Human Subcutaneous Adipose Tissue Research Articles

Adipose-Derived Stem Cells for Future Regenerative System Medicine

BACKGROUND: The potential use of stem cell-based therapies for repair and regeneration of various tissues and organs offers a paradigm shift that may provide alternative therapeutic solutions for a number of disease. Despite the advances, the availability of stem cells remaining a challenge for both scientist and clinicians in pursuing regenerative medicine. CONTENT: Subcutaneous human adipose tissue is an abundant and accessible cell source for applications in tissue engineering and regenerative medicine. Routinely, the adipose issue is digested with collagenase or related lytic enzymes to release a heterogeneous population for stromal vascular fraction (SVF) cells. The SVF cells can be used directly or can be cultured in plastic ware for selection and expansion of an adherent population known as adipose-derived stromal/stem cells (ASCs). Their potential in the ability to differentiate into adipogenic, osteogenic, chondrogenic and other mesenchymal lineages, as well in their other clinically useful properties, includes stimulation of angiogenesis and suppression of inflammation.SUMMARY: Adipose tissue is now recognized as an accessible, abundant and reliable site for the isolation of adult stem cels suitable for the application of tissue engineering and regenerative medicine applications. The past decade has witnessed an explosion of preclinical data relating to the isolation, characterization, cryopreservation, differentiation, and transplantation of freshly isolated stromal vascular fraction cells and adherent, culture-expanded, adipose-derived stromal/stem cells in vitro and in animal models.KEYWORDS: adipose tissue, adult stem cells, regenerative medicine, mesenchymal stem cells

Identification and Functional Characterization of Protein Kinase A Phosphorylation Sites in the Major Lipolytic Protein, Adipose Triglyceride Lipase

Catecholamine-stimulated lipolysis occurs by activating adenylate cyclase and raising cAMP levels, thereby increasing protein kinase A (PKA) activity. This results in phosphorylation and modulated activity of several key lipolytic proteins. Adipose triglyceride lipase (ATGL) is the primary lipase for the initial step in triacylglycerol hydrolysis, and ATGL activity is increased during stimulated lipolysis. Here, we demonstrate that murine ATGL is phosphorylated by PKA at several serine residues in vitro and identify Ser(406) as a functionally important site. ATGL null adipocytes expressing ATGL S406A (nonphosphorylatable) had reduced stimulated lipolysis. Studies in mice demonstrated increased ATGL Ser(406) phosphorylation during fasting and moderate intensity exercise, conditions associated with elevated lipolytic rates. ATGL Ser(404) (corresponding to murine Ser(406)) phosphorylation was increased by β-adrenergic stimulation but not 5'AMP-activated protein kinase activation in human subcutaneous adipose tissue explants, which correlated with lipolysis rates. Our studies suggest that β-adrenergic activation can result in PKA-mediated phosphorylation of ATGL Ser(406), to moderately increase ATGL-mediated lipolysis.

Using Gene Expression to Predict Differences in the Secretome of Human Omental vs. Subcutaneous Adipose Tissue

The objective of this study was to characterize differences in the secretome of human omental compared with subcutaneous adipose tissue using global gene expression profiling. Gene expression was measured using Affymetrix microarrays (Affymetrix, Santa Clara, CA) in subcutaneous and omental adipose tissue in two independent experiments (n = 5 and n = 3 independent subjects; n = 16 arrays in total, 2 for each subject). Predictive bioinformatic algorithms were employed to identify secreted proteins. Microarray analysis identified 22 gene probe sets whose expression was significantly different with a fold change (FC) greater than 5 in expression in both experiments between omental and subcutaneous adipose tissue. Using bioinformatic predictive programs 11 of these 22 probe sets potentially coded for secreted proteins. Pathway network analysis of the secreted proteins showed that three of the proteins are part of a common pathway network. These proteins gremlin 1 (GREM1), pleiotrophin (PTN), and secretory leukocyte peptidase inhibitor (SLPI) are expressed respectively 43×, 23×, and 5× in omental adipose tissue relative to subcutaneous adipose tissue as determined by real-time PCR. The presence of GREM1, PTN, and SLPI protein in human adipose tissue was confirmed by western blotting. All three proteins are expressed in the human Simpson-Golabi-Behmel syndrome (SGBS) preadipocyte cell line. The expression of GREM1, PTN, and SLPI changed with the differentiation of the preadipocytes into mature adipocytes. Gene expression coupled with predictive bioinformatic algorithms have identified several genes coding for secreted proteins which are expressed differently in omental adipose tissue compared to subcutaneous adipose tissue proving a valid alternative approach to help further define the adipocyte secretome.

