Thiazolidinedione derivatives are antidiabetic agents that increase the insulin sensitivity of target tissues in animal models of non-insulin-dependent diabetes mellitus. In vitro, thiazolidinediones promote adipocyte differentiation of preadipocyte and mesenchymal stem cell lines; however, the molecular basis for this adipogenic effect has remained unclear. Here, we report that thiazolidinediones are potent and selective activators of peroxisome proliferator-activated receptor gamma (PPAR gamma), a member of the nuclear receptor superfamily recently shown to function in adipogenesis. The most potent of these agents, BRL49653, binds to PPAR gamma with a Kd of approximately 40 nM. Treatment of pluripotent C3H10T1/2 stem cells with BRL49653 results in efficient differentiation to adipocytes. These data are the first demonstration of a high affinity PPAR ligand and provide strong evidence that PPAR gamma is a molecular target for the adipogenic effects of thiazolidinediones. Furthermore, these data raise the intriguing possibility that PPAR gamma is a target for the therapeutic actions of this class of compounds.

Bile acids repress the transcription of cytochrome P450 7A1 (CYP7A1), which catalyzes the rate-limiting step in bile acid biosynthesis. Although bile acids activate the farnesoid X receptor (FXR), the mechanism underlying bile acid-mediated repression of CYP7A1 remained unclear. We have used a potent, nonsteroidal FXR ligand to show that FXR induces expression of small heterodimer partner 1 (SHP-1), an atypical member of the nuclear receptor family that lacks a DNA-binding domain. SHP-1 represses expression of CYP7A1 by inhibiting the activity of liver receptor homolog 1 (LRH-1), an orphan nuclear receptor that is known to regulate CYP7A1 expression positively. This bile acid-activated regulatory cascade provides a molecular basis for the coordinate suppression of CYP7A1 and other genes involved in bile acid biosynthesis.

Accumulation of cholesterol causes both repression of genes controlling cholesterol biosynthesis and cellular uptake and induction of cholesterol 7␣-hydroxylase, which leads to the removal of cholesterol by increased metabolism to bile acids. Here, we report that LXR␣ and LXR␤, two orphan members of the nuclear receptor superfamily, are activated by 24(S),25-epoxycholesterol and 24(S)-hydroxycholesterol at physiologic concentrations. In addition, we have identified an LXR response element in the promoter region of the rat cholesterol 7␣-hydroxylase gene. Our data provide evidence for a new hormonal signaling pathway that activates transcription in response to oxysterols and suggest that LXRs play a critical role in the regulation of cholesterol homeostasis.Cholesterol (CH) 1 is a major structural constituent of cellular membranes and serves as the biosynthetic precursor for bile acids and steroid hormones. Animal cells can obtain CH endogenously through de novo synthesis from acetyl-CoA or exogenously through receptor-mediated endocytosis of low density lipoproteins. Cells must balance the internal and external sources of CH so as to maintain mevalonate biosynthesis while at the same time avoiding the accumulation of excess CH, which can result in diseases such as atherosclerosis, gallstones, and several lipid storage disorders (1).CH homeostasis is maintained in part through feedback regulation of the low density lipoprotein receptor gene and at least two genes encoding enzymes in the CH biosynthetic pathway, 3-hydroxy-3-methylglutaryl coenzyme A synthase and 3-hydroxy-3-methylglutaryl coenzyme A reductase (1). Although increases in dietary CH lead to the inhibition of expression of these genes in vivo, it remains unclear whether CH or CH metabolites are responsible for this inhibition (2). Experiments performed in vitro using several different cell lines have indicated that derivatives of CH that are oxygenated on the CH side chain are significantly more potent in the suppression of sterol biosynthesis than CH (3). These oxysterols are produced through the actions of P450 enzymes in various metabolic pathways including bile acid synthesis in the liver and sex hormone synthesis in the adrenal glands. The in vitro activities of oxysterols together with their presence in vivo suggests that oxysterols may serve in metabolic feedback loops to regulate CH homeostasis.Although CH and its oxysterol metabolites can repress gene transcription, in at least one instance dietary CH has been shown to stimulate gene expression. Expression of the cholesterol 7␣-hydroxylase (CYP7A) gene, which encodes the enzyme responsible for the initial and rate-limiting step in the conversion of CH to bile acids (4), is up-regulated in rats fed a CH-rich diet (5-7). This stimulatory effect provides a regulatory mechanism whereby excess dietary CH can be converted to more polar bile acids for subsequent removal from the body. Although the molecular mechanism is unknown, induction of CYP7A expression in the presence of CH occurs at the level...

Xenobiotics induce the transcription of cytochromes P450 (CYPs) 2B and 3A through the constitutive androstane receptor (CAR; NR1I3) and pregnane X receptor (PXR; NR1I2), respectively. In this report, we have systematically compared a series of xenobiotics and natural steroids for their effects on mouse and human CAR and PXR. Our results demonstrate dual regulation of PXR and CAR by a subset of compounds that affect CYP expression. Moreover, there are marked pharmacological differences between the mouse (m) and human (h) orthologs of both CAR and PXR. For example, the planar hydrocarbon 1,4-bis[2-(3,5-dichloropyridyl-oxy-)]benzene activates mCAR and hPXR but has little or no activity on hCAR and mPXR. In contrast, the CAR deactivator androstanol activates both mouse and human PXR. Similarly, the PXR activator clotrimazole is a potent deactivator of hCAR. Using radioligand binding and fluorescence resonance energy transfer assays, we demonstrate that several of the compounds that regulate mouse and human CAR, including natural steroids, bind directly to the receptors. Our results suggest that CAR, like PXR, is a steroid receptor that is capable of recognizing structurally diverse compounds. Moreover, our findings underscore the complexity in the physiologic response to xenobiotics.

The human nuclear pregnane X receptor (hPXR) activates cytochrome P450-3A expression in response to a wide variety of xenobiotics and plays a critical role in mediating dangerous drug-drug interactions. We present the crystal structures of the ligand-binding domain of hPXR both alone and in complex with the cholesterol-lowering drug SR12813 at resolutions of 2.5 and 2.75 angstroms, respectively. The hydrophobic ligand-binding cavity of hPXR contains a small number of polar residues, permitting SR12813 to bind in three distinct orientations. The position and nature of these polar residues were found to be critical for establishing the precise pharmacologic activation profile of PXR. Our findings provide important insights into how hPXR detects xenobiotics and may prove useful in predicting and avoiding drug-drug interactions.