Peroxisome proliferator-activated receptor gamma agonists as therapy for chronic airway inflammation

European Journal of Pharmacology 533 (2006) 101 – 109 www.elsevier.com/locate/ejphar

Review

Peroxisome proliferator-activated receptor gamma agonists as therapy for chronic airway inflammation Maria G. Belvisi ⁎, David J. Hele, Mark A. Birrell Respiratory Pharmacology, Airway Diseases, National Heart and Lung Institute, Faculty of Medicine, Imperial College, Guy Scadding Building, Dovehouse Street, London, SW3 6LY UK Accepted 13 December 2005 Available online 3 February 2006

Abstract Peroxisome proliferator-activated receptor gamma (PPARγ) is a ligand-activated transcription factor belonging to the nuclear hormone receptor superfamily. PPARγ regulates several metabolic pathways by binding to sequence-specific PPAR response elements in the promoter region of target genes, including lipid biosynthesis and glucose metabolism. Synthetic PPARγ agonists have been developed, such as the thiazolidinediones rosiglitazone and pioglitazone. These act as insulin sensitizers and are used in the treatment of type 2 diabetes. Recently however, PPARγ ligands have been implicated as regulators of cellular inflammatory and immune responses. They are thought to exert anti-inflammatory effects by negatively regulating the expression of pro-inflammatory genes. Several studies have demonstrated that PPARγ ligands possess anti-inflammatory properties and that these properties may prove helpful in the treatment of inflammatory diseases of the airways. This review will outline the antiinflammatory effects of synthetic and endogenous PPARγ ligands and discuss their potential therapeutic effects in animal models of inflammatory airway disease. © 2006 Elsevier B.V. All rights reserved. Keywords: PPARγ; Airway; Inflammation; Asthma

Contents 1. Introduction . . . . . . . . . . . . . . . . . . 2. PPARγ and gene expression . . . . . . . . . 3. Endogenous PPARγ ligands . . . . . . . . . 4. Synthetic PPARγ ligands . . . . . . . . . . . 5. PPARγ in inflammation and airways disease . 6. PPARγ in animal models of airways disease . 7. New developments in PPAR pharmacology . 8. Conclusion . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . .

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1. Introduction Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors belonging to the nuclear ⁎ Corresponding author. Tel.: +44 207 351 8270; fax: +44 207 351 8173. E-mail address: [email protected] (M.G. Belvisi). 0014-2999/$ - see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.ejphar.2005.12.048

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hormone receptor superfamily related to retinoid, glucocorticoid, and thyroid hormone receptors (Evans, 1988). All members of this superfamily have a similar structural organisation: an amino-terminal region that allows ligandindependent activation, a DNA-binding domain and a liganddependent activation domain (Moras and Gronemeyer, 1998). To date, three different PPAR subtypes have been identified

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PPARα, PPARγ, and PPARβ (also known as PPARδ). The three isoforms are the products of distinct genes and there are two splice isoforms of PPARγ: γ1 and γ2. PPARγ2 is expressed exclusively in adipose tissue whereas PPARγ1 is more widely expressed, although it is most abundant in adipocytes (Vidal-Puig et al., 1996; Fajas et al., 1997). The name PPAR is derived from the fact that activation of PPARα, the first member of the PPAR family to be cloned, results in peroxisome proliferation in rodent hepatocytes (Issemann and Green, 1990). Activation of PPARβ or PPARγ, however, does not elicit this response. PPARs were first identified for their role in lipid and glucose regulation and until recently their actions were thought to be limited to specific tissue types. PPARα is highly expressed in tissues exhibiting high carbolic rates of fatty acids such as the liver, heart, kidney and intestinal mucosa (Braissant et al., 1996). PPARγ is expressed at high levels in adipose tissue, where it plays a critical role in adipocyte differentiation (Spiegelman and Flier, 1996). PPARβ is almost ubiquitously expressed (Braissant et al., 1996) and its role is relatively unknown. Recently PPARγ has been suggested to be an important immunomodulator (Serhan, 1996) and to have potential as a novel anti-inflammatory target for asthma (Serhan and Devchand, 2001; Belvisi et al., 2004) (see Fig. 1). PPARγ has been found in cells of the immune system such as monocytes/macrophages, B and T cells, and dendritic cells (Braissant et al., 1996; Chinetti et al., 1998, 2000; Clark et al., 2000; Faveeuw et al., 2000; Gosset et al., 2001; Marx et al., 1998; Yang et al., 2000). PPARγ ligands have been shown to inhibit the release of pro-inflammatory cytokines from activated macrophages (Jiang et al., 1998) and airway epithelial cells (Wang et al., 2001). Furthermore, PPARγ ligands inhibit vascular smooth muscle cell proliferation (Law et al., 1996) and induce apoptosis in endothelial cells (Bishop-Bailey and Hla, 1999), vascular smooth muscle cells (Bishop-Bailey and Warner, 2003), Tlymphocytes (Clark et al., 2000) and macrophages (Chinetti et al., 1998). Various studies have now been published demonstrating the anti-inflammatory effects of PPARγ ligands in animal models

Fig. 1. Describes the effects of PPARγ agonists on various cell types within the lung.

