Constitutive Activation of Peroxisome Proliferator-activated Receptor-γ Suppresses Pro-inflammatory Adhesion Molecules in Human Vascular Endothelial Cells

Abstract

Peroxisome proliferator-activated receptor-γ (PPAR-γ) is a ligand-activated nuclear receptor that has an essential role in adipogenesis and glucose homeostasis. PPAR-γ is expressed in vascular tissues including endothelial cells (ECs). PPAR-γ activity can be regulated by many pathophysiological and pharmacological agonists. However, the role of PPAR-γ activation in ECs remains unclear. In this study, we examined the effect of the constitutive activation of PPAR-γ on the phenotypic modulation of ECs. Adenovirus-mediated expression of a constitutively active mutant of PPAR-γ resulted in significant ligand-independent activation of PPAR-γ and specific induction of the PPAR-γ target genes. However, PPAR-γ activation significantly suppressed the expression of vascular adhesion molecules in ECs and the ensuing leukocyte recruitment. Furthermore, constitutive activation of PPAR-γ resulted in simultaneous repression of AP-1 and NF-κB activity, which suggests that PPAR-γ may reduce pro-inflammatory phenotypes via, at least in part, suppression of the AP-1 and NF-κB pathways. Therefore, using a gain-of-function approach, our study provides novel evidence showing that constitutive activation of PPAR-γ is sufficient to prevent ECs from converting into a pro-inflammatory phenotype. These results also suggest that, in addition to pharmacological agonists, the genetic modification of the PPAR-γ activity in ECs may be a potential approach for therapeutic intervention in various inflammatory disorders. Peroxisome proliferator-activated receptor-γ (PPAR-γ) is a ligand-activated nuclear receptor that has an essential role in adipogenesis and glucose homeostasis. PPAR-γ is expressed in vascular tissues including endothelial cells (ECs). PPAR-γ activity can be regulated by many pathophysiological and pharmacological agonists. However, the role of PPAR-γ activation in ECs remains unclear. In this study, we examined the effect of the constitutive activation of PPAR-γ on the phenotypic modulation of ECs. Adenovirus-mediated expression of a constitutively active mutant of PPAR-γ resulted in significant ligand-independent activation of PPAR-γ and specific induction of the PPAR-γ target genes. However, PPAR-γ activation significantly suppressed the expression of vascular adhesion molecules in ECs and the ensuing leukocyte recruitment. Furthermore, constitutive activation of PPAR-γ resulted in simultaneous repression of AP-1 and NF-κB activity, which suggests that PPAR-γ may reduce pro-inflammatory phenotypes via, at least in part, suppression of the AP-1 and NF-κB pathways. Therefore, using a gain-of-function approach, our study provides novel evidence showing that constitutive activation of PPAR-γ is sufficient to prevent ECs from converting into a pro-inflammatory phenotype. These results also suggest that, in addition to pharmacological agonists, the genetic modification of the PPAR-γ activity in ECs may be a potential approach for therapeutic intervention in various inflammatory disorders. peroxisome proliferator-activated receptor endothelial cell(s) thiazolidinedione human umbilical cord vein endothelial cell(s) intercellular adhesion molecule vascular adhesion molecule electrophoretic mobility shift assay tumor necrosis factor phorbol 12-myristate 13-acetate herpes simplex virus adenovirus green fluorescence protein glyceraldehyde-3-phosphate dehydrogenase PPAR-responsive element Peroxisome proliferator-activated receptor-γ (PPAR-γ)1 is a ligand-activated nuclear receptor that has an essential role in adipogenesis and glucose homeostasis. PPAR-γ is also expressed in vessel wall tissue, including endothelial cells (ECs) (1Marx N. Bourcier T. Sukhova G.K. Libby P. Plutzky J. Arterioscler. Thromb. Vasc. Biol. 1999; 19: 546-551Crossref PubMed Scopus (338) Google Scholar). However, the role of PPAR-γ activation in ECs remains unclear. Previous studies have shown that PPAR-γ agonists such as 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) and the thiazolidinedione (TZD) class of insulin-sensitizing drugs can modulate the expression of many pro-inflammatory cytokines (2Ricote M., Li, A.C. Willson T.M. Kelly C.J. Glass C.K. Nature. 