Peter Tontonoz

Stimulation of adipogenesis in fibroblasts by PPARγ2, a lipid-activated transcription factor

Peroxisome proliferator-activated receptor gamma 2 (PPAR gamma 2) is an adipocyte-specific nuclear hormone receptor that has recently been identified as a key regulator of two fat cell enhancers. Transcriptional activation by PPAR gamma 2 is potentiated by a variety of lipids and lipid-like compounds, including naturally occurring polyunsaturated fatty acids. We demonstrate here that retroviral expression of PPAR gamma 2 stimulates adipose differentiation of cultured fibroblasts. PPAR activators promote the differentiation of PPAR gamma 2-expressing cells in a dose-dependent manner. C/EBP alpha, a second transcription factor induced during adipocyte differentiation, can cooperate with PPAR gamma 2 to stimulate the adipocyte program dramatically. Our results suggest that the physiologic role of PPAR gamma 2 is to regulate development of the adipose lineage in response to endogenous lipid activators and that this factor may serve to link the process of adipocyte differentiation to systemic lipid metabolism.

15-Deoxy-Δ12,14-Prostaglandin J2 is a ligand for the adipocyte determination factor PPARγ

Abstract Regulation of adipose cell mass is a critical homeostatic process in higher vertebrates. The conversion of fibroblasts into cells of the adipose lineage is induced by expression of the orphan nuclear receptor PPARyγ. This suggests that an endogenous PPARγ ligand may be an important regulator of adipogenesis. By assaying arachidonate metabolites for their capacity to activate PPAR response elements, we have identified 15-deoxy- Δ 12,14 -prostaglandin J 2 as both a PPARγ ligand and an inducer of adipogenesis. Similarly, the thiazolidinedione class of antidiabetic drugs also bind to PPARγ and act as potent regulators of adipocyte development. Thus, adipogenic prostanoids and antidiabetic thiazolidinediones initiate key transcriptional events through a common nuclear receptor signaling pathway. These findings suggest a pivotal role for PPARγ and its endogenous ligand in adipocyte development and glucose homeostasis and as a target for intervention in metabolic disorders.

Fat and Beyond: The Diverse Biology of PPARγ

The nuclear receptor PPARgamma is a ligand-activated transcription factor that plays an important role in the control of gene expression linked to a variety of physiological processes. PPARgamma was initially characterized as the master regulator for the development of adipose cells. Ligands for PPARgamma include naturally occurring fatty acids and the thiazolidinedione (TZD) class of antidiabetic drugs. Activation of PPARgamma improves insulin sensitivity in rodents and humans through a combination of metabolic actions, including partitioning of lipid stores and the regulation of metabolic and inflammatory mediators termed adipokines. PPARgamma signaling has also been implicated in the control of cell proliferation, atherosclerosis, macrophage function, and immunity. Here, we review recent advances in our understanding of the diverse biological actions of PPARgamma with an eye toward the expanding therapeutic potential of PPARgamma agonist drugs.

A PPARγ-LXR-ABCA1 Pathway in Macrophages Is Involved in Cholesterol Efflux and Atherogenesis

Abstract Previous work has implicated PPARγ in the regulation of CD36 expression and macrophage uptake of oxidized LDL (oxLDL). We provide evidence here that in addition to lipid uptake, PPARγ regulates a pathway of cholesterol efflux. PPARγ induces ABCA1 expression and cholesterol removal from macrophages through a transcriptional cascade mediated by the nuclear receptor LXRα. Ligand activation of PPARγ leads to primary induction of LXRα and to coupled induction of ABCA1. Transplantation of PPARγ null bone marrow into LDLR −/− mice results in a significant increase in atherosclerosis, consistent with the hypothesis that regulation of LXRα and ABCA1 expression is protective in vivo. Thus, we propose that PPARγ coordinates a complex physiologic response to oxLDL that involves particle uptake, processing, and cholesterol removal through ABCA1.

PPAR-|[gamma]| dependent and independent effects on macrophage-gene expression in lipid metabolism and inflammation

Peroxisome proliferator-activated receptor-γ (PPAR-γ) is highly expressed in lipid-accumulating macrophages of the coronary artery. In light of this, the wide-spread clinical use of thiazolidinediones (TZDs) in the treatment of type II diabetes raises concerns about the role of PPAR-γ in macrophage function and disease progression. To define the role of PPAR-γ in macrophage biology, we used homologous recombination to create embryonic stem cells that were homozygous for a null mutation in the PPAR-γ gene. We demonstrate here that PPAR-γ is neither essential for nor substantially affects the development of the macrophage lineage both in vitro and in vivo. In contrast, we show it is an important regulator of the scavenger receptor CD36, which has been genetically linked to lipid accumulation in macrophages. Both 15-deoxy-Δ12,14prostaglandin J2 and thiazolidinediones have anti-inflammatory effects that are independent of PPAR-γ. We show that PPAR-γ is required for positive effects of its ligands in modulating macrophage lipid metabolism, but that inhibitory effects on cytokine production and inflammation may be receptor independent.