Depot-specific expression of caveolin-1 in human adipose tissue and their relationship with obesity and insulin resistance

To explore the depot-specific mRNA and protein expression of caveolin-1 (CAV-1) in human subcutaneous and omental adipose tissues and analyze their relationship with obesity and insulin resistance. A total of 41 subjects with normal glucose regulation were recruited. Among them, there were 29 cases with normal body mass index (BMI) and 12 cases with BMI ≥ 24 kg/m(2). All were scheduled to undergone selective abdominal operations. The levels of fasting insulin (FINS) were measured by chemiluminescence enzyme-linked immunosorbent assay (ELISA) kit. Fasting plasma glucose (FPG) was tested by glucose oxidase and home model insulin resistance index (HOMA-IR) calculated. Real-time polymerase chain reaction (RT-PCR) and Western blotting were employed to assess the relative mRNA and protein expression of caveolin-1 in subcutaneous and omental adipose tissues. And the mRNA and protein expression of caveolin-1 from different fat depots were compared and their correlations with BMI and HOMA-IR analyzed. (1) FINS and HOMA-IR were significantly higher in the overweight and obesity group than those in the normal BMI group (FINS: (8.82 ± 3.79) mU/L vs (6.43 ± 4.38) mU/L, P < 0.05, HOMA-1R: 1.91 ± 0.85 vs 1.36 ± 0.72, P < 0.05). (2) The normal BMI group patients had the higher expression levels of caveolin-1 mRNA in omental adipose tissue than overweight counterparts (2.84 ± 0.86 vs 1.02 ± 0.36, P < 0.01). But the difference in subcutaneous adipose tissue was not significant (P > 0.05). (3) The caveolin-1 protein expression in omental adipose tissue of the normal BMI group was higher than that of overweight patients (1.68 ± 0.67 vs 0.73 ± 0.29, P < 0.05). And the difference between two groups was not significant (P > 0.05). (4)The expressions of caveolin-1 mRNA in omental adipose tissue were negatively correlated with BMI, waist circumstance, triglyceride and HOMA-IR (r = -0.441, -0.615, -0.539, -0.688, P < 0.05). No correlations were found between the expressions of caveolin-1 mRNA in subcutaneous adipose tissue with BMI, waist circumstance and HOMA-IR (P > 0.05). Differential depot-specific expressions of caveolin-1 are present in human subcutaneous and omental adipose tissues. A low expression of caveolin-1 in omental adipose tissue may contribute to the pathogenesis of obesity and insulin resistance.

Recycling between cortisol and cortisone in human splanchnic, subcutaneous adipose, and skeletal muscle tissues in vivo.

