The three subtypes of the peroxisome proliferatoractivated receptors (PPAR␣, ␤/␦, and ␥) form heterodimers with the 9-cis-retinoic acid receptor (RXR) and bind to a common consensus response element, which consists of a direct repeat of two hexanucleotides spaced by one nucleotide (DR1). As a first step toward understanding the molecular mechanisms determining PPAR subtype specificity, we evaluated by electrophoretic mobility shift assays the binding properties of the three PPAR subtypes, in association with either RXR␣ or RXR␥, on 16 natural PPAR response elements (PPREs). The main results are as follows. (i) PPAR␥ in combination with either RXR␣ or RXR␥ binds more strongly than PPAR␣ or PPAR␤ to all natural PPREs tested. (ii) The binding of PPAR to strong elements is reinforced if the heterodimerization partner is RXR␥. In contrast, weak elements favor RXR␣ as heterodimerization partner. (iii) The ordering of the 16 natural PPREs from strong to weak elements does not depend on the core DR1 sequence, which has a relatively uniform degree of conservation, but correlates with the number of identities of the 5-flanking nucleotides with respect to a consensus element. This 5-flanking sequence is essential for PPAR␣ binding and thus contributes to subtype specificity. As a demonstration of this, the PPAR␥-specific element ARE6 PPRE is able to bind PPAR␣ only if its 5-flanking region is exchanged with that of the more promiscuous HMG PPRE.

Leptin is thought to exert its actions on energy homeostasis through the long form of the leptin receptor (OB-Rb), which is present in the hypothalamus and in certain peripheral organs, including adipose tissue. In this study, we examined whether leptin has direct effects on the function of brown and white adipose tissue (BAT and WAT, respectively) at the metabolic and molecular levels. The chronic peripheral intravenous administration of leptin in vivo for 4 d resulted in a 1.6-fold increase in the in vivo glucose utilization index of BAT, whereas no significant change was found after intracerebroventricular administration compared with pair-fed control rats, compatible with a direct effect of leptin on BAT. The effect of leptin on WAT fat pads from lean Zucker Fa/ fa rats was assessed ex vivo, where a 9-and 16-fold increase in the rate of lipolysis was observed after 2 h of exposure to 0.1 and 10 nM leptin, respectively. In contrast, no increase in lipolysis was observed in the fat pads from obese fa/fa rats, which harbor an inactivating mutation in the OB-Rb. At the level of gene expression, leptin treatment for 24 h increased malic enzyme and lipoprotein lipase RNA 1.8 Ϯ 0.17 and 1.9 Ϯ 0.14-fold, respectively, while aP2 mRNA levels were unaltered in primary cultures of brown adipocytes from lean Fa/fa rats. Importantly, however, no significant effect of leptin was observed on these genes in brown adipocytes from obese fa/fa animals. The presence of OB-Rb receptors in adipose tissue was substantiated by the detection of its transcripts by RT-PCR, and leptin treatment in vivo and in vitro activated the specific STATs implicated in the signaling pathway of the OB-Rb.Taken together, our data strongly suggest that leptin has direct effects on BAT and WAT, resulting in the activation of the Jak/STAT pathway and the increased expression of certain target genes, which may partially account for the observed increase in glucose utilization and lipolysis in leptintreated adipose tissue. ( J. Clin. Invest. 1997. 100:2858-2864.)

The secreted form of the interleukin-1 receptor antagonist (IL-1Ra) is an acute-phase protein intervening in the counterregulation of inflammatory processes. We previously showed that this cytokine antagonist is upregulated in the serum of obese patients, correlating with BMI and insulin resistance. In this study, we examined the expression pattern of IL-1Ra and showed that it is highly expressed not only in liver and spleen, but also in white adipose tissue (WAT), where it is upregulated in obesity. In WAT of obese humans, IL-1Ra was also markedly increased. Moreover, human WAT explants secreted IL-1Ra into the medium, a process that could be stimulated fivefold by interferon-␤. Finally, lipopolysaccharide administration induced a longlasting expression of IL-1Ra in mouse WAT, suggesting that adipose tissue is an important source of IL-1Ra in both obesity and inflammation. In summary, we demonstrated that WAT is one of the most important sources of IL-1Ra quantitatively, suggesting that this tissue could represent a novel target for anti-inflammatory treatment. Moreover, it can be speculated that IL-1Ra, whose production is markedly increased in WAT in obese individuals, contributes further to weight gain because of its endocrine and paracrine effects on the hypothalamus and adipocytes, respectively. Diabetes