1998; 391: 79-82Crossref PubMed Scopus (3302) Google Scholar), chemokines (3Marx N. Mach F. Sauty A. Leung J.H. Sarafi M.N. Ransohoff R.M. Libby P. Plutzky J. Luster A.D. J. Immunol. 2000; 164: 6503-6508Crossref PubMed Scopus (298) Google Scholar), and adhesion molecules (4Pasceri V., Wu, H.D. Willerson J.T. Yeh E.T. Circulation. 2000; 101: 235-238Crossref PubMed Scopus (504) Google Scholar) in macrophages and many other cell types including ECs. However, the observed anti-inflammatory effects often vary according to the agonists used and are not always consistent with their capacity for receptor binding (5Delerive P. Fruchart J.C. Staels B. J. Endocrinol. 2001; 169: 453-459Crossref PubMed Scopus (669) Google Scholar). Furthermore, recent studies indicate that the receptor-independent effects exist both for 15d-PG-J2 and TZDs (6Chawla A. Barak Y. Nagy L. Liao D. Tontonoz P. Evans R.M. Nat. Med. 2001; 7: 48-52Crossref PubMed Scopus (972) Google Scholar, 7Straus D.S. Pascual G., Li, M. Welch J.S. Ricote M. Hsiang C.H. Sengchanthalangsy L.L. Ghosh G. Glass C.K. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 4844-4849Crossref PubMed Scopus (956) Google Scholar). Therefore, whether constitutive activation of the PPAR-γ per se, regardless of pleiotropic ligand-specific effects, has an anti-inflammatory effect remains to be investigated. The aim of this study was to examine whether a constitutively active PPAR-γ can modulate the endothelial expression of pro-inflammatory phenotype such as the induction of adhesion molecules and recruitment of leukocytes. The constitutively active mutant of PPAR-γ1 (VP-PPAR-γ) was constructed by fusing the herpes virus VP16 activation domain to wild-type PPAR-γ1, and delivered into human umbilical cord vein endothelial cells (HUVECs) with use of a tetracycline-controlled adenoviral system. The overexpression of VP-PPAR-γ resulted in the ligand-independent activation of PPAR-γ and specific induction of the PPAR-γ target genes. However, the induced expression of intercellular adhesion molecule-1 (ICAM-1), vascular adhesion molecule-1 (VCAM-1), and E-selectin was significantly suppressed in the VP-PPAR-γ−transduced ECs. Consequently, endothelial recruitment of monocytes was markedly attenuated by constitutive activation of the PPAR-γ. Results from electrophoretic mobility shift assays (EMSA) and reporter transfection assays demonstrated that VP-PPAR-γ in ECs resulted in a simultaneous decrease in AP-1 and NF-κB activity. These results demonstrate that activation of endothelial PPAR-γ has a potent anti-inflammatory role. HUVECs were harvested by collagenase treatment of umbilical cord veins and cultured on plates coated with collagen. Cells were maintained in M199 supplemented with 20% fetal bovine serum, 20 mm HEPES (pH 7.4), 1 ng/ml of recombinant human fibroblast growth factor, and 90 μg/ml of heparin and antibiotics. In all the experiments, cells within three passages were used. THP-1, a human monocyte cell line (ATCC), was grown in RPMI 1640 containing 10% fetal bovine serum. Recombinant human tumor necrosis factor (TNF)-α was from Becton-Dickinson. Phorbol 12-myristate 13-acetate (PMA) and tetracycline were purchased from Sigma. VP-PPAR-γ contains the 78 amino acid herpes simplex virus (HSV) VP16 transactivation domain fused to the N terminus of mouse PPAR-γ1 (GenBankTM accession number U10374). To generate the adenoviruses expressing VP-PPAR-γ (Ad-VP-PPAR-γ), the cDNA fragment containing VP-PPAR-γ was subcloned into a shuttle plasmid pAdlox and recombined with an E1-and E3-deleted ψ 5 viral DNA in CRE8 cells (8Hardy S. Kitamura M. Harris-Stansil T. Dai Y. Phipps M.L. J. Virol. 