11β-Hydroxysteroid dehydrogenase type 1 (11βHSD1) is a therapeutic target in metabolic syndrome because it catalyses reductase regeneration of cortisol from cortisone in adipose and liver. 11βHSD1 can also catalyze the reverse dehydrogenase reaction in vitro (e.g., if cofactor is limited). We used stable isotope tracers to test the hypothesis that both 11βHSD1-reductase and -dehydrogenase activities occur in human metabolic tissues in vivo. 1,2-[2H]2-Cortisone (d2-cortisone) was validated as a tracer for 11β-dehydrogenase activity and its inhibition by licorice. d2-Cortisone and 9,11,12,12-[2H]4-cortisol (d4-cortisol) (to measure 11β-reductase activity) were coinfused and venous samples obtained from skeletal muscle, subcutaneous adipose (n = 6), and liver (n = 4). Steroids were measured by liquid chromatography–tandem mass spectrometry and arteriovenous differences adjusted for blood flow. Data are means ± SEM. 11β-Reductase and -dehydrogenase activities were detected in muscle (cortisol release 19.7 ± 4.1 pmol/100 mL/min, d3-cortisol 5.9 ± 1.8 pmol/100 mL/min, and cortisone 15.2 ± 5.8 pmol/100 mL/min) and splanchnic (cortisol 64.0 ± 11.4 nmol/min, d3-cortisol 12.9 ± 2.1 nmol/min, and cortisone 19.5 ± 2.8 nmol/min) circulations. In adipose, dehydrogenase was more readily detected than reductase (cortisone release 38.7 ± 5.8 pmol/100 g/min). Active recycling between cortisol and cortisone in metabolic tissues in vivo may facilitate dynamic control of intracellular cortisol but makes consequences of dysregulation of 11βHSD1 transcription in obesity and diabetes unpredictable. Disappointing efficacy of 11βHSD1 inhibitors in phase II studies could be explained by lack of selectivity for 11β-reductase.

Abstract 273: Phenotypic Differences Between Human Subcutaneous and Perivascular Adipose Tissues

Objective: Human adipose lies in specific anatomic depots. Peri-vascular adipose appears unique and impacts blood vessel biology, but direct human data is limited. In view of their theorized roles, we tested the hypothesis that human peri-vascular and subcutaneous adipose tissue hold distinct phenotypic signatures. We also evaluated the impact of clinical parameters on adipose phenotype. Methods: Fresh human peri-vascular and subcutaneous adipose tissues were collected intra-operatively from patients undergoing major lower extremity amputation (n=27) and assayed for protein levels of the adipose-associated mediators IL-6, IL-8, leptin, TNF-α, MCP-1, adiponectin, resistin and PAI-1. Results: Leptin (2.4 fold, p=0.045) and adiponectin (1.8 fold, p=0.007) were significantly more abundant in subcutaneous compared to peri-vascular adipose (Table I). Clinical data were used to model associations between clinical characteristics and mediator levels. Age positively correlated with peri-vascular PAI-1 expression (β=0.64, p=0.042), and hyperlipidemia negatively correlated with peri-vascular adipose adiponectin (β=-1.18, p=0.039). Conclusions: While significant similarity was observed between subcutaneous and peri-vascular adipose from leg amputation patients, the differential in adipose derived hormones leptin and adiponectin provides direct human evidence that these tissue compartments hold distinct biologic roles. Further investigation into the unique nature of peri-vascular adipose may provide meaningful guidance in managing peripheral vascular disease.

Comparison of 3 T MRI and CT for the measurement of visceral and subcutaneous adipose tissue in humans

CT is considered the gold standard imaging modality for measurement of visceral adipose tissue area. However, as CT imaging exposes subjects to ionising radiation, a comparable imaging technique without this exposure is desirable, such as MRI. Therefore, we compared the agreement of measures of visceral adipose tissue and subcutaneous adipose tissue area from single-slice images obtained at the umbilicus using a 3 T MRI scanner with single-slice images obtained via CT scan. 64 images were obtained from 27 subjects who underwent MRI and CT scanning on the same day, after 10-12 hours of fasting. Visceral and subcutaneous adipose tissue depots were manually separated and quantified using a multimodality image-processing software program. We found good agreement between CT and MRI for the measurement of both visceral adipose tissue and subcutaneous adipose tissue. Bland-Altman difference analysis demonstrated a mean bias of -2.9% (as a portion of total abdominal area) for visceral adipose tissue and +0.4% for subcutaneous adipose tissue, as measured by MRI compared with CT. MRI is a safe, accurate and precise imaging modality for measuring both visceral and subcutaneous adipose tissue, making it a favourable alternative to CT for quantification of these adipose depots.

