXCL1, NAMPT, and TNF-α at the level of the peritumor tissue of the melanoma samples compared to the control groups and a correlation of the same with the histopathological prognostic factor of melanoma. Our preliminary study shows that the overexpression of PAI1, LEP, CXCL1, NAMPT, and TNF-α may contribute to the growth and to the local aggressiveness of cutaneous melanoma. It opens the hypothesis of a direct oncogenic role of subcutaneous adipose tissue and adipokines in the tumorigenesis of melanoma.     

Characterization and comparison of adipose tissue‐derived cells from human subcutaneous and omental adipose tissues

Different fat depots contribute differently to disease and function. These differences may be due to the regional variation in cell types and inherent properties of fat cell progenitors. To address the differences of cell types in the adipose tissue from different depots, the phenotypes of freshly isolated adipose tissue‐derived cells (ATDCs) from subcutaneous (SC) and omental (OM) adipose tissues were compared using flow cytometry. Our results showed that CD31^?CD34⁺CD45^?CD90^‐CD105^?CD146⁺ population, containing vascular smooth muscle cells and pericytes, was specifically defined in the SC adipose tissue while no such population was observed in OM adipose tissue. On the other hand, CD31^?CD34⁺CD45^?CD90^?CD105^?CD146^? population, which is an undefined cell population, were found solely in OM adipose tissue. Overall, the SC adipose tissue contained more ATDCs than OM adipose tissue, while OM adipose tissue contained more blood‐derived cells. Regarding to the inherent properties of fat cell progenitors from the two depots, adipose‐derived stem cells (ADSCs) from SC had higher capacity to differentiate into both adipogenic and osteogenic lineages than those from OM, regardless of that the proliferation rates of ADSCs from both depots were similar. The higher differentiation capacity of ADSCs from SC adipose tissue suggests that SC tissue is more suitable cell source for regenerative medicine than OM adipose tissue. Copyright © 2009 John Wiley & Sons, Ltd.     

Pharmacological and nutritional agents promoting browning of white adipose tissue

The role of brown adipose tissue in the regulation of energy balance and maintenance of body weight is well known in rodents. Recently, interest in this tissue has re-emerged due to the realization of active brown-like adipose tissue in adult humans and inducible brown-like adipocytes in white adipose tissue depots in response to appropriate stimuli (“browning process”). Brown-like adipocytes that appear in white fat depots have been called “brite” (from brown-in-white) or “beige” adipocytes and have characteristics similar to brown adipocytes, in particular the capacity for uncoupled respiration. There is controversy as to the origin of these brite/beige adipocytes, but regardless of this, induction of the browning of white fat represents an attractive potential strategy for the management and treatment of obesity and related complications. Here, the different physiological, pharmacological and dietary determinants that have been linked to white-to-brown fat remodeling and the molecular mechanisms involved are reviewed in detail. In the light of available data, interesting therapeutic perspectives can be expected from the use of specific drugs or food compounds able to induce a program of brown fat differentiation including uncoupling protein 1 expression and enhancing oxidative metabolism in white adipose cells. However, additional research is needed, mainly focused on the physiological relevance of browning and its dietary control, where the use of ferrets and other non-rodent animal models with a more similar adipose tissue organization and metabolism to humans could be of much help. This article is part of a Special Issue entitled Brown and White Fat: From Signaling to Disease.     

Stimulation of PAI-1 and adipokines by glucose in human adipose tissue in vitro

Adipokines such as Plasminogen activator inhibitor-1 (PAI-1), interleukin (IL)-8, and tumor necrosis factor (TNF)-alpha are elevated in patients with obesity, insulin resistance, and type 2 diabetes. In the present study, we investigated whether glucose affected the production of these adipokines in human adipose tissue in vitro. Glucose (up to 35mM) increased secretion of PAI-1 (p<0.01) and IL-8 (p<0.01), but not TNF-alpha, in a dose- and time-dependent manner. Half-maximal stimulatory concentration of glucose was about 1mM. Glucosamine (5mM) decreased production of PAI-1 (p<0.05) and IL-8 (p<0.05), indicating that the hexosamine biosynthesis pathway is not involved in the glucose-induced increment in adipokine secretion. The present data demonstrate that glucose increases PAI-1 and IL-8 secretion. However, glucose concentrations above 5mM had no additional effects on adipokine secretion, suggesting that mechanisms other than diabetes/insulin resistance-related hyperglycemia may be involved in the observed elevation of these adipokines.     

