The Interleukin-1 receptor antagonist is a direct target gene of PPARα in liver☆
Introduction
Inflammation describes the comprehensive reaction of the host to various types of injury, which is generally protective and aimed at promoting tissue repair. The inflammatory response is mediated by a diverse group of cytokines and other signaling molecules that are able to profoundly influence cellular function. At the cellular level, numerous signaling pathways and transcription factors conspire in a complex network to produce the appropriate response. In recent years it has become clear that the Peroxisome Proliferator-Activated Receptors (PPARs) modulate this response in a variety of organs. PPARs are members of the superfamily of Nuclear Receptors that play a pivotal role in mediating the effect of small lipophilic ligands on gene transcription [1]. The three isotypes of the PPAR-family, PPARα, PPARβ/δ and PPARγ, have been implicated in numerous processes, including lipid and glucose metabolism, and inflammation. Activation of the receptor occurs by binding of various ligands, ranging from natural compounds such as fatty acids to highly specific synthetic agonists. Upon ligand-activation, binding to so called PPAR Response Elements (PPRE) located in the promoter of target genes results in increased gene transcription. To accomplish activation of gene transcription it is essential that PPAR forms a heterodimer with the Retinoid X Receptor (RXR) [2].
Besides their ability to enhance gene transcription [3], PPARs are also able to suppress gene expression. For example, it has been shown that activated PPARα lowers the expression of several enzymes connected with amino acid metabolism [4], although the mechanism behind the observed down-regulation remains unknown. In addition, the effects of PPARα on inflammation are mainly achieved by suppressing gene expression. Since the initial observation that PPARα−/− mice have a prolonged inflammatory response [5], an important role for PPARs in regulating inflammatory responses has clearly emerged. Although numerous studies have demonstrated the protective and anti-inflammatory effects of PPARα-activation in liver [6], [7], information about the precise molecular mechanisms involved is somewhat limited. One of the mechanisms by which this nuclear receptor exerts its anti-inflammatory action is through modulation of the NF-κB pathway. Physical interaction of PPARα with NF-κB prevents its activation and downstream pro-inflammatory effects [8]. Moreover, PPARα has been shown to up-regulate the expression of IκB, the natural NF-κB inhibitor that prevents the nuclear translocation and activation of the pro-inflammatory transcription factor [9].
Anti-inflammatory properties have also been assigned to the other two PPAR isotypes. Activation of PPARγ controls the inflammatory status of the intestinal tract [10] and is responsible for the down-regulation of a specific subset of pro-inflammatory genes in macrophages [11]. The recent generation of macrophages lacking PPARβ/δ has also revealed a specific role for PPARβ/δ in regulating inflammatory processes [12].
To better understand the regulatory role of PPARα in liver and to identify possible new target genes under the control of PPARα, we studied PPARα-dependent gene expression levels in mouse liver by means of Affymetrix microarray analysis. Activation of PPARα was achieved by treating Wildtype (Wt) and PPARα −/− mice with the synthetic PPARα agonist Wy-14643. While numerous inflammatory genes were found to be down regulated by PPARα activation, the IL-1 receptor antagonist, however, was highly up-regulated by PPARα. Additional experiments indicated that the IL-1 receptor antagonist is a direct target gene of PPARα. Our data suggest that PPARα may modulate inflammation by direct up-regulation of target genes.
Section snippets
Chemicals
Wy14643 was obtained from ChemSyn Laboratories (Lenexa, Kanasa). Cell culture medium, fetal calf serum and penicillin/streptomycin/fungizone were from Cambrex Bioscience (Seraing, Belgium). SYBR green was from Eurogentec (Seraing, Belgium).
The human and mouse antibody against IL-1ra and the recombinant hIL-1β were from R&D systems (R&D Systems Europe Ltd., Abingdon, UK). Otherwise, chemicals were from Sigma–Aldrich (Zwijndrecht, The Netherlands).
Animal experiments
Sv129 PPARα−/− mice and corresponding Wt mice
PPARα, but not PPARβ/δ and PPARγ, is involved in the negative regulation of inflammatory related-genes in liver
To screen for potential novel target genes of PPARα involved in inflammation in liver we performed micro-array analysis. A comparison was made between liver RNA from Wt and PPARα−/− mice treated or not with the synthetic PPARα ligand Wy14643. The expression of a large number of genes involved in inflammation was decreased by Wy14643 in Wt but not PPARα−/− mice (Fig. 1A). As shown in Fig. 1B, the expression of all of these genes was up-regulated during Lipopolysaccharide (LPS)-induced acute
Discussion
Since the initial discovery indicating that PPARα is involved in inflammation [5], considerable progress has been made towards resolving the regulatory role of PPARα during inflammation [8], [9], [20]. It is now clear that PPARα mainly governs inflammation by down-regulating expression of genes, including several acute phase genes.
Here, we show that soluble Interleukin-1 receptor antagonist is a direct target gene of PPARα with a functional PPRE in its promoter, suggesting that PPARα may also
Acknowledgements
We thank Jolanda van der Meijde for performing the microarray experiment and René Bakker for assisting with the animal experiments. We thank Dr. Mark Leibowitz (Ligand Pharmaceuticals) for the kind gift of Lg100268. This study was supported by the Centre for Human Nutrigenomics, with additional support by the Netherlands Organization for Scientific Research (NWO), the Wageningen Center for Food Sciences, the Dutch Nutrigenomics Consortium, the Swiss National Science Foundation and Etat de Vaud
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The authors who have taken part in this study declared that they have no relationship with the manufacturers of the drugs involved either in the past or present and did not receive funding from the manufacturers and from any source to carry out their research.