Expression of Clock Genes in Human Subcutaneous and Visceral Adipose Tissues

Disrupted circadian rhythms are associated with obesity and metabolic alterations, but little is known about the participation of peripheral circadian clock machinery in these processes. The aim of the present study was to analyze RNA expression of clock genes in subcutaneous (SAT) and visceral (VAT) adipose tissues of male and female subjects in AM (morning) and PM (afternoon) periods, and its interactions with body mass index (BMI). Ninety-one subjects (41 ± 11 yrs of age) presenting a wide range of BMI (21.4 to 48.6 kg/m2) were included. SAT and VAT biopsies were obtained from patients undergoing abdominal surgeries. Clock genes expressions were evaluated by qRT-PCR. The only clock gene that showed higher expression (p < .0001) in SAT in comparison to VAT was PER1 of female (372%) and male (326%) subjects. Different patterns of expression between the AM and PM periods were observed, in particular REV-ERBα, which was reduced (p < .05) at the PM period in SAT and VAT of both women and men (women: ∼53% lower; men: ∼78% lower), whereas CLOCK expression was not altered. Relationships between clock genes were different in SAT vs. VAT. BMI was negatively correlated with SATPER1 (r = −.549; p = .001) and SATPER2 (r = −.613; p = .0001) and positively with VATCLOCK (r = .541; p = .001) and VATBMAL1 (r = .468; p = .007) only in women. These data suggest that the circadian clock machinery of adipose tissue depots differs between female and male subjects, with a sex-specific effect observed for some genes. BMI correlated with clock genes, but at this moment it is not possible to establish the cause-effect relationship. (Author correspondence: mzanquetta@gmail.com)

The novel endocrine disruptor tolylfluanid impairs insulin signaling in primary rodent and human adipocytes through a reduction in insulin receptor substrate-1 levels

Emerging data suggest that environmental endocrine disrupting chemicals may contribute to the pathophysiology of obesity and diabetes. In a prior work, the phenylsulfamide fungicide tolylfluanid (TF) was shown to augment adipocyte differentiation, yet its effects on mature adipocyte metabolism remain unknown. Because of the central role of adipose tissue in global energy regulation, the present study tested the hypothesis that TF modulates insulin action in primary rodent and human adipocytes. Alterations in insulin signaling in primary mammalian adipocytes were determined by the phosphorylation of Akt, a critical insulin signaling intermediate. Treatment of primary murine adipose tissue in vitro with 100nM TF for 48h markedly attenuated acute insulin-stimulated Akt phosphorylation in a strain- and species-independent fashion. Perigonadal, perirenal, and mesenteric fat were all sensitive to TF-induced insulin resistance. A similar TF-induced reduction in insulin-stimulated Akt phosphorylation was observed in primary human subcutaneous adipose tissue. TF treatment led to a potent and specific reduction in insulin receptor substrate-1 (IRS-1) mRNA and protein levels, a key upstream mediator of insulin's diverse metabolic effects. In contrast, insulin receptor-β, phosphatidylinositol 3-kinase, and Akt expression were unchanged, indicating a specific abrogation of insulin signaling. Additionally, TF-treated adipocytes exhibited altered endocrine function with a reduction in both basal and insulin-stimulated leptin secretion. These studies demonstrate that TF induces cellular insulin resistance in primary murine and human adipocytes through a reduction of IRS-1 expression and protein stability, raising concern about the potential for this fungicide to disrupt metabolism and thereby contribute to the pathogenesis of diabetes.

The nicotinic acetylcholine receptor α7 in subcutaneous mature adipocytes: downregulation in human obesity and modulation by diet-induced weight loss