Objective-Obesity is associated with an increased risk for cardiovascular disease. Although it is known that white adipose tissue (WAT) produces numerous proinflammatory and proatherogenic cytokines and chemokines, it is unclear whether adipose-derived chemotactic signals affect the chronic inflammation in atherosclerosis. Methods and Results-Histological examination showed that perivascular WAT (pWAT) is in close proximity to vascular walls, particularly at sites that have a tendency to develop atherosclerosis. In rodents, the amount of pWAT is markedly increased by a high-fat diet. At a functional level, supernatant from subcutaneous and pWAT strongly induced the chemotaxis of peripheral blood leukocytes. The migration of granulocytes and monocytes was mostly mediated by interleukin-8 and monocyte chemoattractant protein-1, respectively, whereas both chemokines contributed to the migration of activated T cells. Moreover, pWAT produces these chemokines, as shown by immunohistochemistry and by explant culture. The accumulation of macrophages and T cells at the interface between pWAT and the adventitia of human atherosclerotic aortas may reflect this prochemotactic activity of pWAT. Conclusions-Human pWAT has chemotactic properties through the secretion of different chemokines, and we propose that pWAT might contribute to the progression of obesity-associated atherosclerosis. (Arterioscler Thromb Vasc Biol. 2005;25:2594-2599.)Key Words: chemokines Ⅲ perivascular adipose tissue Ⅲ chemotaxis Ⅲ obesity Ⅲ inflammation O besity is characterized by an excess of body fat and is associated with an increased risk for cardiovascular disease. 1 Obesity is also linked to a state of chronic inflammation 2 associated with the production of pro-and antiinflammatory cytokines secreted by white adipose tissue (WAT), such as interleukin (IL)-6, tumor necrosis factor (TNF)-␣, IL-1␤, and the IL-1 receptor antagonist. 3 WAT is also known to produce chemokines such as IL-8 (also called CXCL8) 4,5 and monocyte chemoattractant protein-1 (MCP-1, also called CCL2). 5,6 Atherosclerosis is characterized by the accumulation of macrophages and T lymphocytes in the wall of large arteries, a process in which chemokines play an important role. 7,8 Many of the chemoattractant factors involved in the development of atherosclerosis are expressed in the atherosclerotic plaque and produced by endothelial and smooth muscle cells. 8 The importance of chemokines in the development of atherosclerosis has been demonstrated in a low-density lipoprotein (LDL) receptor-deficient mouse model where the invalidation of the MCP-1 gene prevented lipid deposition and macrophage infiltration in the aortic wall. 8 Similarly, irradiated LDL receptor-deficient mice received a transplant of bone marrow cells lacking the CXCR2 (murine IL-8 homologue receptor), resulting in the reduced accumulation of monocytes/macrophages in the vascular intima. 9 Because obesity is a metabolic and chronic inflammatory condition associated with cardiovascular complications, it is tempti...

The peroxisome proliferator-activated receptors (PPARs) are a subgroup of nuclear receptors activated by fatty acids and eicosanoids. In addition, they are subject to phosphorylation by insulin, resulting in the activation of PPAR␣, while inhibiting PPAR␥ under certain conditions. However, it was hitherto unclear whether the stimulatory effect of insulin on PPAR␣ was direct and by which mechanism it occurs. We now demonstrate that amino acids 1-92 of hPPAR␣ contain an activation function (AF)-1-like domain, which is further activated by insulin through a pathway involving the mitogen-activated protein kinases p42 and p44. Further analysis of the amino-terminal region of PPAR␣ revealed that the insulin-induced trans-activation occurs through the phosphorylation of two mitogen-activated protein kinase sites at positions 12 and 21, both of which are conserved across evolution. The characterization of a strong AF-1 region in PPAR␣, stimulating transcription one-fourth as strongly as the viral protein VP16, is compatible with the marked basal transcriptional activity of this isoform in transfection experiments. However, it is intriguing that the activity of this AF-1 region is modulated by the phosphorylation of two serine residues, both of which must be phosphorylated in order to activate transcription. This is in contrast to PPAR␥2, which was previously shown to be phosphorylated at a single site in a motif that is not homologous to the sites now described in PPAR␣. Although the molecular details involved in the phosphorylation-dependent enhancement of the transcriptional activity of PPAR␣ remain to be elucidated, we demonstrate that the effect of insulin on the AF-1 region of PPAR␣ can be mimicked by the addition of triiodothyronine receptor ␤1, a strong binder of corepressor proteins. In addition, a triiodothyronine receptor ␤1 mutant deficient in interacting with corepressors is unable to activate PPAR␣. These observations suggest that the AF-1 region of PPAR␣ is partially silenced by corepressor proteins, which might interact in a phosphorylation-dependent manner.