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We have identified Wnt10b as a potent inhibitor of adipogenesis that must be suppressed for preadipocytes to differentiate in vitro. Here, we demonstrate that a specific inhibitor of glycogen synthase kinase 3, CHIR 99021, mimics Wnt signaling in preadipocytes. CHIR 99021 stabilizes free cytosolic ␤-catenin and inhibits adipogenesis by blocking induction of CCAAT/enhancer-binding protein ␣ and peroxisome proliferatoractivated receptor ␥. Preadipocyte differentiation is inhibited when 3T3-L1 cells are exposed to CHIR 99021 for any 24 h period during the first 3 days of adipogenesis. Consistent with this time frame of inhibition, expression of Wnt10b mRNA is suppressed upon induction of differentiation, with a 50% decline by 6 h and complete inhibition by 36 h. Of the agents used to induce differentiation, exposure of 3T3-L1 cells to methyl-isobutylxanthine or cAMP is sufficient to suppress expression of Wnt10b mRNA. Inhibition of adipogenesis by Wnt10b is likely mediated by Wnt receptors, Frizzled 1, 2, and/or 5, and co-receptors low density lipoprotein receptor-related proteins 5 and 6. These receptors, like Wnt10b, are highly expressed in preadipocytes and stromal vascular cells. Finally, we demonstrate that disruption of extracellular Wnt signaling by expression of secreted Frizzled related proteins causes spontaneous adipocyte conversion.White adipose tissue is an important depot for short and long term energy storage. In addition, adipose tissue is an endocrine organ that regulates energy homeostasis through secretion of leptin, Acrp30/adiponectin, resistin, and other factors (1). The study of adipocyte biology has been greatly facilitated by development of immortalized preadipocyte lines (e.g. 3T3-L1) (2) that differentiate into adipocytes and recapitulate many of the metabolic and endocrine functions of adipocytes in vivo. Analysis of differentiation in preadipocyte lines and in animals has led to a paradigmatic model of the genetic program of adipogenesis (3-5). In response to stimulators of adipogenesis, two transcription factors, CCAAT/enhancer-binding protein ␤ (C/ EBP␤) 1 and C/EBP␦ are rapidly and transiently induced. These proteins then stimulate expression of the key adipogenic transcription factors, C/EBP␣ and peroxisome proliferator-activated receptor ␥ (PPAR␥), which together induce expression of the complement of genes necessary to create the adipocyte phenotype. Investigations of factors that regulate progression through this adipogenic program are important for our understanding of adipocyte differentiation. Differentiation of preadipocytes into adipocytes is regulated by a balance of local and endocrine factors that either stimulate or inhibit differentiation (6). Well known factors that stimulate differentiation of preadipocyte lines include glucocorticoid agonists, high concentrations of insulin to stimulate the insulinlike growth factor 1 receptor, PPAR␥ agonists, and agents that elevate cAMP (3). Factors that counter these positive stimuli include Wnt10b, tumor necrosis factor ␣ (TNF␣), ...

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Wnt signaling maintains preadipocytes in an undifferentiated state. When Wnt signaling is enforced, 3T3-L1 preadipocytes no longer undergo adipocyte conversion in response to adipogenic medium. Here we used microarray analyses to identify subsets of genes whose expression is aberrant when differentiation is blocked through enforced Wnt signaling. Furthermore, we used the microarray data to identify potentially important adipocyte genes and chose one of these, the liver X receptor ␣ (LXR␣), for further analyses. Our studies indicate that enforced Wnt signaling blunts the changes in gene expression that correspond to mitotic clonal expansion, suggesting that Wnt signaling inhibits adipogenesis in part through dysregulation of the cell cycle. Experiments designed to uncover the potential role of LXR␣ in adipogenesis revealed that this transcription factor, unlike CCAAT/enhancer binding protein ␣ and peroxisome proliferator-activated receptor gamma, is not adipogenic but rather inhibits adipogenesis if inappropriately expressed and activated. However, LXR␣ has several important roles in adipocyte function. Our studies show that this nuclear receptor increases basal glucose uptake and glycogen synthesis in 3T3-L1 adipocytes. In addition, LXR␣ increases cholesterol synthesis and release of nonesterified fatty acids. Finally, treatment of mice with an LXR␣ agonist results in increased serum levels of glycerol and nonesterified fatty acids, consistent with increased lipolysis within adipose tissue. These findings demonstrate new metabolic roles for LXR␣ and increase our understanding of adipogenesis.Adipocytes play a central role in energy balance, both as a reservoir, storing and releasing fuel, and as endocrine cells, secreting factors that regulate whole-body energy metabolism (49). Understanding this cell type is becoming increasingly important because of the rising incidence of obesity and its associated disorder, type II diabetes. One widely used model for studying the adipocyte is the 3T3-L1 cell line, which was derived from disaggregated mouse embryos and selected based on the propensity of these cells to differentiate into adipocytes in culture (14). Over the last 30 years, this cell line has proven to be a faithful model for studying adipocyte biology, particularly adipogenesis and energy metabolism.