It is known that cholinergic anti-inflammatory reflex regulates inflammation in peripheral tissues. Nicotinic acetylcholine receptors (nAChRs) are mediators of this anti-inflammatory pathway and also non-neuronal cells express functional nAChrs. A role for α7-subtype acetylcholine cholinergic receptor (α7nAChR) in insulin sensitivity improvement has already been shown in rodents both in vivo and in vitro. However, no data are available on α7nAChR expression in human adipocytes. To investigate the expression and protein content of α7nAChR in human subcutaneous adipose tissue (SAT) and in isolated mature adipocytes. A total of 39 SAT biopsy specimens obtained from obese and normal-weight subjects were used to assess α7nAChR messenger RNA levels and to stimulate α7nAChR with a specific agonist and antagonist in vitro. Additional SATs from eight non-diabetic obese subjects were also studied, before and after a 3-month lifestyle intervention. α7nAChR expression was significantly lower in the SAT of obese subjects compared with that of normal-weight subjects. In mature adipocytes isolated from morbidly obese subjects (body mass index > 40 kg m(-2)), α7nAChR expression was 75% lower compared with adipocytes from normal-weight subjects. In adipocytes of obese subjects, α7nAChR was downregulated also at protein level. In eight non-diabetic obese subjects, a lifestyle intervention (3 months of diet and physical activity) induced a significant weight loss and an increase in α7nAChR SAT expression. In vitro stimulation of adipocytes with the specific α7nAChR agonist PNU282987 induced a significant anti-inflammatory effect. Furthermore, a similar downregulation of the inflammatory profile, associated with a significant increase in α7nAChR protein level, was observed after genistein stimulation. These results provide evidence that α7nAChR expression levels are significantly decreased in obese subjects, and that this receptor modulates inflammatory gene expression in human adipocytes. The upregulation of α7nAChR by genistein stimulation opens new insights for the management of low-grade inflammation linked to human obesity.

Changes in Gene Expression Profile in Human Subcutaneous Adipose Tissue during Significant Weight Loss

Objective: To analyze the expression of peroxisome proliferator-activated receptor-γ1 and 2 (PPARγ1 and 2), 11β-hydroxysteroid dehydrogenase type 1 (11βHSD1), and leptin in adipose tissue (AT) of obese women during weight loss following Roux-en-Y gastric bypass (RYGB) and to compare these levels with those obtained in AT of nonobese subjects. Methods: Gene expression was determined by real-time RT-PCR prior to surgery and at 3, 6, and 12 months after RYGB. Results: All obese patients lost weight, reaching a mean BMI of 29.3 ± 1.0 kg/m² at 1 year after surgery (–33.9 ± 1.5% of their initial body weight). In obese subjects leptin and 11βHSD1 were over-expressed, whereas PPARγ1 was expressed at lower levels compared to controls. After surgery, leptin and 11βHSD1 gene expression decreased, whereas PPARγ1 expression increased. At 12 months after RYGB, these 3 genes had reached levels similar to the controls. In contrast, PPARγ2 gene expression was not different between groups and types of tissue and remained unchanged during weight loss. We found a positive correlation between BMI and levels of gene expression of leptin and 11βHSD1. Conclusion: Gene expression of leptin, PPARγ1, and 11βHSD1 in AT is modified in human obesity. This default is completely corrected by RYGB.

Lipid and Gene Interactions during Differentiation of Human Subcutaneous Adipose Tissue Stromal Vascular Cells

Examination of the changes in cellular lipid composition, allows for an understanding in their contribution to adipocyte differentiation in health and disease. In the future the analysis of body lipid composition could offer an additional tool to diagnosing disease with the predictive capacity for prognosis. Exposition of cells to excess of energy substrates causes activation of evolutionarily conserved adaptive mechanisms in endoplasmic reticulum stress. In preadipocytes, metabolic stress, through the activation of endoplasmic reticulum stress connected with modification of lipid composition and lipid droplet formation leads to differentiation. The aim of the study was to examine changes in cellular lipid composition in parallel with gene expression and enzyme activity during preadipocyte differentiation using “omics” results. Differentiation condition lead to activation of lipid droplets related protein expression and lipid droplet formation in preadipocytes. The predominant increase of the phospholipids, plasmalogens and cholesterol amounts during cell differentiation was observed. This was accompanied by stimulation of de novo synthesis of saturated fatty acids incorporated into sphingolipids and with reduction in amount of arachidonic acid. Our results indicated that the changes in lipid composition of differentiating stromal vascular fraction cells changed in parallel with gene expression and matched the functional requirements.

