Jaswinder K. Sethi

TNF‐α and adipocyte biology

Dyslipidemia and insulin resistance are commonly associated with catabolic or lipodystrophic conditions (such as cancer and sepsis) and with pathological states of nutritional overload (such as obesity‐related type 2 diabetes). Two common features of these metabolic disorders are adipose tissue dysfunction and elevated levels of tumour necrosis factor‐alpha (TNF‐α). Herein, we review the multiple actions of this pro‐inflammatory adipokine on adipose tissue biology. These include inhibition of carbohydrate metabolism, lipogenesis, adipogenesis and thermogenesis and stimulation of lipolysis. TNF‐α can also impact the endocrine functions of adipose tissue. Taken together, TNF‐α contributes to metabolic dysregulation by impairing both adipose tissue function and its ability to store excess fuel. The molecular mechanisms that underlie these actions are discussed.

Thematic review series: adipocyte biology. Adipose tissue function and plasticity orchestrate nutritional adaptation.

This review focuses on adipose tissue biology and introduces the concept of adipose tissue plasticity and expandability as key determinants of obesity-associated metabolic dysregulation. This concept is fundamental to our understanding of adipose tissue as a dynamic organ at the center of nutritional adaptation. Here, we summarize the current knowledge of the mechanisms by which adipose tissue can affect peripheral energy homeostasis, particularly in the context of overnutrition. Two mechanisms emerge that provide a molecular understanding for obesity-associated insulin resistance. These are a) the dysregulation of adipose tissue expandability and b) the abnormal production of adipokines. This knowledge has the potential to pave the way for novel therapeutic concepts and strategies for managing and/or correcting complications associated with obesity and the metabolic syndrome.

Adipogenesis and WNT signalling.

An inability of adipose tissue to expand consequent to exhausted capacity to recruit new adipocytes might underlie the association between obesity and insulin resistance. Adipocytes arise from mesenchymal precursors whose commitment and differentiation along the adipocytic lineage is tightly regulated. These regulatory factors mediate cross-talk between adipose cells, ensuring that adipocyte growth and differentiation are coupled to energy storage demands. The WNT family of autocrine and paracrine growth factors regulates adult tissue maintenance and remodelling and, consequently, is well suited to mediate adipose cell communication. Indeed, several recent reports, summarized in this review, implicate WNT signalling in regulating adipogenesis. Manipulating the WNT pathway to alter adipose cellular makeup, therefore, constitutes an attractive drug-development target to combat obesity-associated metabolic complications.

Pharmacological Inhibition of Glucosylceramide Synthase Enhances Insulin Sensitivity

A growing body of evidence implicates ceramide and/or its glycosphingolipid metabolites in the pathogenesis of insulin resistance. We have developed a highly specific small molecule inhibitor of glucosylceramide synthase, an enzyme that catalyzes a necessary step in the conversion of ceramide to glycosphingolipids. In cultured 3T3-L1 adipocytes, the iminosugar derivative N-(5′-adamantane-1′-yl-methoxy)-pentyl-1-deoxynojirimycin (AMP-DNM) counteracted tumor necrosis factor-α–induced abnormalities in glycosphingolipid concentrations and concomitantly reversed abnormalities in insulin signal transduction. When administered to mice and rats, AMP-DNM significantly reduced glycosphingolipid but not ceramide concentrations in various tissues. Treatment of ob/ob mice with AMP-DNM normalized their elevated tissue glucosylceramide levels, markedly lowered circulating glucose levels, improved oral glucose tolerance, reduced A1C, and improved insulin sensitivity in muscle and liver. Similarly beneficial metabolic effects were seen in high fat–fed mice and ZDF rats. These findings provide further evidence that glycosphingolipid metabolites of ceramide may be involved in mediating the link between obesity and insulin resistance and that interference with glycosphingolipid biosynthesis might present a novel approach to the therapy of states of impaired insulin action such as type 2 diabetes.

Differential Lipid Partitioning Between Adipocytes and Tissue Macrophages Modulates Macrophage Lipotoxicity and M2/M1 Polarization in Obese Mice