1997; 71: 1842-1849Crossref PubMed Google Scholar). The expression of the inserted gene was driven by a 7 × tet/minimal cytomegalovirus promoter that was further under the control of an artificial tetracycline-responsive transactivator (tTA). The adenoviruses expressing green fluorescence protein (Ad-GFP) and tTA were previously described (9Wang N. Verna L. Liao H. Ballard A. Zhu Y. Stemerman M.B. Arterioscler. Thromb. Vasc. Biol. 2001; 21: 1414-1420Crossref PubMed Scopus (35) Google Scholar). The adenoviruses were plaque-purified, expanded, titrated in 293 cells, and purified by cesium chloride methods. For adenovirus-mediated gene transfer, confluent HUVECs were exposed to adenoviral vectors at a multiplicity-of-infection rate of ∼50–100 for 2 h (Ad-tTA was co-infected to induce the tetracycline controllable expression). After the viruses were washed off, infected cells were further incubated for the indicated time in the presence or absence of tetracycline. The PPRE-TK-Luc is a luciferase reporter containing the herpes virus thymidine kinase promoter (−105/+51) downstream of three copies of PPAR response elements from the acyl-CoA oxidase gene. The pCMX-VP16 is a plasmid encoding the VP16 transactivation domain. Plasmids for the yeast transcription factor Gal4 and 4 × UAS-TK-luc were described previously (9Wang N. Verna L. Liao H. Ballard A. Zhu Y. Stemerman M.B. Arterioscler. Thromb. Vasc. Biol. 2001; 21: 1414-1420Crossref PubMed Scopus (35) Google Scholar). pAP-1-luc and pNF-κB-luc are luciferase reporters containing 7 × AP-1 binding sites or 5 × NF-κB sites in tandem (Stratagene, La Jolla, CA). The ICAM-1 promoter reporter ICAM-1 (−445)-luc was constructed by subcloning a 445-bp 5′-flanking region of the human ICAM-1 gene (provided by Thomas Parks, Boehringer Ingelheim Pharmaceuticals, Ridgefield, CT) (10Ledebur H.C. Parks T.P. J. Biol. Chem. 1995; 270: 933-943Abstract Full Text Full Text PDF PubMed Scopus (519) Google Scholar) into pGL3-basic luciferase reporter plasmid (Promega) (9Wang N. Verna L. Liao H. Ballard A. Zhu Y. Stemerman M.B. Arterioscler. Thromb. Vasc. Biol. 2001; 21: 1414-1420Crossref PubMed Scopus (35) Google Scholar). Mutagenesis of the AP-1/NF-κB site in the human ICAM-1 promoter region was performed with the QuikChangeTM polymerase chain reaction-based method (Stratagene) as described previously (9Wang N. Verna L. Liao H. Ballard A. Zhu Y. Stemerman M.B. Arterioscler. Thromb. Vasc. Biol. 2001; 21: 1414-1420Crossref PubMed Scopus (35) Google Scholar). Mutagenic primers were used to introduce mutations (lowercase letters) at the AP-1-like site (−321, in relation to the start codon) and the NF-κB-like site (−223) (underlined) in the ICAM-1 promoter region: AP-1m, 5′-GCGGTGTAGACCGTGgTgCcAGCTTAGCCTGGCCG-3′ and NF-κBm, 5′-CGATTGCTTTAGCTTGGAAAcattGGAGCTGAAGCGGCC-3′. Following transformation, amplification, selection, and screening, the mutagenic exchanges were confirmed by sequencing. Plasmids were introduced into HUVECs with use of a cationic lipid-based transfection reagent Targefect (Targeting System, San Diego, CA). At the indicated time posttransfection, cell lysates were harvested to measure luciferase activity. The β-galactosidase activity was also measured to normalize transfection efficiency. Total RNA was isolated using Trizol reagent (Invitrogen), fractionated on a formaldehyde-agarose gel, transferred to a nylon membrane, and hybridized to random-primed cDNA probes for human ICAM-1, VCAM-1, E-selectin, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) genes. The cDNA probes for human CD36 were synthesized by reverse-transcriptase PCR as previously described (11Han J. Hajjar D.P. Tauras J.M. Feng J. Gotto A.M., Jr. Nicholson A.C. J. Biol. Chem. 2000; 275: 1241-1246Abstract Full Text Full Text PDF PubMed Scopus (168) Google Scholar). Sequences of the primers were CD36 (forward): 5′-ATGGGCTGTGACCGGAACT (285–304); (reverse): 5′-ACAGACCAACTGTGGTAG (871–889). Cellular proteins were extracted from HUVECs as previously described (12Wang N. Verna L. Hardy S. Forsayeth J. Zhu Y. Stemerman M.B. Arterioscler. Thromb. Vasc. Biol. 1999; 19: 2078-2084Crossref PubMed Scopus (68) Google Scholar). Protein concentration was measured with the BCA protein assay kit (Pierce). Protein samples were resolved on SDS-PAGE, transferred onto ImmobilonTM-P membrane (Millipore, Bedford, Massachusetts) and analyzed with use of a rabbit polyclonal