CD105 Protein Depletion Enhances Human Adipose-derived Stromal Cell Osteogenesis through Reduction of Transforming Growth Factor β1 (TGF-β1) Signaling

Clinically available sources of bone for repair and reconstruction are limited by the accessibility of autologous grafts, infectious risks of cadaveric materials, and durability of synthetic substitutes. Cell-based approaches for skeletal regeneration can potentially fill this need, and adipose tissue represents a promising source for development of such therapies. Here, we enriched for an osteogenic subpopulation of cells derived from human subcutaneous adipose tissue utilizing microfluidic-based single cell transcriptional analysis and fluorescence-activated cell sorting (FACS). Statistical analysis of single cell transcriptional profiles demonstrated that low expression of endoglin (CD105) correlated with a subgroup of adipose-derived cells with increased osteogenic gene expression. FACS-sorted CD105(low) cells demonstrated significantly enhanced in vitro osteogenic differentiation and in vivo bone regeneration when compared with either CD105(high) or unsorted cells. Evaluation of the endoglin pathway suggested that enhanced osteogenesis among CD105(low) adipose-derived cells is likely due to identification of a subpopulation with lower TGF-β1/Smad2 signaling. These findings thus highlight a potential avenue to promote osteogenesis in adipose-derived mesenchymal cells for skeletal regeneration.

Adipogenesis of Human Adipose-Derived Stem Cells Within Three-Dimensional Hollow Fiber-Based Bioreactors

To further differentiate adipose-derived stem cells (ASCs) into mature adipocytes and create three-dimensional (3D) adipose tissue in vitro, we applied multicompartment hollow fiber-based bioreactor technology with decentral mass exchange for more physiological substrate gradients and integral oxygenation. We hypothesize that a dynamic 3D perfusion in such a bioreactor will result in longer-term culture of human adipocytes in vitro, thus providing metabolically active tissue serving as a diagnostic model for screening drugs to treat diabetes. ASCs were isolated from discarded human abdominal subcutaneous adipose tissue and then inoculated into dynamic 3D culture bioreactors to undergo adipogenic differentiation. Insulin-stimulated glucose uptake from the medium was assessed with and without TNF-alpha. 3D adipose tissue was generated in the 3D-bioreactors. Immunohistochemical staining indicated that 3D-bioreactor culture displayed multiple mature adipocyte markers with more unilocular morphologies as compared with two-dimensional (2D) cultures. Results of real-time polymerase chain reaction showed 3D-bioreactor treatment had more efficient differentiation in fatty acid-binding protein 4 expression. Repeated insulin stimulation resulted in increased glucose uptake, with a return to baseline between testing. Importantly, TNF-alpha inhibited glucose uptake, an indication of the metabolic activity of the tissue. 3D bioreactors allow more mature adipocyte differentiation of ASCs compared with traditional 2D culture and generate adipose tissue in vitro for up to 2 months. Reproducible metabolic activity of the adipose tissue in the bioreactor was demonstrated, which is potentially useful for drug discovery. We present here, to the best of our knowledge for the first time, the development of a coherent 3D high density fat-like tissue consisting of unilocular structure from primary adipose stem cells in vitro.

Influence of menopause on adipose tissue clock gene genotype and its relationship with metabolic syndrome in morbidly obese women

Menopausal women exhibit a loss of circadian coordination, a process that runs parallel with a redistribution of adipose tissue. However, the specific genetic mechanisms underlying these alterations have not been studied. Thus, the aim of the present study was to determine whether the development of menopause induces an alteration of the genes that control biological rhythms in human subcutaneous (SAT) and visceral (VAT) adipose tissue, and whether changes in clock gene expression are involved in the increased risk of developing metabolic syndrome (MetS), which is frequently associated with menopause. To this end, VAT and SAT biopsies were taken in pre- (n = 7) and postmenopausal (n = 7) women at similar hours in the morning. RNA was extracted, and a microarray analysis was made. Data were confirmed by quantitative real-time polymerase chain reaction. Western blot and immunohistochemical analysis were also performed. When clock gene expression was compared between both groups of women, data in SAT showed that expression of the core clock gene period3 was significantly higher in postmenopausal women, while casein kinase-1δ, E1A-binding protein and cAMP-responsive element were preferentially expressed in the premenopausal group. In VAT, period2 (PER2) and v-myc myelocytomatosis viral oncogene expressions were significantly higher in the postmenopausal group. Western blot analysis indicated that PER2 and PER3 protein expression was also increased in postmenopausal women. In addition, several genes, including PER2, were differentially expressed depending on whether or not the patient met the MetS criteria. We conclude that menopause transition induces several changes in the genotype of the adipose tissue chronobiological machinery related to an increased risk of developing MetS.