of arthritis, ischaemia–reperfusion, inflammatory bowel disease, Alzheimer's disease and lupus nephritis (Sher and Pillinger, 2005). Evidence of anti-inflammatory effects in the airways is now starting to appear in the literature. Our group has reported that the activation of PPARγ has potent antiinflammatory effects on human airway smooth muscle cells in culture. In particular, PPARγ activation inhibited the release of survival factors and the proliferation of human airway smooth muscle cells as well as reducing the release of inflammatory mediators and inducing apoptosis (Patel et al., 2003). In an animal model of airway inflammation we have demonstrated a reduction in lipopolysaccharide-induced neutrophilia and a reduction in chemoattractants and survival factors (Birrell et al., 2004). Recently several other groups have published work demonstrating that PPARγ agonists have beneficial effects on airway inflammation in various animal models of asthma (Ueki et al., 2004). Therefore, the potential exists to exploit PPARγ agonists as putative treatments for chronic inflammatory diseases of the airways. 2. PPARγ and gene expression PPARγ regulates gene expression by binding as a heterodimer with the retinoid X receptor (RXR), a member of the nuclear hormone receptor superfamily activated by binding of 9-cis-retinoic acid. The RXR family comprises three different gene isoforms: RXRα, RXRβ, and RXRγ. RXR is widely expressed in several tissues and cells including adipose tissue, liver, kidneys, small intestine, cardiac myocytes, and monocytes/macrophages (Dubuquoy et al., 2002). The PPARγ/RXR heterodimer binds to sequence-specific PPAR response elements in the promoter region of target genes and acts as a transcriptional regulator. To prevent PPARγ/RXR binding to DNA, high-affinity complexes are formed between the inactive PPARγ/RXR heterodimers and co-repressor molecules, such as nuclear receptor co-repressor or silencing mediator for retinoic receptors. On ligand binding and activation, these co-repressors are displaced and the heterodimer is free to bind to the response element in the promoter region of the relevant target genes, resulting in either activation or suppression of a specific gene. Recruitment of co-activator proteins is required for transcriptional interaction of PPARγ with motifs in the PPAR response elements (Desvergne and Wahli, 1999). PPARγ can also function in a DNA-binding-independent manner to transrepress different target genes. It has been suggested that PPARγ may exert anti-inflammatory effects by negatively regulating the expression of pro-inflammatory genes induced during macrophage differentiation and activation. Several mechanisms for PPARγ-agonist-dependent transcriptional transrepression of pro-inflammatory macrophage responses have been suggested. Competition for limited amounts of shared transcriptional co-activators may play a role in transrepression where activated PPARγ/RXR heterodimer reduces the availability of co-activators required for gene induction by other transcriptional factors (Kodera et al., 2000).

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Thus, without distinct cofactors, transcription factors, such as activated protein-1 (AP-1), nuclear factor-кB (NF-кB), nuclear factor of activated T cells (NFAT) or signal transducer and activator of transcription (STAT), cannot cause gene expression. In lipopolysaccharide/interferon-γ (IFN-γ) stimulated macrophages, attenuation of inducible nitric oxide synthase expression in response to PPARγ activation has been explained by targeting of the two general co-activators CREB-binding protein and p300 by PPARγ (Li et al., 2000). PPARγ/RXR complexes may cause a functional inhibition by directly binding to transcription factors, preventing them from inducing gene transcription. This method of transrepression has been reported for proteins of the NF-кB family, such as p50 and p65 (Chung et al., 2000) and NFAT where PPARγ sequestration of NFAT accounted for suppression of T cell proliferation and activation (Yang et al., 2000). Another mechanism, which may result in transrepression, involves regulation of the mitogen-activated protein kinase (MAPK) cascade. PPARγ/RXR heterodimers may inhibit phosphorylation and activation of several members of the MAPK family. In a study carried out in PPARγ+/− heterozygous mice, activation of c-Jun-N-terminal kinase and p38 was significantly reduced when compared with wild-type littermates (Desreumaux et al., 2001). 3. Endogenous PPARγ ligands PPARγ is bound and activated by a variety of lipophilic ligands, including long-chain polyunsaturated fatty acids and several eicosanoids. The essential fatty acids arachidonic acid, gamolenic acid, docosahexanoic acid, and eicosapentaenoic acid, as well as modified oxidised lipids 9- and 13-hydroxyoctadecadienoic acid and 12- and 15-hydroxyeicosatetraenoic acid, bind to and activate PPARγ (Willson et al., 2000). 15deoxy-Δ12,14-PGJ2 (15d-PGJ2) has been recognised as the endogenous ligand for PPARγ and is thought to be responsible for many of its anti-inflammatory actions (Sher and Pillinger, 2005). 15d-PGJ2 is also the PPARγ agonist which has been most frequently studied experimentally (Forman et al., 1995). The cyclopentenone prostaglandin 15d-PGJ2 was first discovered in 1983, (Fitzpatrick and Wynalda, 1983) but received relatively little attention until two independent groups reported that it was capable of activating PPARγ (Forman et al., 1995; Kliewer et al., 1995). Accumulating evidence now suggests that 15d-PGJ2 is the endogenous PPARγ ligand (Sher and Pillinger, 2005). 15d-PGJ2 and other PPARγ ligands have been shown to possess anti-inflammatory activity in a wide range of inflammatory disease models and recently 15d-PGJ2 has been shown to significantly limit lung injury in an animal model of pulmonary fibrosis, a disease characterised by inflammatory cell infiltration (Genovese et al., 2005). A reduction in neutrophil influx, oedema, histological parameters and mortality were observed thus supporting the anti-inflammatory potential of PPARγ ligands. 15d-PGJ2 has been widely used as a pharmacological tool for defining the role of PPARγ and it is important to note that it can induce a variety of PPARγ-independent responses, and indeed a

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recent study demonstrated that 15d-PGJ2 exerts its antiinflammatory effect in rat chondrocytes by a PPARγ-independent mechanism, which could be conferred to a partial inhibition of inhibitor κBα degradation (Boyault et al., 2004). 15d-PGJ2 has also been shown to induce responses in cells devoid of the receptor (Chawla et al., 2001). 4. Synthetic PPARγ ligands In addition to natural ligands, a wide range of synthetic PPARγ agonists have been developed. The most widely used belong to the thiazolidinedione or glitazone class of antidiabetic drugs used in the treatment of type 2 diabetes (YkiJarvinen, 2004). These include rosiglitazone, pioglitazone, ciglitazone, and troglitazone. The first to be developed, troglitazone, has since been withdrawn from the market following the emergence of a serious hepatotoxicity in some patients. The two PPARγ agonists currently available for the treatment of type 2 diabetes in the United States are rosiglitazone and pioglitazone. Thiazolidinediones exert their insulin-sensitising and hypoglycaemic effects through stimulation of PPARγ (Berger et al., 1996). The involvement of PPARγ in the pharmacological effects of thiazolidinediones has been supported by studies showing that their binding affinity to PPARγ closely parallels their in vivo hypoglycaemic potency (Willson et al., 2000). In the last few years, there have been numerous reports indicating that the therapeutic benefits of PPARγ agonists may go far beyond their use in diabetes with increasing evidence of anti-inflammatory activities in a range of disease models from Alzheimer's to pancreatitis and evidence is now emerging of the potential benefits of PPARγ ligands in models of inflammatory airways disease. 5. PPARγ in inflammation and airways disease Inflammation plays an important role in many airways diseases and is associated with the release of pro-inflammatory cytokines and oxygen free radicals from activated inflammatory cells such as neutrophils, eosinophils, monocytes and macrophages. There is increasing evidence to suggest that PPARγ acts as an anti-inflammatory agent by negatively regulating the expression of pro-inflammatory genes induced in response to inflammatory cell activation (Von Knethen and Brune, 2003) and by inhibiting the cytokines, chemoattractants and cell survival factors involved (Sher and Pillinger, 2005; Patel et al., 2003; Birrell et al., 2004) (see Table 1 and Fig. 2). Thus, the development of PPARγ agonists has been suggested as a novel anti-inflammatory target for diseases of the airways such as asthma, chronic obstructive pulmonary disease (COPD), acute respiratory distress syndrome (ARDS) and pulmonary fibrosis (Serhan and Devchand, 2001; Patel et al., 2003; Birrell et al., 2004; Burgess et al., 2005). PPARγ is expressed in human and murine monocytes/ macrophages (Ricote et al., 1998a) and a link has been established between the state of monocyte/macrophage differentiation and activation and PPARγ expression. In