antibody to PPAR-γ (H-100, Santa Cruz, California) and an horseradish peroxidase-conjugated secondary antibody (sheep anti-rabbit, 1:5000, Sigma) followed by ECL detection (AmershamBiosciences). The nuclear extracts were first mixed with 1 μg of poly (dI-dC) in DNA binding buffer and incubated at room temperature for 10 min.32P-labeled oligonucleotides were then added to the reaction and incubated for 20 min at room temperature. The oligonucleotides were end labeled with use of T4 polynucleotide kinase and [γ-32P]adenosine triphosphate (ICN). The sequences of oligonucleotides are as follows (sequences of the complementary strands are omitted): 1) PPRE (Aco oxidase), 5′-GGGGACCAGGACAAAGGTCACGTTCGGGAGCT-3′; 2) AP-1 (consensus), 5′-CGCTTGATGAGTCAGCCGGAA-3′; and 3) NF-κB (consensus), 5′-AGTTGAGGGGACTTTCCCAGGC-3′. Human monocytic cells, THP-1, were infected with Ad-GFP and Ad-tTA. Twenty-four hours post-infection, the THP-1 cells were washed and co-incubated for 30 min with control or activated HUVECs. Unbound leukocytes were removed by washing, the adhered cells were fixed with 4% paraformaldehyde, and the number of adhered cells was visualized by fluorescence microscopy as described previously (9Wang N. Verna L. Liao H. Ballard A. Zhu Y. Stemerman M.B. Arterioscler. Thromb. Vasc. Biol. 2001; 21: 1414-1420Crossref PubMed Scopus (35) Google Scholar). To quantify the THP-1 adhesion, the cells (THP-1 and HUVECs) were harvested, solubilized, and measured with use of a fluorescence concentration analyzer (Pandex, IDEXX) as previously described (13Smalley D.M. Lin J.H. Curtis M.L. Kobari Y. Stemerman M.B. Pritchard K.A., Jr. Arterioscler. Thromb. Vasc. Biol. 1996; 16: 585-590Crossref PubMed Scopus (100) Google Scholar). Quantitative data are expressed as mean ± S.E. Statistical analysis was performed with use of the Student’s t test. Differences were considered significant when probability values were less than 0.05. To examine the role of PPAR-γ in EC phenotypic modulation, we constructed a constitutively active form of PPAR-γ by fusing a herpes virus VP16 transactivation domain to the wild-type PPAR-γ1 cDNA. A recombinant adenovirus expressing VP-PPAR-γ was made to ensure a ubiquitous yet controllable expression of VP-PPAR-γ in ECs (Fig. 1A). As shown by RNA hybridization and immunoblotting (Fig. 1B), overexpression of VP-PPAR-γ was achieved by infection of HUVECs with Ad-VP-PPAR-γ and Ad-tTA and switched off by the addition of tetracycline in the culture medium. Furthermore, the overexpression of VP-PPAR-γ specifically induced DNA binding to Aco-PPRE, which could be blocked by a specific cold competitor (Fig.2A). VP-PPAR-γ did not bind to the sequences for other transcription factors such as AP-1, NF-κB, AP-2, and GATA (data not shown). In addition, a transient reporter assay was performed to assess the functionality of VP-PPAR-γ to induce the PPRE-dependent gene expression. As shown in Fig. 2B, adenovirus-mediated expression of VP-PPAR-γ caused a more than 10-fold increase in the PPRE-driven luciferase expression in the absence of exogenous PPAR-γ ligands. It is thus demonstrated that adenovirus-mediated expression of VP-PPAR-γ can be used as a model for constitutive PPAR-γ activation in to the PPRE and the gene in results showing overexpression of VP-PPAR-γ the binding to PPRE oligonucleotides μg of nuclear extracts per induction of the in HUVECs infected with Ad-VP-PPAR-γ and Ad-tTA in the presence or absence of tetracycline. Cells in plates were with the plasmid for β-galactosidase was to normalize the transfection efficiency. were activity, expressed as the induction of the activity. mean ± S.E. of three experiments, performed in VP-PPAR-γ activity the induced expression of adhesion molecules is a phenotypic of EC activation and a of many pro-inflammatory we examined the effect of the PPAR-γ constitutive activation on the gene expression of ICAM-1, VCAM-1, and E-selectin in response to pro-inflammatory agonists and results that overexpression of the VP-PPAR-γ significantly suppressed the induction of adhesion molecules by and expression of the gene was not In the expression of