Transcriptional Regulation of the Platelet-derived Growth Factor α Receptor Gene via CCAAT/Enhancer-binding Protein-δ in Vascular Smooth Muscle Cells

Inflammatory cytokines stimulate the proliferation of vascular smooth muscle cells (VSMC) and play a pivotal role in the pathogenesis of vascular diseases including atherosclerosis and restenosis. Mitogenic response of interleukin-1β (IL-1β) on VSMC is thought to be mediated by induction of endogenous platelet-derived growth factor (PDGF), especially PDGF-AA. Although the action of PDGF-AA is mediated by its specific receptor, PDGFα-receptor (PDGFαR), very little is known about the regulatory mechanism of PDGFαR gene expression in VSMC. To understand the mechanism, we studied the transcriptional control of the PDGFαR gene in VSMC after treatment with IL-1β. IL-1β (10 ng/ml) drastically increased both PDGFαR and CCAAT/enhancer-binding protein δ (C/EBPδ) mRNA levels in a time dependent manner. A rapid induction of C/EBPδ mRNA within 30 min was followed by slower emergence of PDGFαR mRNA, which reached the maximum level in 12 h, whereas C/EBPδ mRNA was detectable at 30 min and reached the maximum level at 3 h. Electromobility shift and supershift assays revealed that IL-1β markedly increased DNA-protein complex, which was mainly composed of C/EBPβ and/or -δ. Both Western blotting and immunohistochemistry demonstrated that either C/EBPβ or -δ expression was induced by IL-1β exclusively in nuclei of VSMC. On the other hand, overexpression of C/EBPδ specifically transactivated the promoter activity of the PDGFαR gene and significantly enhanced VSMC proliferation in PDGF-treated cells. We conclude that induction of PDGFαR expression is mainly mediated by C/EBPδ expression in VSMC, and a high level of C/EBPδ expression may be involved in the pathogenesis of atherosclerosis and restenosis. Inflammatory cytokines stimulate the proliferation of vascular smooth muscle cells (VSMC) and play a pivotal role in the pathogenesis of vascular diseases including atherosclerosis and restenosis. Mitogenic response of interleukin-1β (IL-1β) on VSMC is thought to be mediated by induction of endogenous platelet-derived growth factor (PDGF), especially PDGF-AA. Although the action of PDGF-AA is mediated by its specific receptor, PDGFα-receptor (PDGFαR), very little is known about the regulatory mechanism of PDGFαR gene expression in VSMC. To understand the mechanism, we studied the transcriptional control of the PDGFαR gene in VSMC after treatment with IL-1β. IL-1β (10 ng/ml) drastically increased both PDGFαR and CCAAT/enhancer-binding protein δ (C/EBPδ) mRNA levels in a time dependent manner. A rapid induction of C/EBPδ mRNA within 30 min was followed by slower emergence of PDGFαR mRNA, which reached the maximum level in 12 h, whereas C/EBPδ mRNA was detectable at 30 min and reached the maximum level at 3 h. Electromobility shift and supershift assays revealed that IL-1β markedly increased DNA-protein complex, which was mainly composed of C/EBPβ and/or -δ. Both Western blotting and immunohistochemistry demonstrated that either C/EBPβ or -δ expression was induced by IL-1β exclusively in nuclei of VSMC. On the other hand, overexpression of C/EBPδ specifically transactivated the promoter activity of the PDGFαR gene and significantly enhanced VSMC proliferation in PDGF-treated cells. We conclude that induction of PDGFαR expression is mainly mediated by C/EBPδ expression in VSMC, and a high level of C/EBPδ expression may be involved in the pathogenesis of atherosclerosis and restenosis. vascular smooth muscle cell(s) platelet-derived growth factor interleukin platelet-derived growth factor α-receptor platelet-derived growth factor