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Table 1 Effects of PPARγ ligands on various cell types Cell type

Ligand

Effect

Monocyte Macrophage

PPARγ PPARγ

Eosinophils

Troglitazone

T cells

PPARγ

Inhibited cytokines Inhibited cytokines Induced apoptosis Reduced iNOS expression Reduced transcription factors Reduced COX-2 expression Reduced cytokines, chemoattractants and cell survival Induced apoptosis Inhibited activation and proliferation Inhibited cytokines Inhibited cytokines Reduced MMP activity

Airway epithelium

PPARγ Rosiglitazone, pioglitazone Dendritic cells PPARγ Endothelium PPARγ Airway smooth PPARγ muscle Myofibroblasts General

Reduced migration Induced apoptosis Reduced cytokines Inhibited proliferation Induced apoptosis 15d-PGJ2, rosiglitazone, Reduced differentiation ciglitazone and collagen production PPARγ Inhibited release of survival factors in several cell types

Abbreviations: iNOS, inducible nitric oxide synthase; COX-2, cycloxygenase-2; MMP, matrix metalloproteinase.

human peripheral blood monocytes PPARγ is expressed at low levels. However, during differentiation into macrophages and activation the levels are much higher (Chinetti et al., 1998). Activation of monocytes/macrophages with macrophage colony-stimulating factor (MCSF) or granulocyte macrophage CSF (GM-CSF) induces an increased expression of PPARγ (Chinetti et al., 1998; Ricote et al., 1998b). Activation of PPARγ itself can lead to differentiation of monocytes to macrophages (Tontonoz et al., 1998). Recent studies show that PPARγ is expressed by human alveolar macrophages and using PPARγ agonists the authors have demonstrated that PPARγ plays an anti-inflammatory role in these cells by inhibiting cytokine production, increasing CD36 expression and enhancing the phagocytosis of apoptotic neutrophils, an essential process for the resolution of inflammation (Asada et al., 2003). This data supports the potential of PPARγ ligands in the treatment of inflammatory disorders of the lung. Our group was the first to show that human airway smooth muscle cells express PPARγ and PPARα and that treatment with endogenous and synthetic PPARγ ligands could inhibit serum-induced growth of these cells and promote apoptosis (Patel et al., 2003). This study also revealed that PPARγ activation inhibited the release of the cytokines G-CSF and GM-CSF. The effect on cell growth and G-CSF was greater than that produced by a glucocorticoid, the current drug class of choice for the treatment of inflammatory airways disease. This suggests that PPARγ agonists may provide a novel alternative approach to the treatment of inflammatory diseases of the airways and one that has advantages over the therapies currently used.

PPARγ agonists suppress the production of the inflammatory cytokine interleukin (IL)-8 in airway epithelial cells (Wang et al., 2001) thus suggesting the possibility of reducing leukocyte recruitment and airway inflammation. PPARγ agonists also inhibit other inflammatory cytokines such as, IL-1β, IL-6 and tumour necrosis factor (TNF)-α, in stimulated human peripheral blood monocytes (Jiang et al., 1998). PPARγ is markedly up-regulated in activated peritoneal macrophages and PPARγ ligands inhibit the expression of inducible nitric oxide synthase, gelatinase B and scavenger receptor A genes, in part by antagonising the activities of the transcription factors AP-1, STAT and NF-κB (Ricote et al., 1998a). The expression of PPARγ in macrophages is upregulated by IL-4 and IL-4 also enhances the activation of PPARγ via the production of endogenous PPARγ ligands such as 13-hydroxyoctadecadienoic acid and 12- and 15hydroxyeicosatetraenoic acid, (Huang et al., 1999). 15dprostaglandin J2 (15d-PGJ2) and 13-hydroxyoctadecadienoic acid inhibited lipopolysaccharide-induced IL-10 and IL-12 production by macrophages (Azuma et al., 2001). PPARγ activation suppresses cycloxygenase-2 expression by preventing activation and translocation of NF-κB (Inoue et al., 2000; Maggi et al., 2000). Eosinophils may play a pivotal role in the development of allergic diseases such as asthma. IL-5 and eotaxin are critical cytokines/chemokines for eosinophil activation. Recent studies have demonstrated the expression of PPARγ by human eosinophils and the effects of PPARγ agonists on these cells. The PPARγ agonist, troglitazone reduced both IL-5-stimulated eosinophil survival and eotaxin-directed eosinophil chemotaxis suggesting a role for PPARγ agonists in the treatment of allergic diseases such as asthma (Ueki et al., 2004).

Fig. 2. Points of intervention of PPARγ or PPARγ ligands after inflammatory stimulus initiates signal transduction in a cell. Lines in red indicate the consequences of inflammatory stimulus, lines in blue indicate the possible points of anti-inflammatory intervention of PPARγ.