OBJECTIVE Obesity-associated insulin resistance is characterized by a state of chronic, low-grade inflammation that is associated with the accumulation of M1 proinflammatory macrophages in adipose tissue. Although different evidence explains the mechanisms linking the expansion of adipose tissue and adipose tissue macrophage (ATM) polarization, in the current study we investigated the concept of lipid-induced toxicity as the pathogenic link that could explain the trigger of this response. RESEARCH DESIGN AND METHODS We addressed this question using isolated ATMs and adipocytes from genetic and diet-induced murine models of obesity. Through transcriptomic and lipidomic analysis, we created a model integrating transcript and lipid species networks simultaneously occurring in adipocytes and ATMs and their reversibility by thiazolidinedione treatment. RESULTS We show that polarization of ATMs is associated with lipid accumulation and the consequent formation of foam cell–like cells in adipose tissue. Our study reveals that early stages of adipose tissue expansion are characterized by M2-polarized ATMs and that progressive lipid accumulation within ATMs heralds the M1 polarization, a macrophage phenotype associated with severe obesity and insulin resistance. Furthermore, rosiglitazone treatment, which promotes redistribution of lipids toward adipocytes and extends the M2 ATM polarization state, prevents the lipid alterations associated with M1 ATM polarization. CONCLUSIONS Our data indicate that the M1 ATM polarization in obesity might be a macrophage-specific manifestation of a more general lipotoxic pathogenic mechanism. This indicates that strategies to optimize fat deposition and repartitioning toward adipocytes might improve insulin sensitivity by preventing ATM lipotoxicity and M1 polarization.

Tumour necrosis factor- α inhibits adipogenesis via a β -catenin/TCF4(TCF7L2)-dependent pathway

Tumour necrosis factor-α (TNF-α), a proinflammatory cytokine, is a potent negative regulator of adipocyte differentiation. However, the mechanism of TNF-α-mediated antiadipogenesis remains incompletely understood. In this study, we first confirm that TNF-α inhibits adipogenesis of 3T3-L1 preadipocytes by preventing the early induction of the adipogenic transcription factors peroxisome proliferator-activated receptor-γ (PPARγ) and CCAAT/enhancer binding protein-α (C/EBPα). This suppression coincides with enhanced expression of several reported mediators of antiadipogenesis that are also targets of the Wnt/β-catenin/T-cell factor 4 (TCF4) pathway. Indeed, we found that TNF-α enhanced TCF4-dependent transcriptional activity during early antiadipogenesis, and promoted the stabilisation of β-catenin throughout antiadipogenesis. We analysed the effect of TNF-α on adipogenesis in 3T3-L1 cells in which β-catenin/TCF signalling was impaired, either via stable knockdown of β-catenin, or by overexpression of dominant-negative TCF4 (dnTCF4). The knockdown of β-catenin enhanced the adipogenic potential of 3T3-L1 preadipocytes and attenuated TNF-α-induced antiadipogenesis. However, β-catenin knockdown also promoted TNF-α-induced apoptosis in these cells. In contrast, overexpression of dnTCF4 prevented TNF-α-induced antiadipogenesis but showed no apparent effect on cell survival. Finally, we show that TNF-α-induced antiadipogenesis and stabilisation of β-catenin requires a functional death domain of TNF-α receptor 1 (TNFR1). Taken together these data suggest that TNFR1-mediated death domain signals can inhibit adipogenesis via a β-catenin/TCF4-dependent pathway.

IGF-Binding Protein-2 Protects Against the Development of Obesity and Insulin Resistance

Proliferation of adipocyte precursors and their differentiation into mature adipocytes contributes to the development of obesity in mammals. IGF-I is a potent mitogen and important stimulus for adipocyte differentiation. The biological actions of IGFs are closely regulated by a family of IGF-binding proteins (IGFBPs), which exert predominantly inhibitory effects. IGFBP-2 is the principal binding protein secreted by differentiating white preadipocytes, suggesting a potential role in the development of obesity. We have generated transgenic mice overexpressing human IGFBP-2 under the control of its native promoter, and we show that overexpression of IGFBP-2 is associated with reduced susceptibility to obesity and improved insulin sensitivity. Whereas wild-type littermates developed glucose intolerance and increased blood pressure with aging, mice overexpressing IGFBP-2 were protected. Furthermore, when fed a high-fat/high-energy diet, IGFBP-2–overexpressing mice were resistant to the development of obesity and insulin resistance. This lean phenotype was associated with decreased leptin levels, increased glucose sensitivity, and lower blood pressure compared with wild-type animals consuming similar amounts of high-fat diet. Our in vitro data suggest a direct effect of IGFBP-2 preventing adipogenesis as indicated by the ability of recombinant IGFBP-2 to impair 3T3-L1 differentiation. These findings suggest an important, novel role for IGFBP-2 in obesity prevention.

Transmembrane tumor necrosis factor (TNF)-alpha inhibits adipocyte differentiation by selectively activating TNF receptor 1.