an target gene of was constitutive activation of PPAR-γ the pro-inflammatory in ECs. The induction of adhesion molecules is essential for leukocyte to the vessel wall and for inflammatory in various and Therefore, we the adhesion assay to whether the constitutive activation of PPAR-γ such a in The monocytic cells were infected to a green fluorescence protein for use in and by fluorescence microscopy or fluorescence As shown in adhesion was induced by HUVECs with or However, constitutive activation of PPAR-γ in ECs significantly attenuated the recruitment of leukocytes by pro-inflammatory is that an increase in PPAR-γ activity is sufficient to the expression of adhesion molecules as a to prevent the from converting into a pro-inflammatory To the constitutive activation of PPAR-γ the repression of the adhesion we examined the effect of VP-PPAR-γ on the activation of AP-1 and NF-κB, which are to be the transcription factors endothelial As in the constitutive activation of PPAR-γ markedly both and gene expression in the transient reporter However, did not the yeast transcription factor gene expression. effect was also demonstrated in results from In the cells, the AP-1 and NF-κB binding activity (Fig. ICAM-1 promoter assays were performed to whether the simultaneous of AP-1 and NF-κB by the constitutively active PPAR-γ can for the suppression of the vascular adhesion As shown in the induced ICAM-1 promoter activity was significantly by of the VP-PPAR-γ not the VP16 activation However, the AP-1 and NF-κB binding sites the ICAM-1 promoter induction as as the effect of the PPAR-γ. demonstrated that a simultaneous of AP-1 and NF-κB activity is sufficient to the ICAM-1 induction and may to the effect of VP-PPAR-γ on the induction of the adhesion molecules and the ensuing ICAM-1 induction AP-1 and NF-κB assays were performed by of ICAM-1 (−445)-luc or ICAM-1 with pCMX-VP16 or After HUVECs were exposed to or control for h harvested for luciferase and β-galactosidase induction ± 3) of activity. VP-PPAR-γ an anti-inflammatory role for PPAR-γ has many of the PPAR-γ reduce inflammatory in and in (3Marx N. Mach F. Sauty A. Leung J.H. Sarafi M.N. Ransohoff R.M. Libby P. Plutzky J. Luster A.D. J. Immunol. 2000; 164: 6503-6508Crossref PubMed Scopus (298) Google V., Wu, H.D. Willerson J.T. Yeh E.T. Circulation. 2000; 101: 235-238Crossref PubMed Scopus (504) Google Scholar). and PPAR-γ such as 15d-PG-J2 and TZDs were shown to the expression of pro-inflammatory in monocyte and other cell have demonstrated to be in of many inflammatory including A.C. Willson T.M. Glass C.K. J. 2000; PubMed Scopus Google Scholar), Y. M. Y. A. D. M. T. H. J. 2000; PubMed Scopus Google Scholar) and inflammatory A. S. J. 1999; PubMed Scopus Google Scholar). However, the anti-inflammatory of the PPAR-γ not their to the receptor and often pleiotropic other than PPAR-γ agonists, receptor-independent have For of the gene expressing PPAR-γ in macrophages did not or Furthermore, of PPAR-γ can in macrophages PPAR-γ (6Chawla A. Barak Y. Nagy L. Liao D. Tontonoz P. Evans R.M. Nat. Med. 2001; 7: 48-52Crossref PubMed Scopus (972) Google Scholar). the is whether PPAR-γ the various receptor-independent effects of a of is by sufficient to a pro-inflammatory phenotype. In this study, using a regulated adenoviral to the constitutively active PPAR-γ in we demonstrated a potent effect of this receptor on the endothelial expression of pro-inflammatory adhesion molecules and the ensuing leukocyte recruitment. provides novel evidence for a role in endothelial phenotypic which suggests that modification of the PPAR-γ receptor may be a potential anti-inflammatory The EC expression of adhesion molecules and recruitment of leukocytes to the vessel wall are in the response and in inflammatory N. J. Med. 1996; PubMed Scopus Google Scholar). The expression of adhesion molecules has to be regulated by various agonists for which is expressed in vessel tissues including the However, with to a role for PPAR-γ in the induction of endothelial adhesion data have that 15d-PG-J2 and markedly attenuated the expression of and ICAM-1 not E-selectin and in HUVECs (4Pasceri V., Wu, H.D. Willerson J.T. Yeh E.T. Circulation. 