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Matrix metalloproteinases (MMPs) are known to be involved in airway wall remodelling and are thought to play a role in the development of chronic inflammatory diseases of the airways. In a study using human bronchial epithelial cells Hetzel et al. (2003) found that PPARγ was expressed and was functionally active in these cells. Activation of PPARγ by rosiglitazone or pioglitazone significantly reduced TNF-alpha and PMAinduced MMP-9 gelatinolytic activity, but did not alter the expression of tissue inhibitor of MMPs type 1, the endogenous inhibitor of MMP-9. They also demonstrated a decrease in MMP-9 mRNA expression following treatment with PPARγ which resulted from the inhibition of NF-кB activation in these cells (Hetzel et al., 2003). Limiting the expression of matrix degrading MMP-9 by PPARγ activation might have therapeutic potential in the treatment of chronic inflammatory diseases of the respiratory system. Myofibroblasts are one of the key effector cells in pulmonary fibrosis and are the primary source of extracellular matrix production. Burgess et al. (2005) have shown that both endogenous (15d-PGJ2) and synthetic (ciglitazone and rosiglitazone) PPARγ agonists inhibit transforming growth factor (TGF)-β driven myofibroblast differentiation. PPARγ agonists also potently attenuated TGF-β driven Type I collagen protein production (Burgess et al., 2005). Thus, PPARγ agonists may provide potential therapy for fibrotic diseases of the lung. Dendritic cells are powerful antigen-presenting cells with a unique capacity to stimulate naïve T cells and the migration of dendritic cells from the epithelia to the lymphoid organs represents a tightly regulated series of events involved in the induction of the immune response. A recent study has shown that PPARγ activation reduces the spontaneous migration of antigen bearing lung dendritic cells (Angeli et al., 2003) suggesting a potential role for PPARγ agonists in the treatment of allergic asthma. In T cells, PPARγ activation inhibits IL-2 production via a mechanism believed to involve transrepression of NFAT (Yang et al., 2000). PPARγ activation has also been reported to down-regulate CCR2, the receptor for monocyte chemoattractant protein-1, in circulating rat monocytes (Ishibashi et al., 2002). As mentioned above PPARγ also has a role in apoptosis in several cell types. Chinetti et al. (1998) have reported that PPARγ agonists induce apoptosis in TNF-α/ interferon-γ-stimulated macrophages by interfering with the anti-apoptotic NF-κB signalling pathway and many other groups have reported the induction of apoptosis in a variety of cell types such as endothelial cells, smooth muscle cells, T cells and macrophages.

treating inflammatory airways disease remains to be seen. Recently studies have been published outlining the effect of synthetic and endogenous PPARγ agonists in a variety of airways disease models (see Table 2). Allergic asthma is characterised by cytokine production, inflammatory cell influx, airway hyperresponsiveness, increased mucus production and airway remodelling and several groups have employed animal models of asthma to determine the effects of synthetic and natural PPARγ agonists on these axes. Trifillieff et al. (2003) showed that intranasally administered agonists of PPARα (GW 9578) and PPARγ (GI 262570), but not PPARδ (GW 501516), inhibited allergen-induced bronchoalveolar lavage eosinophil and lymphocyte influx in ovalbumin sensitised and challenged mice. In a similar murine model of allergic asthma, orally administered ciglitazone, a potent synthetic PPARγ ligand, significantly reduced lung inflammation and mucus production. T cells from these ciglitazone treated mice produced less interferon-γ, IL-4, and IL-2 upon allergen challenge in vitro (Mueller et al., 2003). In a study where PPAR agonists were administered by aerosol it has been demonstrated that PPARα and PPARγ activated receptors decreased antigen-induced airway hyperresponsiveness, lung inflammation, eosinophilia, cytokine production, GATA-3 expression and serum levels of antigen-specific IgE in a murine model of human asthma (Woerly et al., 2003). A further study where ciglitazone was administered by aerosol in a murine model of asthma demonstrated a reduction in airway hyperresponsiveness, basement membrane thickness, mucus production, collagen deposition and TGF-β synthesis (Honda et al., 2004). Also using a murine model of asthma Lee et al. (2005) Table 2 Effects of PPARγ ligands in animal models of airway inflammation Model

Ligand

Effect

Murine asthma

GI 262570

Murine asthma

Ciglitazone

Murine asthma

Rosiglitazone

Murine asthma

Rosiglitazone

LPS-induced neutrophilia

Rosiglitazone

Reduced eosinophil influx Reduced lymphocyte influx (Trifillieff et al., 2003) Reduced lung inflammation, mucus production (Mueller et al., 2003), AHR, eosinophilia, cytokine production, serum IgE, basement membrane thickness, collagen deposition and TGF-β synthesis (Honda et al., 2004) Reduced lung inflammation, piogliatazone AHR and cytokine production Adv-PPARγ (Lee et al., 2005) Inhibited dendritic cell migration reduced T cell proliferation, increased IL-10 levels and reduced eosinophil recruitment (Hammad et al., 2004) Reduced neutrophilia

6. PPARγ in animal models of airways disease As described above there is now an encouraging body of in vitro based evidence suggesting that PPARγ activation or treatment with PPARγ agonists may translate into beneficial effects in animal models of airway disease and potentially into putative treatments for human airway disease. Several pharmaceutical companies are developing newer and more potent PPARγ agonists for various inflammatory indications (Oates et al., 2002) but whether this approach proves to be beneficial in

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SB219994 Pulmonary fibrosis

15d-PGJ2 rosiglitazone

Reduced chemoattractants reduced survival factors (Birrell et al., 2004) Reduced neutrophilia, oedema, lung injury and mortality reduced iNOS production (Genovese et al., 2005)

Abbreviations: iNOS, inducible nitric oxide synthase; AHR, airways hyperresponsiveness; TGF-β, transforming growth factor-β; Adv-PPARγ, adenovirus carrying PPARγ cDNA.