Differential expression of adipose tissue proteins between obesity-susceptible and -resistant rats fed a high-fat diet

One of the major questions in the field of obesity is why some humans become obese (obesity prone, OP) and others resist the development of obesity (obesity resistant, OR) when exposed to a high-calorie diet, which has not been completely studied. Therefore, in the present study, in order to gain insight into the molecular mechanisms underlying this propensity, we have performed a comparative analysis of protein expression profiles in white adipose tissue (WAT) and brown adipose tissue (BAT) of rats fed a high-fat diet by 2-DE and MALDI-TOF-MS. Protein mapping of homogenates revealed significant alterations to a number of proteins; 60 and 70 proteins were differentially regulated in BAT and WAT, respectively. For careful interpretation of proteomic results, we categorized the identified proteins into two groups by analysis of both average spot density of pooled six rat adipose tissues and individual spot density of each adipose tissue of six rats as a function of body weight. One of the most striking findings of this study was that significant changes of Ehd1 and laminin receptor in BAT as well as antiquitin, DJ-1 protein, and paraoxonase 2 in WAT were found for the first time in obese rats. In addition, we confirmed the increased expression of some thermogenic enzymes and decreased lipogenic enzymes in adipose tissues of OR rats by immunoblot analysis. To our knowledge, this is the first proteomic study of profiling of protein modulation in OP and OR rats, thereby providing the first global evidence for different propensities to obesity between OP and OR rats.     

Acute and chronic saturated fatty acid treatment as a key instigator of the TLR-mediated inflammatory response in human adipose tissue, in vitro

A post-prandial increase in saturated fatty acids (SFAs) and glucose (Glc) activates an inflammatory response, which may be prolonged following restoration of physiological SFAs and Glc levels — a finding referred to as ‘metabolic memory'.This study examined chronic and oscillating SFAs and Glc on the inflammatory signalling pathway in human adipose tissue (AT) and adipocytes (Ads) and determined whether Ads are subject to “metabolic memory.”Abdominal (Abd) subcutaneous (Sc) explants and Ads were treated with chronic low glucose (L-Glc): 5.6 mM and high glucose (H-Glc): 17.5 mM, with low (0.2 mM) and high (2 mM) SFA for 48 h. Abd Sc explants and Ads were also exposed to the aforementioned treatment regimen for 12-h periods, with alternating rest periods of 12 h in L-Glc.Chronic treatment with L-Glc and high SFAs, H-Glc and high SFAs up-regulated key factors of the nuclear factor-κB (NFκB) pathway in Abd Sc AT and Ads (TLR4, NFκB; P<.05), whilst down-regulating MyD88. Oscillating Glc and SFA concentrations increased TLR4, NFκB, IKKβ (P<.05) in explants and Ads and up-regulated MyD88 expression (P<.05). Both tumor necrosis factor α and interleukin 6 (P<.05) secretion were markedly increased in chronically treated Abd Sc explants and Ads whilst, with oscillating treatments, a sustained inflammatory effect was noted in absence of treatment.Therefore, SFAs may act as key instigators of the inflammatory response in human AT via NFκB activation, which suggests that short-term exposure of cells to uncontrolled levels of SFAs and Glc leads to a longer-term inflammatory insult within the Ad, which may have important implications for patients with obesity and Type 2 diabetes.     

Osteogenic differentiation of multipotent mesenchymal stromal cells isolated from human bone marrow and subcutaneous adipose tissue

Cellular populations with phenotypes similar to multipotent mesenchymal stromal cells were isolated from two different sources, including human bone marrow (BM) and subcutaneous adipose tissue (SAT). Comparative analysis of the efficiency of differentiation in the direction of osteogenesis has revealed morphological changes confirmed by staining with Alizarin red and von Kossa in bone marrow cells at the 14th day and in adipose tissue cells at the 28th day of cultivation in the medium with inductors. Analysis of expression of the osteopontin, osteocalcin, and bone sialoprotein genes in RT-PCR reactions has detected essential differences in the potential of these cells to differentiate into bone tissue cells. Cells isolated from BM of both the control and experimental groups were positive for octeopontin (OP) on the 14th day. Unlike these cells, in cells isolated from SAT in medium without an inductor, no product of OP gene expression was identified. In the cells subjected to differentiation, OP appeared at day 14. In the BM cells, octeocalcin (OC) was found at the 14th day, while the bone sialoprotein (BS) was found at the 21st day of cultivation in induction medium. In cells isolated from SAT, OC, and BS were not detected, even at the 28th day after the beginning of induction.     