2000; 101: 235-238Crossref PubMed Scopus (504) Google Scholar). that, in the or was by the PPAR-γ and by L.L. Arterioscler. Thromb. Vasc. Biol. 1999; 19: PubMed Scopus Google Scholar). described as both and ICAM-1 expression in an tumor cell line D. 1999; PubMed Scopus Google Scholar). At the is for It is that in cell types and may for the observed In addition, the results may also be to the receptor-independent However, we have shown that constitutive activation of PPAR-γ in ECs the expression of adhesion molecules by or can be that pharmacological activation of PPAR may an anti-inflammatory effect the receptor-independent effects are As a transcription factor, the for PPAR-γ to gene expression is binding to the specific the PPAR-responsive element in the promoter region of a target gene. as the constitutively active PPAR-γ induced the gene expression of which has a PPRE in promoter and is considered to be a target gene P. Nagy L. Evans R.M. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). of the CD36 expression demonstrated both functionality and of the expressed However, the PPRE was not in the 5′-flanking of the ICAM-1, VCAM-1, and E-selectin the for the of vascular adhesion molecules by VP-PPAR-γ may not be the as that for the CD36 have that the constitutive activation of PPAR-γ the AP-1 and NF-κB activation in ECs. In addition, the VP-PPAR-γ AP-1 and NF-κB DNA which may to in their It is that both AP-1 and NF-κB in the of ICAM-1 and other pro-inflammatory T. J. 2001; PubMed Scopus Google Scholar). and NF-κB-like have in the 445-bp of the 5′-flanking region of the ICAM-1 gene and to be for ICAM-1 induction by and (10Ledebur H.C. Parks T.P. J. Biol. Chem. 1995; 270: 933-943Abstract Full Text Full Text PDF PubMed Scopus (519) Google Scholar, N. Verna L. Hardy S. Forsayeth J. Zhu Y. Stemerman M.B. Arterioscler. Thromb. Vasc. Biol. 1999; 19: 2078-2084Crossref PubMed Scopus (68) Google Scholar). In this study, we demonstrated that mutations at sites are sufficient to the ICAM-1 induction as as the effect by Therefore, is that the simultaneous of AP-1 and NF-κB may for of the by which the constitutively active PPAR-γ endothelial adhesion molecules and pro-inflammatory In the study, the constitutively active PPAR-γ1 is a containing the VP16 activation domain from of this in ECs to PPAR-γ activation in the absence of exogenous or approach is to the receptor-independent effects that PPAR agonists may The anti-inflammatory effect described is to be to a effect of the VP16 transactivation domain VP-PPAR-γ did not the transcription as demonstrated by the use of a reporter gene (Fig. more gene induction of ICAM-1 was by the VP-PPAR-γ not by the VP16 activation domain (Fig. In addition, other studies that infection pro-inflammatory vascular adhesion molecules and activity A. P. P. N. J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar, J. Virol. 2001; PubMed Scopus Google Scholar, J. Med. 1996; PubMed Scopus Google Scholar, G. B. J. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). the that the anti-inflammatory effect is with the constitutive activation of PPAR-γ. PPAR-γ such as 15d-PG-J2 are to AP-1 and NF-κB DNA binding both the and D.S. Pascual G., Li, M. Welch J.S. Ricote M. Hsiang C.H. Sengchanthalangsy L.L. Ghosh G. Glass C.K. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 4844-4849Crossref PubMed Scopus (956) Google Scholar). In this the PPAR-γ constitutive activation model described in this study may the in a ligand-activated The receptor-independent may for pleiotropic effects of pharmacological PPAR agonists. For the time the to a of transcription factors unclear. These are for the expression of VP-PPAR-γ in this study be in the pathophysiological of this receptor in vessel our that a constitutively active form of PPAR-γ endothelial activation and leukocyte recruitment may an approach to control the vascular inflammatory In we have demonstrated a anti-inflammatory role for PPAR-γ. PPAR-γ activity in ECs may have potential in the treatment of various pro-inflammatory including and inflammatory