Free Fatty Acid Storage in Human Visceral and Subcutaneous Adipose Tissue

OBJECTIVEBecause direct adipose tissue free fatty acid (FFA) storage may contribute to body fat distribution, we measured FFA (palmitate) storage rates and fatty acid (FA) storage enzymes/proteins in omental and abdominal subcutaneous fat.RESEARCH DESIGN AND METHODSElective surgery patients received a bolus of [1-14C]palmitate followed by omental and abdominal subcutaneous fat biopsies to measure direct FFA storage. Long chain acyl-CoA synthetase (ACS) and diacylglycerol acyltransferase activities, CD36, fatty acid-binding protein, and fatty acid transport protein 1 were measured.RESULTSPalmitate tracer storage (dpm/g adipose lipid) and calculated palmitate storage rates were greater in omental than abdominal subcutaneous fat in women (1.2 ± 0.8 vs. 0.7 ± 0.4 μmol ⋅ kg adipose lipid−1 ⋅ min−1, P = 0.005) and men (0.7 ± 0.2 vs. 0.2 ± 0.1, P < 0.001), and both were greater in women than men (P < 0.0001). Abdominal subcutaneous adipose tissue palmitate storage rates correlated with ACS activity (women: r = 0.66, P = 0.001; men: r = 0.70, P = 0.007); in men, CD36 was also independently related to palmitate storage rates. The content/activity of FA storage enzymes/proteins in omental fat was dramatically lower in those with more visceral fat. In women, only omental palmitate storage rates were correlated (r = 0.54, P = 0.03) with ACS activity.CONCLUSIONSSome adipocyte FA storage factors correlate with direct FFA storage, but sex differences in this process in visceral fat do not account for sex differences in visceral fatness. The reduced storage proteins in those with greater visceral fat suggest that the storage factors we measured are not a predominant cause of visceral adipose tissue accumulation.

Proteomic Profiling of Adipose Tissue from Zmpste24−/− Mice, a Model of Lipodystrophy and Premature Aging, Reveals Major Changes in Mitochondrial Function and Vimentin Processing

Lipodystrophy is a major disease involving severe alterations of adipose tissue distribution and metabolism. Mutations in genes encoding the nuclear envelope protein lamin A or its processing enzyme, the metalloproteinase Zmpste24, cause diverse human progeroid syndromes that are commonly characterized by a selective loss of adipose tissue. Similarly to humans, mice deficient in Zmpste24 accumulate prelamin A and display phenotypic features of accelerated aging, including lipodystrophy. Herein, we report the proteome and phosphoproteome of adipose tissue as well as serum metabolome in lipodystrophy by using Zmpste24(-/-) mice as experimental model. We show that Zmpste24 deficiency enhanced lipolysis, fatty acid biogenesis and β-oxidation as well as decreased fatty acid re-esterification, thus pointing to an increased partitioning of fatty acid toward β-oxidation and away from storage that likely underlies the observed size reduction of Zmpste24-null adipocytes. Besides the mitochondrial proteins related to lipid metabolism, other protein networks related to mitochondrial function, including those involved in tricarboxylic acid cycle and oxidative phosphorylation, were up-regulated in Zmpste24(-/-) mice. These results, together with the observation of an increased mitochondrial response to oxidative stress, support the relationship between defective prelamin A processing and mitochondrial dysfunction and highlight the relevance of oxidative damage in lipoatrophy and aging. We also show that absence of Zmpste24 profoundly alters the processing of the cytoskeletal protein vimentin and identify a novel protein dysregulated in lipodystrophy, High-Mobility Group Box-1 Protein. Finally, we found several lipid derivates with important roles in energy balance, such as Lysophosphatidylcholine or 2-arachidonoylglycerol, to be dysregulated in Zmpste24(-/-) serum. Together, our findings in Zmpste24(-/-) mice may be useful to unveil the mechanisms underlying adipose tissue dysfunction and its overall contribution to body homeostasis in progeria and other lipodystrophy syndromes as well as to develop novel strategies to prevent or ameliorate these diseases.