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determined the effect of not only the PPARγ agonists, rosiglitazone and pioglitazone, but also an adenovirus carrying PPARγ cDNA (AdPPARγ), on allergen-induced bronchial inflammation and airway hyperresponsiveness. Administration of the agonists or the AdPPARγ reduced bronchial inflammation and airway hyperresponsiveness. Expression of PPARγ was increased by ovalbumin inhalation, and the increase was further enhanced by the administration of the PPARγ agonists or the AdPPARγ. Levels of IL-4, IL-5, IL-13, and eosinophil cationic protein were increased after ovalbumin inhalation, and the increased levels were significantly reduced by the administration of the PPARγ agonists or the AdPPARγ. The results also showed that the administration of PPARγ agonists or the AdPPARγ up-regulated phosphatase and tensin homologue deleted on chromosome ten (PTEN) expression in the lungs of the ovalbumin treated animals. This up-regulation correlated with a decrease in phosphatidylinositol 3-kinase activity. These findings demonstrate a protective role for PPARγ in the pathogenesis of the asthma phenotype and suggest that this may occur via the regulation of PTEN expression (Lee et al., 2005). However, it should be noted that in another recent publication which showed an increase in protein expression of PPARγ after antigen challenge, a reduction in protein expression resulted after treatment with a PPARγ agonist (Honda et al., 2004). This group also claimed cause and effect suggesting that further studies are needed to clarify the situation. Dendritic cells have an important role to play in the development of allergic asthma. Angeli et al. (2003) demonstrated that PPARγ activation inhibits the migration of dendritic cells from the airway mucosa to the thoracic lymph nodes and also profoundly reduces the priming of antigen-specific T lymphocytes in the draining lymph nodes. In a follow up study this group demonstrated that the PPARγ agonist rosiglitazone reduced the proliferation of antigenspecific T cells in the draining mediastinal lymph nodes and dramatically increased T cell production of the immunoregulatory cytokine IL-10. A reduction in the recruitment of eosinophils into the bronchoalveolar lavage fluid was also observed in this study suggesting that PPAR-gamma activation prevents induction of Th2-dependent eosinophilic airway inflammation and might contribute to immune homeostasis in the lung (Hammad et al., 2004). The wide spectrum of anti-inflammatory effects demonstrated in these studies offers compelling evidence that PPARγ agonists may provide a new therapeutic modality for the treatment of allergic asthma. A role for PPAR activation in the treatment of COPD has been suggested (Patel et al., 2003); however, this potential has only recently been examined in vivo. Respiratory diseases such as COPD and ARDS are thought to involve an influx of neutrophils into the airways and using an LPS-induced model of airway neutrophilia Birrell et al. (2004) examined the effect of two structurally different PPARγ agonists, rosiglitazone and SB219994. The ligands both inhibited airway neutrophilia and the associated chemoattractant/survival factors, keratinocytederived chemokine (murine IL-8 and G-CSF), suggesting that

this approach may be beneficial in the treatment of airway diseases involving airway neutrophilia. Pulmonary fibrosis is a progressive interstitial lung disease characterised by inflammatory cell infiltration, fibroblast proliferation and excessive deposition of extracellular matrix proteins in the lung parenchyma. In a study using bleomycininduced lung injury to provide an animal model of lung fibrosis, recent evidence has shown that the PPARγ agonist rosiglitazone and the endogenous PPARγ agonist 15d-PGJ2 provided significant protection, reducing neutrophil infiltration, oedema, lung injury, weight loss and mortality rate. In addition, the two PPARγ agonists inhibited the increase in immunoreactivity to nitrotyrosine, poly (ADP ribose) polymerase and inducible nitric oxide synthase seen in this model (Genovese et al., 2005). Therefore the in vivo data supports the view that PPARγ agonists may have potential for the therapy of chronic inflammatory disorders of the airways such as asthma, COPD, ARDS and pulmonary fibrosis. 7. New developments in PPAR pharmacology The thiazolidinediones rosiglitazone and pioglitazone, which are full agonists of PPARγ, have significant clinical efficacy but their use is associated with adverse side effects including plasma volume expansion, haemodilution, oedema, increased adiposity and weight gain (Yki-Jarvinen, 2004). The have also been shown to induce cardiac enlargement in some preclinical species (Berger et al., 2003; Arakawa et al., 2004). These undesirable effects, and the potential to cause congestive heart failure in the subset of diabetic patients with underlying heart disease, limit the clinical application of these drugs. Consequently, identification of PPARγ agonists that have an improved therapeutic index has remained an important goal. This has resulted in the development of selective PPARγ modulators and several of these have now been identified that possess desirable efficacy and improved tolerance. Although these molecules have yet to be tested in models of airway disease they do represent a significant step forward in terms of safety, while retaining the efficacy demonstrated by the thiazolidinediones. The first described was nTZDpa, a potent, selective, non-thiazolidinedione PPARγ modulator (Berger et al., 2003). nTZDpa is a partial agonist at PPARγ in cell based transactivation assays, and has decreased adipogenic activity and an attenuated gene signature in adipocytes in culture. These novel activities result from the unique physical interaction of nTZDpa with PPARγ, which provides diminished conformational stability of the receptor compared with full agonists. In diabetic models it proved as effective as a potent thiazolidinedione full agonist of PPAR-γ, but without the excessive gain in body weight, heart weight and increase in adipose tissue associated with the full agonist. Compound 24 is another structurally novel potent selective PPARγ modulator that has useful efficacy in animal models of diabetes and retains a comparatively improved therapeutic index in rodents with regard to weight gain, adiposity and

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cardiac hypertrophy despite high systemic plasma exposures (Acton et al., 2005). Similarly, a recent report described FK614, a non-thiazolidinedione selective PPARγ modulator that is currently in Phase II clinical trials. This compound has insulin-sensitizing activity similar to that of PPARγ full agonists but induces less hemodilution and cardiomegaly (Minoura et al., 2004). It will be interesting to see if these selective PPARγ modulators are effective in animal models of inflammatory airway disease and whether they will provide a safer alternative treatment for diseases such as asthma and COPD in the clinic. 8. Conclusion There is an unmet clinical need in inflammatory diseases of the airways. Asthma although well controlled by steroid and β agonist treatment has a long term inflammatory component which is not well controlled. COPD is a chronic inflammatory disease of the airways in search of an effective treatment as are diseases such as pulmonary fibrosis and ARDS. It has been suggested that PPARγ agonists may go some way to providing an effective treatment for these chronic inflammatory diseases and the evidence presented here adds weight to that argument. PPARγ does appear to be involved in immune regulation and does appear to possess antiinflammatory properties. Using synthetic and natural PPARγ agonists many groups have demonstrated the anti-inflammatory potential of these compounds both in cell based assays and in vitro studies. Added to this is a growing body of in vivo evidence obtained in many different models of human inflammatory disease such as arthritis, atherosclerosis and inflammatory bowel disease which further supports their use. However, recent evidence has demonstrated the effectiveness of both synthetic and natural PPARγ agonists in models of human inflammatory airway diseases such as asthma, COPD, ARDS and pulmonary fibrosis suggesting that this class of compounds may be exploited for the treatment of these debilitating diseases. Future developments will be watched with interest. References Acton III, J.J., Black, R.M., Jones, A.B., Moller, D.E., Colwell, L., Doebber, T.W., Macnaul, K.L., Berger, J., Wood, H.B., 2005. Benzoyl 2-methyl indoles as selective PPARgamma modulators. Bioorg. Med. Chem. Lett. 15, 357–362. Angeli, V., Hammad, H., Staels, B., Capron, M., Lambrecht, B.N., Trottein, F., 2003. Peroxisome proliferator-activated receptor gamma inhibits the migration of dendritic cells: consequences for the immune response. J. Immunol. 170, 5295–5301. Arakawa, K., Ishihara, T., Aoto, M., Inamasu, M., Kitamura, K., Saito, A., 2004. An antidiabetic thiazolidinedione induces eccentric cardiac hypertrophy by cardiac volume overload in rats. Clin. Exp. Pharmacol. Physiol. 31, 8–13. Asada, K., Sasaki, S., Suda, T., Chida, K., Nakamura, H., 2003. Antiinflammatory roles of peroxisome proliferator-activated receptor gamma in human alveolar macrophages. Am. J. Respir. Crit. Care Med. 169, 195–200. Azuma, Y., Shinohara, M., Wang, P.L., Ohura, K., 2001. 15-deoxy-delta(12,14)prostaglandin J(2) inhibits IL-10 and IL-12 production by macrophages. Biochem. Biophys. Res. Commun. 283, 344–346.