β-receptor CCAAT/enhancer-binding protein cycloheximide Excessive or uncontrolled replication and migration of vascular smooth muscle cells (VSMC)1are critical events involved in a number of vascular diseases including atherosclerosis, hypertension, and restenosis that often occurs after balloon angioplasty (1Mulvany M.J. Hansen P.K. Aalkjaer C. Circ. Res. 1978; 43: 854-864Crossref PubMed Scopus (434) Google Scholar, 2Spaet T.H. Stemerman M.B. Veith F.J. Lejnieks I. Circ. Res. 1975; 36: 58-70Crossref PubMed Scopus (115) Google Scholar, 3Ross R. Nature. 1993; 362: 801-809Crossref PubMed Scopus (10004) Google Scholar). Morphologic studies of the sequencing events in the arterial wall of animals with artificially induced hypercholesterolemia showed that macrophages are present in all processes of the formation of atherosclerotic lesions (4Jonasson L. Holm J. Skalli O. Bondjers G. Hansson G.K. Arteriosclerosis. 1986; 6: 131-138Crossref PubMed Google Scholar, 5Fowler S. Berberian P.A. Shio H. Goldfischer S. Wolinsky H. Circ. Res. 1980; 46: 520-530Crossref PubMed Google Scholar, 6Gown A.M. Tsukada T. Ross R. Am. J. Pathol. 1986; 125: 191-207PubMed Google Scholar, 7Joris I. Zand T. Nunnari J.J. Krolikowski F.J. Majno G. Am. J. Pathol. 1983; 113: 341-358PubMed Google Scholar). The normal function of the macrophage is to act not only as an antigen-presenting cell to T lymphocytes but also as a source of several growth factors such as platelet-derived growth factor (PDGF), basic fibroblast growth factor, tumor necrosis factor α, and transforming growth factor β1, which are generally not expressed in the normal artery, whereas they are up-regulated in the lesions of atherosclerosis (3Ross R. Nature. 1993; 362: 801-809Crossref PubMed Scopus (10004) Google Scholar). Thus, the macrophage is thought to be a principal inflammatory mediator of cells in the atheromatous plaque microenvironment. Interleukin (IL)-1β is one of the major secretory products of activated macrophage and can induce proliferation of cultured fibroblasts and VSMC (8Schmidt J.A. Mizel S.B. Cohen D. Green I. J. Immunol. 1982; 128: 2177-2182PubMed Google Scholar, 9Bitterman P.B. Wewers M.D. Rennard S.I. Adelberg S. Crystal R.G. J. Clin. Invest. 1986; 77: 700-708Crossref PubMed Scopus (175) Google Scholar, 10Libby P. Wyler D.J. Janicka M.W. Dinarello C.A. Arteriosclerosis. 1985; 5: 186-191Crossref PubMed Google Scholar, 11Libby P. Warner S.J.C. Friedman G.B. J. Clin. Invest. 1988; 81: 487-498Crossref PubMed Scopus (417) Google Scholar). Previous studies (12Raines E.W. Dower S.K. Ross R. Science. 1989; 243: 393-396Crossref PubMed Scopus (518) Google Scholar, 13Ikeda U. Ikeda M. Oohara T. Kano S. Yaginuma T. Atherosclerosis. 1990; 84: 183-188Abstract Full Text PDF PubMed Scopus (65) Google Scholar, 14Bonin P.D. Fici G.J. Singh J.P. Exp. Cell. Res. 1989; 181: 475-482Crossref PubMed Scopus (54) Google Scholar) have demonstrated that mitogenic activity of IL-1β for fibroblasts and VSMC is mediated indirectly via an autocrine loop by causing the release of PDGF-AA, which then specifically binds to the PDGF α-receptor (PDGFαR) subtype on cell surface. Furthermore, recent studies (15Bonner J.C. Lindroos P.M. Rice A.B. Moomaw C.R. Morgan D.L. Am. J. Physiol. 