Histone Deacetylase 9 Is a Negative Regulator of Adipogenic Differentiation

Differentiation of preadipocytes into mature adipocytes capable of efficiently storing lipids is an important regulatory mechanism in obesity. Here, we examined the involvement of histone deacetylases (HDACs) and histone acetyltransferases (HATs) in the regulation of adipogenesis. We find that among the various members of the HDAC and HAT families, only HDAC9 exhibited dramatic down-regulation preceding adipogenic differentiation. Preadipocytes from HDAC9 gene knock-out mice exhibited accelerated adipogenic differentiation, whereas HDAC9 overexpression in 3T3-L1 preadipocytes suppressed adipogenic differentiation, demonstrating its direct role as a negative regulator of adipogenesis. HDAC9 expression was higher in visceral as compared with subcutaneous preadipocytes, negatively correlating with their potential to undergo adipogenic differentiation in vitro. HDAC9 localized in the nucleus, and its negative regulation of adipogenesis segregates with the N-terminal nuclear targeting domain, whereas the C-terminal deacetylase domain is dispensable for this function. HDAC9 co-precipitates with USF1 and is recruited with USF1 at the E-box region of the C/EBPα gene promoter in preadipocytes. Upon induction of adipogenic differentiation, HDAC9 is down-regulated, leading to its dissociation from the USF1 complex, whereas p300 HAT is up-regulated to allow its association with USF1 and accumulation at the E-box site of the C/EBPα promoter in differentiated adipocytes. This reciprocal regulation of HDAC9 and p300 HAT in the USF1 complex is associated with increased C/EBPα expression, a master regulator of adipogenic differentiation. These findings provide new insights into mechanisms of adipogenic differentiation and document a critical regulatory role for HDAC9 in adipogenic differentiation through a deacetylase-independent mechanism.

Persistent organic pollutant (POPs) levels in human visceral and subcutaneous adipose tissue on an obese Portuguese population—Metabolic improvement after bariatric surgery versus POPs burden

Persistent organic pollutant (POPs) levels in human visceral and subcutaneous adipose tissue on an obese Portuguese population—Metabolic improvement after bariatric surgery versus POPs burden

Human subcutaneous adipose tissue Glut 4 mRNA expression in obesity and type 2 diabetes

Cellular resistance to insulin caused by reduced glucose transport and metabolism is a primary defect leading to the development of metabolic disease. While the etiology of insulin resistance is multifactorial, reduced insulin action is associated with impaired activity of the glucose transporter GLUT4 in insulin-sensitive tissues. Yet, the role of adipose tissue GLUT4 deregulation in the pathogenesis of insulin resistance, obesity, and diabetes is still unclear. In this study, we assessed the relative GLUT4 level in human subcutaneous adipose tissue from obese, diabetic, and diabetic obese versus control subjects, using a real-time PCR method. GLUT4 mRNA levels were considerably decreased among type 2 diabetic patients compared with those of the controls (P<0.01), whereas no such difference was found between obese and normal-weight controls. Multiple linear regressions analysis in both diabetic non-obese and diabetic obese groups showed a negative correlation between GLUT4 mRNA expression and both markers of obesity or insulin resistance (P<0.01). However, in obese group, GLUT4 was inversely associated only with HOMA-IR (P<0.01). Our findings showed that adipose GLUT4 gene expression changes were more related to insulin resistance and type 2 diabetes rather than to obesity.