107

Belvisi, M.G., Hele, D.J., Birrell, M.A., 2004. New advances and potential therapies for the treatment of asthma. BioDrugs 18, 211–223. Berger, J., Bailey, P., Biswas, C., Cullinan, C.A., Doebber, T.W., Hayes, N.S., Saperstein, R., Smith, R.G., Leibowitz, M.D., 1996. Thiazolidinediones produce a conformational change in peroxisomal proliferator-activated receptor-gamma: binding and activation correlate with antidiabetic actions in db/db mice. Endocrinology 137, 4189–4195. Berger, J.P., Petro, A.E., Macnaul, K.L., Zhang, B.B., Richards, K., Elbrecht, A., Johnson, B.A., Zhou, G., Doebber, T.W., Biswas, C., Parikh, M., Sharma, N., Tanen, M.R., Thompson, G.M., Ventre, J., Adams, A.D., Mosley, R., Surwit, R.S., Moller, D.E., 2003. Distinct properties and advantages of a novel peroxisome proliferator-activated receptor [gamma] selective modulator. Mol. Endocrinol. 17, 662–676. Birrell, M.A., Patel, H.J., McCluskie, K., Wong, S., Leonard, T., Yacoub, M.H., Belvisi, M.G., 2004. PPAR-γ agonists as therapy for diseases involving airway neutrophilia. Eur. Respir. J. 24, 18–23. Bishop-Bailey, D., Hla, T., 1999. Endothelial cell apoptosis induced by the peroxisome proliferator-activated receptor (PPAR) ligand 15-deoxy-Δ12, 14prostaglandin J2. J. Biol. Chem. 274, 17042–17048. Bishop-Bailey, D., Warner, T.D., 2003. PPARgamma ligands induce prostaglandin production in vascular smooth muscle cells: indomethacin acts as a peroxisome proliferator-activated receptor-gamma antagonist. FASEB J. 17, 1925–1927. Boyault, S., Bianchi, A., Moulin, D., Morin, S., Francois, M., Netter, P., Terlain, B., Bordji, K., 2004. 15-Deoxy-delta(12,14)-prostaglandin J(2) inhibits IL1beta-induced IKK enzymatic activity and IkappaBalpha degradation in rat chondrocytes through a PPARgamma-independent pathway. FEBS Lett. 572, 33–40. Braissant, O., Foufelle, F., Scotto, C., Dauca, M., Wahli, W., 1996. Differential expression of peroxisome proliferator-activated receptors (PPARs): tissue distribution of PPAR-alpha,-beta, and-gamma in the adult rat. Endocrinology 137, 354–366. Burgess, H.A., Daugherty, L.E., Thatcher, T.H., Lakatos, H.F., Redonnet, M., Phipps, R.P., Sime, P.J., 2005. PPAR{gamma} agonists inhibit TGF-{beta} induced pulmonary myofibroblast differentiation and collagen production: implications for therapy of lung fibrosis. Am. J. Physiol., Lung Cell Mol. Physiol. 288, L1146–L1153. Chawla, A., Barak, Y., Nagy, L., Liao, D., Tontonoz, P., Evans, R.M., 2001. PPAR-gamma dependent and independent effects on macrophage-gene expression in lipid metabolism and inflammation. Nat. Med. 7, 48–52. Chinetti, G., Griglio, S., Antonucci, M., Torra, I.P., Delerive, P., Majd, Z., Fruchart, J.C., Chapman, J., Najib, J., Staels, B., 1998. Activation of proliferator-activated receptors alpha and gamma induces apoptosis of human monocyte-derived macrophages. J. Biol. Chem. 273, 25573–25580. Chinetti, G., Fruchart, J.C., Staels, B., 2000. Peroxisome proliferator-activated receptors (PPARs): nuclear receptors at the crossroads between lipid metabolism and inflammation. Inflamm. Res. 49, 497–505. Chung, S.W., Kang, B.Y., Kim, S.H., Pak, Y.K., Cho, D., Trinchieri, G., Kim, T.S., 2000. Oxidized low density lipoprotein inhibits interleukin-12 production in lipopolysaccharide-activated mouse macrophages via direct interactions between peroxisome proliferator-activated receptor-gamma and nuclear factor-kappa. B. J. Biol. Chem. 275, 32681–32687. Clark, R.B., Bishop-Bailey, D., Estrada-Hernandez, T., Hla, T., Puddington, L., Padula, S.J., 2000. The nuclear receptor PPAR gamma and immunoregulation: PPAR gamma mediates inhibition of helper T cell responses. J. Immunol. 164, 1364–1371. Desreumaux, P., Dubuquoy, L., Nutten, S., Peuchmaur, M., Englaro, W., Schoonjans, K., Derijard, B., Desvergne, B., Wahli, W., Chambon, P., Leibowitz, M.D., Colombel, J.F., Auwerx, J., 2001. Attenuation of colon inflammation through activators of the retinoid X receptor (RXR)/ peroxisome proliferator-activated receptor gamma (PPARgamma) heterodimer. A basis for new therapeutic strategies. J. Exp. Med. 827–838. Desvergne, B., Wahli, W., 1999. Peroxisome proliferator-activated receptors: nuclear control of metabolism. Endocr. Rev. 20, 649–688. Dubuquoy, L., Dharancy, S., Nutten, S., Pettersson, S., Auwerx, J., Desreumaux, P., 2002. Role of peroxisome proliferator-activated receptor gamma and retinoid X receptor heterodimer in hepatogastroenterological diseases. Lancet 360, 1410–1418.

