Peroxisome proliferator-activated receptor-gamma ligands inhibit adipocyte 11beta -hydroxysteroid dehydrogenase type 1 expression and activity

Peroxisome proliferator-activated receptor-gamma (PPARgamma) has been shown to play an important role in the regulation of expression of a subclass of adipocyte genes and to serve as the molecular target of the thiazolidinedione (TZD) and certain non-TZD antidiabetic agents. Hypercorticosteroidism leads to insulin resistance, a variety of metabolic dysfunctions typically seen in diabetes, and hypertrophy of visceral adipose tissue. In adipocytes, the enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD-1) converts inactive cortisone into the active glucocorticoid cortisol and thereby plays an important role in regulating the actions of corticosteroids in adipose tissue. Here, we show that both TZD and non-TZD PPARgamma agonists markedly reduced 11beta-HSD-1 gene expression in 3T3-L1 adipocytes. This diminution correlated with a significant decrease in the ability of the adipocytes to convert cortisone to cortisol. The half-maximal inhibition of 11beta-HSD-1 mRNA expression by the TZD, rosiglitazone, occurred at a concentration that was similar to its K(d) for binding PPARgamma and EC(50) for inducing adipocyte differentiation thereby indicating that this action was PPARgamma-dependent. The time required for the inhibitory action of the TZD was markedly greater for 11beta-HSD-1 gene expression than for leptin, suggesting that these genes may be down-regulated by different molecular mechanisms. Furthermore, whereas regulation of PPARgamma-inducible genes such as phosphoenolpyruvate carboxykinase was maintained when cellular protein synthesis was abrogated, PPARgamma agonist inhibition of 11beta-HSD-1 and leptin gene expression was ablated, thereby supporting the conclusion that PPARgamma affects the down-regulation of 11beta-HSD-1 indirectly. Finally, treatment of diabetic db/db mice with rosiglitazone inhibited expression of 11beta-HSD-1 in adipose tissue. This decrease in enzyme expression correlated with a significant decline in plasma corticosterone levels. In sum, these data indicate that some of the beneficial effects of PPARgamma antidiabetic agents may result, at least in part, from the down-regulation of 11beta-HSD-1 expression in adipose tissue.     

GLUT4 repression in response to oxidative stress is associated with reciprocal alterations in C/EBP alpha and delta isoforms in 3T3-L1 adipocytes

Insulin responsiveness of adipocytes is acquired during normal adipogenesis, and is essential for maintaining whole-body insulin sensitivity. Differentiated adipocytes exposed to oxidative stress become insulin resistant, exhibiting decreased expression of genes like the insulin-responsive glucose transporter GLUT4. Here we assessed the effect of oxidative stress on DNA binding capacity of C/EBP isoforms known to participate in adipocyte differentiation, and determine the relevance for GLUT4 gene regulation. By electrophoretic mobility shift assay, nuclear proteins from oxidized adipocytes exhibited decreased binding of C/EBPalpha-containing dimers to a DNA oligonucleotide harboring the C/EBP binding sequence from the murine GLUT4 promoter. C/EBPdelta-containing dimers were increased, while C/EBPbeta-dimers were unchanged. These alterations were mirrored by a 50% decrease and a 2-fold increase in the protein content of C/EBPalpha and C/EBPdelta, respectively. In oxidized cells, GLUT4 protein and mRNA levels were decreased, and a GLUT4 promoter segment containing the C/EBP binding site partially mediated oxidative stress-induced repression of a reported gene. The antioxidant lipoic acid protected against oxidation-induced decrease in GLUT4 and C/EBPalpha mRNA, but did not prevent the increase in C/EBPdelta mRNA. We propose that oxidative stress induces adipocyte insulin resistance partially by affecting the expression of C/EBPalpha and delta, resulting in altered C/EBP-dimer composition potentially occupying the GLUT4 promoter.     

Dexamethasone reverses TGF-beta-mediated inhibition of primary rat preadipocyte differentiation

Dexamethasone and transforming growth factor-beta (TGF-beta) show contrary effects on differentiation of adipocytes. Dexamethasone stimulates adipocyte differentiation whereas TGF-beta inhibits it. In the present study, we investigated whether dexamethasone could reverse the TGF-beta-mediated inhibition of preadipocyte differentiation. Primary rat preadipocytes, obtained from Sprague-Dawley rats, were pretreated with dexamethasone in the presence or absence of TGF-beta, prior to the induction of differentiation. Co-treatment of dexamethasone and TGF-beta before inducing differentiation reversed the TGF-beta-mediated inhibition of preadipocyte differentiation. In order to elucidate the mechanism by which dexamethasone reversed the effect of TGF-beta on the inhibition of preadipocyte differentiation, the expression of CCAAT/enhancer binding protein-alpha (C/EBPalpha) and peroxisome proliferator-activated receptor gamma (PPARgamma) was examined. Dexamethasone increased C/EBPalpha and PPARgamma expression in the absence of TGF-beta and also recovered the TGF-beta-mediated suppression of C/EBPalpha expression in preadipocytes. Its effect was sustained in differentiated adipocytes as well. However, those effects were not observed in 3T3-L1 preadipocytes or differentiated adipocytes. These results indicate that dexamethasone reverses the TGF-beta-mediated suppression of adipocyte differentiation by regulating the expression of C/EBPalpha and PPARgamma, which is dependent on the cellular context.     

Adipogenic differentiating agents regulate expression of fatty acid binding protein and CD36 in the J744 macrophage cell line

Adipocyte fatty acid binding protein (aP2) is a key mediator of intracellular transport and metabolism of fatty acids. Its expression during adipocyte differentiation is regulated through the actions of peroxisome proliferator-activated receptor gamma (PPARgamma) and CCAAT/enhancer binding protein alpha (C/EBPalpha). Macrophages also express aP2, and the lack of macrophage aP2 significantly reduces atherosclerotic lesion size in hypercholesterolemic mice. We investigated the regulation of expression of macrophage aP2 and CD36, a fatty acid membrane binding protein and scavenger receptor, in response to the adipogenic agents isobutylmethylxanthine (IBMX), insulin, and dexamethasone, a combination of agents shown to induce fibroblast-to-adipocyte differentiation. Treatment of J774 macrophages with adipogenic agents significantly induced aP2 mRNA expression, while CD36 expression was inhibited. Dexamethasone was essential and sufficient to induce aP2 expression, and insulin had a synergistic effect. However, IBMX antagonized induced-aP2 expression. aP2 protein expression and [14C]oleic acid uptake by macrophages were also increased by dexamethasone. Unlike what occurs in adipocytes, adipogenic agents had mixed effects on the expression of PPARgamma and C/EBPalpha in macrophages. Our data demonstrate differences in the regulation of aP2 in adipocytes and macrophages and show that macrophage aP2 expression by adipogenic agents is independent of the PPARgamma and/or C/EBPalpha signaling pathway.