Brown adipose tissue: from thermal physiology to bioenergetics

Brown adipose tissue is an organ in mammals specialized for the generation of heat. The tissue plays an important role in thermoregulatory heat production (nonshivering thermogenesis), and in nutritional energetics (through the process of diet-induced thermogenesis). Much of the current interest in brown adipose tissue has been catalysed by the postulate (1970’s) that a reduced capacity for thermogenesis underlies the development of obesity. Heat is generated in brown fat by a controlled uncoupling of oxidative phosphorylation, a process regulated by a tissue-specific mitochondrial uncoupling protein,M _r 32–33,000. The immunological identification of uncoupling protein is now used as a biochemical criterion for distinguishing brown fat from white adipose tissue. The gene coding for uncoupling protein has been cloned in several species, and a number of factors regulating the expression of the gene, as well as the amount and activity of the protein itself, have been documented. In addition to its direct role in heat production, brown adipose tissue has some notable general metabolic properties, such as in the conversion of thyroxine to triiodothyronine. An overview of the biology of brown adipose tissue is presented in this article, with an emphasis on some recent developments.     

CLA differently regulates adipogenesis in stromal vascular cells from porcine subcutaneous adipose and skeletal muscle

Conjugated linoleic acid (CLA), a mixture of isomers of linoleic acid, has previously been shown to be able to decrease porcine subcutaneous (SC) adipose tissue levels while increasing the count of intramuscular (IM) adipose tissue in vivo. However, the underlying mechanisms through which it acts are poorly understood. The objective of this study was to investigate the different effects of CLA on adipogenesis in cultured SC adipose tissue and IM stromal vascular cells obtained from neonatal pigs. As shown here, trans-10, cis-12 CLA decreased the expression of adipocyte-specific genes as well as adipose precursor cell numbers and the accumulation of lipid in cultured SC adipose tissue stromal vascular cells. However, the cis-9, trans-11 CLA did not alter adipogenesis in SC cultures. On the other hand, both CLA isomers increased the expression of adipocyte-specific genes in IM cultures, together with the increasing accumulation of lipid and Oil Red O-stained cells. Collectively, these data show that CLA decreases SC adipose tissue but increases IM adipose tissue by different regulation of adipocyte-specific gene expression. These results suggest that adipogenesis in IM adipocytes differs from that in SC adipocytes.     

Visualization of sympathetic neural innervation in human white adipose tissue

Obesity, defined as an excess of adipose tissue that adversely affects health, is a major cause of morbidity and mortality. However, to date, understanding the structure and function of human adipose tissue has been limited by the inability to visualize cellular components due to the innate structure of adipocytes, which are characterized by large lipid droplets. Combining the iDISCO and the CUBIC protocols for whole tissue staining and optical clearing, we developed a protocol to enable immunostaining and clearing of human subcutaneous white adipose tissue (WAT) obtained from individuals with severe obesity. We were able to perform immunolabelling of sympathetic nerve terminals in whole WAT and subsequent optical clearing by eliminating lipids to render the opaque tissue completely transparent. We then used light sheet confocal microscopy to visualize sympathetic innervation of human WAT from obese individuals in a three-dimensional manner. We demonstrate the visualization of sympathetic nerve terminals in human WAT. This protocol can be modified to visualize other structures such as blood vessels involved in the development, maintenance and function of human adipose tissue in health and disease.     

GDF1 is a novel mediator of macrophage infiltration in brown adipose tissue of obese mice

We previously demonstrated a marked upregulation in the bone morphogenic protein (BMP)/growth differentiation factor (GDF) family member, GDF5, which is capable of promoting brown adipogenesis, in brown adipose tissue (BAT) of obese mice. In this study, we identified other GDF family members, besides GDF5 that are responsive to different obesogenic signals in BAT using inborn and acquired obesity animal models. In BAT from leptin-deficient ob/ob mice, GDF1 expression was preferentially downregulated, whereas the expression of several other genes in the BMP/GDF family, including GDF5, was upregulated. Moreover, in cultured brown adipocytes exposed to tunicamycin and hydrogen peroxide, at concentrations not affecting cellular viability, GDF1 expression was significantly downregulated. Recombinant GDF1 failed to significantly alter brown adipogenesis, despite the promoted phosphorylation of Smad1/5/8 in cultured brown adipocytes, but accelerated Smad1/5/8 phosphorylation with a concomitant increase in the number of migrating cells during exposure in a manner sensitive to activin-like kinase inhibitors in macrophagic RAW264.7 cells. Similarly, accelerated migration was observed in murine peritoneal macrophages exposed to GDF1. These results indicate that obesity could lead to predominant downregulation of GDF1 expression in BAT, which can modulate cellular migration through a mechanism relevant to activation of the downstream Smad signaling pathway in adjacent macrophages.     