108

M.G. Belvisi et al. / European Journal of Pharmacology 533 (2006) 101–109

Evans, R.M., 1988. The steroid and thyroid hormone receptor superfamily. Science 240, 889–895. Fajas, L., Auboeuf, D., Raspe, E., Schoonjans, K., Lefebvre, A.M., Saladin, R., Najib, J., Laville, M., Fruchart, J.C., Deeb, S., Vidal-Puig, A., Flier, J., Briggs, M.R., Staels, B., Vidal, H., Auwerx, J., 1997. The organization, promoter analysis, and expression of the human PPARgamma gene. J. Biol. Chem. 272, 18779–18789. Faveeuw, C., Fougeray, S., Angeli, V., Fontaine, J., Chinetti, G., Gosset, P., Delerive, P., Maliszewski, C., Capron, M., Staels, B., Moser, M., Trottein, F., 2000. Peroxisome proliferator-activated receptor gamma activators inhibit interleukin-12 production in murine dendritic cells. FEBS Lett. 486, 261–266. Fitzpatrick, F.A., Wynalda, M.A., 1983. Albumin-catalyzed metabolism of prostaglandin D2. Identification of products formed in vitro. J. Biol. Chem. 258, 11713–11718. Forman, B.M., Tontonoz, P., Chen, J., Brun, R.P., Spiegelman, B.M., Evans, R.M., 1995. 15-deoxy-delta 12, 14-prostaglandin J2 is a ligand for the adipocyte determination factor PPAR gamma. Cell 83, 803–812. Genovese, T., Cuzzocrea, S., Di Paola, R., Mazzon, E., Mastruzzo, C., Catalano, P., Sortino, M., Crimi, N., Caputi, A.P., Thiemermann, C., Vancheri, C., 2005. Effect of rosiglitazone and 15-deoxy-Delta12, 14-prostaglandin J2 on bleomycin-induced lung injury. Eur. Respir. J. 25, 225–234. Gosset, P., Charbonnier, A.S., Delerive, P., Fontaine, J., Staels, B., Pestel, J., Tonnel, A.B., Trottein, F., 2001. Peroxisome proliferator-activated receptor gamma activators affect the maturation of human monocyte-derived dendritic cells. Eur. J. Immunol. 31, 2857–2865. Hammad, H., de Heer, H.J., Soullie, T., Angeli, V., Trottein, F., Hoogsteden, H.C., Lambrecht, B.N., 2004. Activation of peroxisome proliferatoractivated receptor-gamma in dendritic cells inhibits the development of eosinophilic airway inflammation in a mouse model of asthma. Am. J. Pathol. 164, 263–271. Hetzel, M., Walcher, D., Grub, M., Bach, H., Hombach, V., Marx, N., 2003. Inhibition of MMP-9 expression by PPARgamma activators in human bronchial epithelial cells. Thorax 58, 778–783. Honda, K., Marquillies, P., Capron, M., Dombrowicz, D., 2004. Peroxisome proliferator-activated receptor gamma is expressed in airways and inhibits features of airway remodeling in a mouse asthma model. J. Allergy Clin. Immunol. 113, 882–888. Huang, J.T., Welch, J.S., Ricote, M., Binder, C.J., Willson, T.M., Kelly, C., 1999. Interleukin-4-dependent production of PPAR-gamma ligands in macrophages by 12/15-lipoxygenase. Nature 400, 378–382. Inoue, H., Tanabe, T., Umesono, K., 2000. Feedback control of cyclooxygenase2 expression through PPARgamma. J. Biol. Chem. 275, 28028–28032. Ishibashi, M., Egashira, K., Hiasa, K., Inoue, S., Ni, W., Zhao, Q., Usui, M., Kitamoto, S., Ichiki, T., Takeshita, A., 2002. Antiinflammatory and antiarteriosclerotic effects of pioglitazone. Hypertension 40, 687–693. Issemann, I., Green, S., 1990. Activation of a member of the steroid hormone receptor superfamily by peroxisome proliferators. Nature 347, 645–650. Jiang, C., Ting, A.T., Seed, B., 1998. PPAR-gamma agonists inhibit production of monocyte inflammatory cytokines. Nature 391, 82–86. Kliewer, S.A., Lenhard, J.M., Willson, T.M., Patel, I., Morris, D.C., Lehmann, J.M., 1995. A prostaglandin J2 metabolite binds peroxisome proliferatoractivated receptor gamma and promotes adipocyte differentiation. Cell 83, 813–819. Kodera, Y., Takeyama, K., Murayama, A., Suzawa, M., Masuhiro, Y., Kato, S., 2000. Ligand type-specific interactions of peroxisome proliferator-activated receptor gamma with transcriptional coactivators. J. Biol. Chem. 275, 33201–33204. Law, R.E., Meehan, W., Xi, X.-P., Graf, K., Wuthrich, D.A., Coats, W., Faxon, D., Hsueh, W.A., 1996. Troglitazone inhibits vascular smooth muscle cell growth and intimal hyperplasia. J. Clin. Invest. 98, 1897–1905. Lee, K.S., Park, S.J., Hwang, P.H., Yi, H.K., Song, C.H., Chai, O.H., Kim, J.S., Lee, M.K., Lee, Y.C., 2005. PPAR-gamma modulates allergic inflammation through up-regulation of PTEN. FASEB J. 19, 1033–1035. Li, M., Pascual, G., Glass, C.K., 2000. Peroxisome proliferator-activated receptor gamma-dependent repression of the inducible nitric oxide synthase gene. Mol. Cell Biol. 20, 4699–4707.