Reciprocal regulation of inflammation and lipid metabolism by liver X receptors

Macrophages have important roles in both lipid metabolism and inflammation and are central to the pathogenesis of atherosclerosis. The liver X receptors (LXRs) are established mediators of lipid-inducible gene expression, but their role in inflammation and immunity is unknown. We demonstrate here that LXRs and their ligands are negative regulators of macrophage inflammatory gene expression. Transcriptional profiling of lipopolysaccharide (LPS)-induced macrophages reveals reciprocal LXR-dependent regulation of genes involved in lipid metabolism and the innate immune response. In vitro, LXR ligands inhibit the expression of inflammatory mediators such as inducible nitric oxide synthase, cyclooxygenase (COX)-2 and interleukin-6 (IL-6) in response to bacterial infection or LPS stimulation. In vivo, LXR agonists reduce inflammation in a model of contact dermatitis and inhibit inflammatory gene expression in the aortas of atherosclerotic mice. These findings identify LXRs as lipid-dependent regulators of inflammatory gene expression that may serve to link lipid metabolism and immune functions in macrophages.

Synthetic LXR ligand inhibits the development of atherosclerosis in mice

The nuclear receptors LXRα and LXRβ have been implicated in the control of cholesterol and fatty acid metabolism in multiple cell types. Activation of these receptors stimulates cholesterol efflux in macrophages, promotes bile acid synthesis in liver, and inhibits intestinal cholesterol absorption, actions that would collectively be expected to reduce atherosclerotic risk. However, synthetic LXR ligands have also been shown to induce lipogenesis and hypertriglyceridemia in mice, raising questions as to the net effects of these compounds on the development of cardiovascular disease. We demonstrate here that the nonsteroidal LXR agonist GW3965 has potent antiatherogenic activity in two different murine models. In LDLR−/− mice, GW3965 reduced lesion area by 53% in males and 34% in females. A similar reduction of 47% was observed in male apoE−/− mice. Long-term (12-week) treatment with LXR agonist had differential effects on plasma lipid profiles in LDLR−/− and apoE−/− mice. GW3965 induced expression of ATP-binding cassettes A1 and G1 in modified low-density lipoprotein-loaded macrophages in vitro as well as in the aortas of hyperlipidemic mice, suggesting that direct actions of LXR ligands on vascular gene expression are likely to contribute to their antiatherogenic effects. These observations provide direct evidence for an atheroprotective effect of LXR agonists and support their further evaluation as potential modulators of human cardiovascular disease.

Liver X receptors as integrators of metabolic and inflammatory signaling

The liver X receptors (LXRs) are nuclear receptors that play central roles in the transcriptional control of lipid metabolism. LXRs function as nuclear cholesterol sensors that are activated in response to elevated intracellular cholesterol levels in multiple cell types. Once activated, LXRs induce the expression of an array of genes involved in cholesterol absorption, efflux, transport, and excretion. In addition to their function in lipid metabolism, LXRs have also been found to modulate immune and inflammatory responses in macrophages. Synthetic LXR agonists promote cholesterol efflux and inhibit inflammation in vivo and inhibit the development of atherosclerosis in animal models. The ability of LXRs to integrate metabolic and inflammatory signaling makes them particularly attractive targets for intervention in human metabolic disease.

Direct and Indirect Mechanisms for Regulation of Fatty Acid Synthase Gene Expression by Liver X Receptors

The nuclear receptors LXRα and LXRβ have been implicated in the control of lipogenesis and cholesterol homeostasis. Ligand activation of these receptors in vivoinduces expression of the LXR target gene SREBP-1cand increases plasma triglyceride levels. Expression of fatty acid synthase (FAS), a central enzyme in de novo lipogenesis and an established target of the SREBP-1 pathway, is also induced by LXR ligands. The effects of LXR ligands on FAS expression have been proposed to be entirely secondary to the induction of SREBP-1c. We demonstrate here that LXRs regulate FAS expression through direct interaction with the FAS promoter as well as through activation of SREBP-1c expression. Induction of FAS expression in HepG2 cells by LXR ligands is reduced, but not abolished, under conditions where SREBP processing is suppressed. Moreover, LXR ligands induce FAS expression in CHO-7 cells without altering expression of SREBP-1. We demonstrate that in addition to tandem SREBP sites, the FASpromoter contains a high affinity binding site for the LXR/RXR heterodimer that is conserved in diverse animal species including birds, rodents, and humans. The LXR and SREBP binding sites independently confer LXR responsiveness on the FASpromoter, and maximal induction requires both transcription factors. Transient elevation of plasma triglyceride levels in mice treated with a synthetic LXR agonist correlates with transient induction of hepatic FAS expression. These results indicate that the LXR signaling pathway modulates FAS expression through distinct but complementary mechanisms and suggest that the FAS gene may be a critical target in the control of lipogenesis by LXRs.

ApoE Promotes the Proteolytic Degradation of Aβ

Apolipoprotein E is associated with age-related risk for Alzheimers disease and plays critical roles in Abeta homeostasis. We report that ApoE plays a role in facilitating the proteolytic clearance of soluble Abeta from the brain. The endolytic degradation of Abeta peptides within microglia by neprilysin and related enzymes is dramatically enhanced by ApoE. Similarly, Abeta degradation extracellularly by insulin-degrading enzyme is facilitated by ApoE. The capacity of ApoE to promote Abeta degradation is dependent upon the ApoE isoform and its lipidation status. The enhanced expression of lipidated ApoE, through the activation of liver X receptors, stimulates Abeta degradation. Indeed, aged Tg2576 mice treated with the LXR agonist GW3965 exhibited a dramatic reduction in brain Abeta load. GW3965 treatment also reversed contextual memory deficits. These data demonstrate a mechanism through which ApoE facilitates the clearance of Abeta from the brain and suggest that LXR agonists may represent a novel therapy for AD.