Adipocyte differentiation of multipotent cells established from human adipose tissue

In this study multipotent adipose-derived stem cells isolated from human adipose tissue (hMADS cells) were shown to differentiate into adipose cells in serum-free, chemically defined medium. During the differentiation process, hMADS cells exhibited a gene expression pattern similar to that described for rodent clonal preadipocytes and human primary preadipocytes. Differentiated cells displayed the key features of human adipocytes, i.e., expression of specific molecular markers, lipolytic response to agonists of beta-adrenoreceptors (beta2-AR agonist > beta1-AR agonist > beta3-AR agonist) and to the atrial natriuretic peptide, insulin-stimulated glucose transport, and secretion of leptin and adiponectin. hMADS cells were able to respond to drugs as inhibition of adipocyte differentiation was observed in the presence of prostaglandin F2alpha, tumour necrosis factor-alpha, and nordihydroguaiaretic acid, a natural polyhydroxyphenolic antioxidant. Thus, for the first time, human adipose cells with normal karyotype and indefinite life span have been established. They represent a novel and valuable tool for studies of fat tissue development and metabolism.     

Brown adipose tissue: Updates in cellular and molecular biology

Brown adipose tissue (BAT) is mainly composed of adipocytes, it is highly vascularized and innervated, and can be activated in adult humans. Brown adipocytes are responsible for performing non-shivering thermogenesis, which is exclusively mediated by uncoupling protein (UCP) -1 (a protein found in the inner mitochondrial membrane), the hallmark of BAT, responsible for the uncoupling of the proton leakage from the ATP production, therefore, generating heat (i.e. thermogenesis). Besides UCP1, other compounds are essential not only to thermogenesis, but also to the proliferation and differentiation of BAT, including peroxisome proliferator-activated receptor (PPAR) family, PPARgamma coactivator 1 (PGC1)-alpha, and PRD1-BF-1-RIZ1 homologous domain protein containing protein (PRDM) -16. The sympathetic nervous system centrally regulates thermogenesis through norepinephrine, which acts on the adrenergic receptors of BAT. This bound leads to the initialization of the many pathways that may activate thermogenesis in acute and/or chronic ways. In summary, this mini-review aims to demonstrate the latest advances in the knowledge of BAT.     

Attenuated metabolism is a hallmark of obesity as revealed by comparative proteomic analysis of human omental adipose tissue

Obesity is recognized as an epidemic health problem worldwide. In humans, the accumulation of omental rather than subcutaneous fat appears to be tightly linked to insulin resistance, type 2 diabetes and cardiovascular disease. Differences in gene expression profiles in the adipose tissue comparing non-obese and obese subjects have been well documented. However, to date, no comparative proteomic studies based on omental fat have investigated the influence of obesity in protein expression. In this work, we searched for proteins differentially expressed in the omental fat of non-obese and obese subjects using 2D-DIGE and MS. Forty-four proteins, several of which were further studied by immunoblotting and immunostaining analyses, showed significant differences in the expression levels in the two groups of subjects. Our findings reveal a clearly distinctive proteomic profile between obese and non-obese subjects which emphasizes: i) reduced metabolic activity in the obese fat, since most down-regulated proteins were engaged in metabolic pathways; and ii) morphological and structural cell changes in the obese fat, as revealed by the functions exerted by most up-regulated proteins. Interestingly, transketolase and aminoacylase-1 represent newly described molecules involved in the pathophysiology of obesity, thus opening up new possibilities in the study of obesity.     

Continuous flow microcalorimetric measurement of heat production in white adipose tissue from obese (ob/ob) mice and their lean littermates

Heat production, free fatty acid and glycerol release from white adipose tissue fat pads from obese (ob/ob) mice and their lean littermates are determined. Heat production was significantly lower in obese mice compared to lean mice when expressed on wet weight basis but not when expressed on DNA basis. Noradrenaline significantly increased the heat production in fat pads from both groups of animals. However, the increase in heat production due to noradrenaline addition in fat pads from lean mice was significantly higher than in fat pads from obese mice. The release of free fatty acids and glycerol before incubation with noradrenaline was similar from fat pads from both groups of animals. Addition of noradrenaline to the fat pads increased the release of free fatty acids and glycerol in both groups of animals, but the increase was significantly larger from fat pads from lean mice. In the absence of noradrenaline the free fatty acid/glycerol ratio (mol/mol) in the effluent was 7.9:1 and 4.8:1 for lean mice and obese mice, respectively. In the presence of noradrenaline the ratio decreased to 3:1 for both groups of animals.