Maggi Jr., L.B., Sadeghi, H., Weigand, C., Scarim, A.L., Heitmeier, M.R., Corbett, J.A., 2000. Anti-inflammatory actions of 15-deoxy-delta 12,14prostaglandin J2 and troglitazone: evidence for heat shock-dependent andindependent inhibition of cytokine-induced inducible nitric oxide synthase expression. Diabetes 49, 346–355. Marx, N., Sukhova, G., Murphy, C., Libby, P., Plutzky, J., 1998. Macrophages in human atheroma contain PPARgamma: differentiation-dependent peroxisomal proliferator-activated receptor gamma (PPARgamma) expression and reduction of MMP-9 activity through PPARgamma activation in mononuclear phagocytes in vitro. Am. J. Pathol. 153, 17–23. Minoura, H., Takeshita, S., Ita, M., Hirosumi, J., Mabuchi, M., Kawamura, I., Nakajima, S., Nakayama, O., Kayakiri, H., Oku, T., Ohkubo-Suzuki, A., Fukagawa, M., Kojo, H., Hanioka, K., Yamasaki, N., Imoto, T., Kobayashi, Y., Mutoh, S., 2004. Pharmacological characteristics of a novel nonthiazolidinedione insulin sensitizer, FK614. Eur. J. Pharmacol. 494, 273–281. Moras, D., Gronemeyer, H., 1998. The nuclear receptor ligand-binding domain: structure and function. Curr. Opin. Cell Biol. 10, 384–391. Mueller, C., Weaver, V., Vanden Heuvel, J.P., August, A., Cantorna, M.T., 2003. Peroxisome proliferator-activated receptor gamma ligands attenuate immunological symptoms of experimental allergic asthma. Arch. Biochem. Biophys. 418, 186–196. Oates, J.C., Reilly, C.M., Crosby, M.B., Gilkeson, G.S., 2002. Peroxisome proliferator-activated receptor γ agonists: potential use for treating chronic inflammatory diseases. Arthritis Rheum. 46, 598–605. Patel, H.J., Belvisi, M.G., Bishop-Bailey, Yacoub, M.H., Mitchell, J.A., 2003. Activation of peroxisome proliferator-activated receptors in human airway smooth muscle cells has a superior anti-inflammatory profile to corticosteroids: relevance for chronic obstructive pulmonary disease therapy. J. Immunol. 170, 2663–2669. Ricote, M., Li, A.C., Willson, T.M., Kelly, C.J., Glass, C.K., 1998a. The peroxisome proliferator-activated receptor-gamma is a negative regulator of macrophage activation. Nature 391, 79–82. Ricote, M., Huang, J., Fajas, L., Li, A., Welch, J., Najib, J., Witztum, J.L., Auwerx, J., Palinski, W., Glass, C.K., 1998b. Expression of the peroxisome proliferator-activated receptor gamma (PPARgamma) in human atherosclerosis and regulation in macrophages by colony stimulating factors and oxidized low density lipoprotein. Proc. Natl. Acad. Sci. U. S. A. 95, 7614–7619. Serhan, C.N., 1996. Inflammation. Signalling the fat controller. Nature 384, 23–24. Serhan, C.N., Devchand, P.R., 2001. Novel anti-inflammatory targets for asthma. A role for PPARγ? Am. J. Respir. Cell Mol. Biol. 24, 658–661. Sher, J.U., Pillinger, M.H., 2005. 15d-PGJ2: the anti-inflammatory prostaglandin? Clin. Immunol. 114, 100–109. Spiegelman, B.M., Flier, J.S., 1996. Adipogenesis and obesity: rounding out the big picture. Cell 87, 377–389. Tontonoz, P., Nagy, L., Alvarez, J.G., Thomazy, V.A., Evans, R.M., 1998. PPARgamma promotes monocyte/macrophage differentiation and uptake of oxidized LDL. Cell 93, 241–252. Trifillieff, A., Bench, A., Hanley, M., Bayley, D., Campbell, E., Whittaker, P., 2003. PPAR-α and -γ but not -δ agonists inhibit airway inflammation in a murine model of asthma: in vitro evidence for an NF-κB-independent model. Br. J. Pharmacol. 139, 163–171. Ueki, S., Matsuwaki, Y., Kayaba, H., Oyamada, H., Kanda, A., Usami, A., Saito, N., Chihara, J., 2004. Peroxisome proliferator-activated receptor gamma regulates eosinophil functions: a new therapeutic target for allergic airway inflammation. Int. Arch. Allergy Immunol. 134 (Suppl 1), 30–36. Vidal-Puig, A., Jiminez-Linan, M., Lowell, B.B., Hamann, A., Hu, E., Spiegelman, B., Flier, J.S., Moller, D.E., 1996. Regulation of PPAR gamma gene expression by nutrition and obesity in rodents. J. Clin. Invest. 97, 2553–2561. Von Knethen, A., Brune, B., 2003. PPARgamma—an important regulator of monocyte/macrophage function. Arch. Immunol. Ther. Exp. (Warsz.) 51, 219–226. Wang, A.C., Dai, X., Luu, B., Conrad, D.J., 2001. Peroxisome proliferatoractivated receptor-gamma regulates airway epithelial cell activation. Am. J. Respir. Cell Mol. Biol. 24, 688–693.

M.G. Belvisi et al. / European Journal of Pharmacology 533 (2006) 101–109 Willson, T.M., Brown, P.J., Sternbach, D.D., Henke, B.R., 2000. The PPARs: from orphan receptors to drug discovery. J. Med. Chem. 43, 527–550. Woerly, G., Honda, K., Loyens, M., Papin, J.P., Auwerx, J., Staels, B., Capron, M., Dombrowicz, D., 2003. Peroxisome proliferator-activated receptors alpha and gamma down-regulate allergic inflammation and eosinophil activation. J. Exp. Med. 198, 411–421.

109

Yang, X.A., Wang, L.H., Chen, T., Hodge, D.R, Resau, J.H., DaSilva, L., Farrar, W.L., 2000. Activation of human T lymphocytes is inhibited by peroxisome proliferator-activated receptor gamma (PPARgamma) agonists. PPARgamma co-association with transcription factor NFAT. J. Biol. Chem. 275, 4541–4544. Yki-Jarvinen, H., 2004. Thiazolidinediones. N. Engl. J. Med. 351, 1106–1118.