Tumor necrosis factor α (TNFα) is a potent cytokine with multiple biological activities and exists in two forms as follows: a 17-kDa soluble form that is a cleaved product of the 26-kDa transmembrane form (mTNFα). It has been suggested that the transmembrane form of TNFα is mainly responsible for localized responses via cell-cell contact. Here, we have examined the activities of transmembrane TNFα in cultured adipocytes. A non-cleavable transmembrane form of TNFα (mTNFΔ1–9K11E) was expressed in several preadipocyte cell lines using retroviral gene transfer. In wild type preadipocytes carrying both TNF receptors, expression of mTNFΔ1–9K11E resulted in inhibition of the differentiation program. The extent of this varied depending on the nature and strength of the adipogenic stimuli. The TNF receptor responsible for this function was determined by expressing mTNFΔ1–9K11E in preadipocyte cell lines lacking either TNF receptor 1 (TNFR1), 2 (TNFR2), or both. In order to confirm the results in the same cellular background, TNF receptors were also reconstituted in the cell lines lacking corresponding receptors. These experiments demonstrated that TNFR1 was necessary and sufficient for mediating mTNFΔ1–9K11E-induced inhibition of adipogenesis and that this action was similar to that of soluble TNFα. In conclusion, our results indicate that mTNFΔ1–9K11E is biologically active in cultured adipocytes and can alter the adipogenic program of these cells by selectively activating TNFR1. This may have physiological implications where local TNFα actions are thought to be generated at sites such as adipose tissue.

The Wnt antagonist Dickkopf-1 and its receptors are coordinately regulated during early human adipogenesis

Secretion of Wnts by adipose cells has an important role in the control of murine adipogenesis. We present the first evidence that a Wnt antagonist, Dickkopf 1 (Dkk1), is secreted by human preadipocytes and promotes adipogenesis. DKK1 mRNA increases six hours after onset of human adipogenesis and this is followed by an increase in Dkk1 protein. With further differentiation, the mRNA and protein levels progressively decline such that they are undetectable in mature adipocytes. The transient induction in DKK1 correlates with downregulation of cytoplasmic and nuclear β-catenin levels, this being a surrogate marker of canonical Wnt signalling, and Wnt/β-catenin transcriptional activity. In addition, constitutive expression of Dkk1 in 3T3-L1 preadipocytes promotes their differentiation, further supporting the functional significance of increased Dkk1 levels during human adipogenesis. Concomitant downregulation of the Dkk1 receptors LRP5 and LRP6 is likely to potentiate the ability of Dkk1 to inhibit Wnt signalling and promote differentiation. Notably, Dkk1 is not expressed in primary murine preadipocytes or cell lines. The involvement of Dkk1 in human but not murine adipogenesis indicates that inter-species differences exist in the molecular control of this process. Given the public health importance of disorders of adipose mass, further knowledge of the pathways involved specifically in human adipocyte differentiation might ultimately be of clinical relevance.

11β-Hydroxysteroid Dehydrogenase Type 1 Regulates Glucocorticoid-Induced Insulin Resistance in Skeletal Muscle

OBJECTIVE Glucocorticoid excess is characterized by increased adiposity, skeletal myopathy, and insulin resistance, but the precise molecular mechanisms are unknown. Within skeletal muscle, 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) converts cortisone (11-dehydrocorticosterone in rodents) to active cortisol (corticosterone in rodents). We aimed to determine the mechanisms underpinning glucocorticoid-induced insulin resistance in skeletal muscle and indentify how 11β-HSD1 inhibitors improve insulin sensitivity. RESEARCH DESIGN AND METHODS Rodent and human cell cultures, whole-tissue explants, and animal models were used to determine the impact of glucocorticoids and selective 11β-HSD1 inhibition upon insulin signaling and action. RESULTS Dexamethasone decreased insulin-stimulated glucose uptake, decreased IRS1 mRNA and protein expression, and increased inactivating pSer307 insulin receptor substrate (IRS)-1. 11β-HSD1 activity and expression were observed in human and rodent myotubes and muscle explants. Activity was predominantly oxo-reductase, generating active glucocorticoid. A1 (selective 11β-HSD1 inhibitor) abolished enzyme activity and blocked the increase in pSer307 IRS1 and reduction in total IRS1 protein after treatment with 11DHC but not corticosterone. In C57Bl6/J mice, the selective 11β-HSD1 inhibitor, A2, decreased fasting blood glucose levels and improved insulin sensitivity. In KK mice treated with A2, skeletal muscle pSer307 IRS1 decreased and pThr308 Akt/PKB increased. In addition, A2 decreased both lipogenic and lipolytic gene expression. CONCLUSIONS Prereceptor facilitation of glucocorticoid action via 11β-HSD1 increases pSer307 IRS1 and may be crucial in mediating insulin resistance in skeletal muscle. Selective 11β-HSD1 inhibition decreases pSer307 IRS1, increases pThr308 Akt/PKB, and decreases lipogenic and lipolytic gene expression that may represent an important mechanism underpinning their insulin-sensitizing action.