1998; 274: L72-L80Crossref PubMed Google Scholar, 16Lindroos P.M. Coin P.G. Badgett A. Morgan D.L. Bonner J.C. Am. J. Respir. Cell. Mol. Biol. 1998; 16: 283-292Crossref Scopus (59) Google Scholar) have also demonstrated that IL-1β can up-regulate PDGFαR expression in rat lung fibroblasts, thereby enhancing PDGF-mediated mitogenesis and chemotaxis of lung fibroblasts. Although the pathophysiological implications of IL-1β-induced PDGFαR expression are beginning to be recognized, little is known about the molecular mechanism involved. Therefore, we have investigated the molecular mechanism of PDGFαR gene transcription in VSMC and obtained results indicating that IL-1β induces PDGFαR gene expression via a trans-acting nuclear factor, CCAAT/enhancer-binding protein δ (C/EBPδ). Actinomycin D and cycloheximide (CHX) were purchased from Sigma, and recombinant mouse IL-1β was from Roche Molecular Biochemicals (Tokyo, Japan). [α-32P]dCTP (110 TBq/mmol) and [γ-32P]ATP (220 TBq/mmol) were obtained from Amersham Pharmacia Biotech (Tokyo, Japan). Affinity-purified rabbit polyclonal antibodies for PDGFαR and C/EBPα, -β, and -δ raised against peptidic epitopes corresponding to amino acid residues of human PDGFαR (residues 951–1,089), rat C/EBPα (residues 253–265), rat C/EBPβ (residues 258–276), and rat C/EBPδ (residues 247–268) were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA). Expression vectors of C/EBPα, -β, and -δ (designated EBPα, -β, and -δ, respectively) were generous gifts of Dr. Steven L. McKnight. VSMC were isolated from the thoracic aorta of male Harlan Sprague-Dawley rats (Charles River Japan Inc., Kanagawa, Japan), weighing 280–320 g, by the method described previously (17Inui H. Kitami Y. Kondo T. Inagami T. J. Biol. Chem. 1993; 268: 17045-17050Abstract Full Text PDF PubMed Google Scholar). Cells were seeded onto 100-mm dishes at a density of 1 × 106 per dish and maintained in Dulbecco's modified Eagle's medium with 10% heat-inactivated fetal calf serum at 37 °C in a humidified atmosphere of 95% air, 5% CO2. VSMC were passaged every 4–7 days, and experiments were performed on cells at 3–10 passages from primary culture. In preparation for experiments, confluent cells, which exhibited a typical hill and valley pattern of smooth muscle cells in culture, were made quiescent by placing them in a defined serum-free medium containing insulin (10 μg/ml), transferrin (10 μg/ml), and sodium selenite (10 ng/ml) for 48 h. This medium has been shown to maintain VSMC in a quiescent and noncatabolic state for an extended period of time (18Libby P. O'Brien K.V. J. Cell. Physiol. 1983; 115: 217-223Crossref PubMed Scopus (184) Google Scholar). A 0.6-kilobase pair fragment of rat PDGFαR cDNA (19Kitami Y. Inui H. Uno S. Inagami T. J. Clin. Invest. 1995; 96: 558-567Crossref PubMed Scopus (45) Google Scholar), 1.1-kilobase pair Nco I fragment of EBPα, 0.4-kilobase pair Nco I fragment of EBPβ or 1.0-kilobase pair Eco RI–Bam HI fragment of EBPδ was used as a probe for Northern blotting. Each DNA fragment was labeled with [α-32P]dCTP using the random primer method. Total cellular RNA extraction from VSMC and Northern blot analysis were carried out by the methods described previously (19Kitami Y. Inui H. Uno S. Inagami T. J. Clin. Invest. 1995; 96: 558-567Crossref PubMed Scopus (45) Google Scholar, 20Kitami Y. Hiwada K. Kokubu T. J. Hypertens. 1989; 7: 727-731Crossref PubMed Scopus (20) Google Scholar). Nuclear extracts were prepared from VSMC according to the method described by Dignam et al. (21Dignam J.D. Lebovitz R.M. Roeder R.G. Nucleic Acids Res. 1983; 11: 1475-1489Crossref PubMed Scopus (9164) Google Scholar). After protein concentrations were determined using Bio-Rad Protein Assay Reagent, nuclear extracts were divided into small aliquots, quickly frozen in liquid nitrogen, and stored at −80 °C. For electrophoretic mobility shift assay and supershift assay, a double-stranded oligodeoxynucleotide probe for the consensus sequence of C/EBP was generated by annealing two complementary oligodeoxynucleotides corresponding to the nucleotide sequence spanning −165 to −138 in the 5′-flanking region of the rat PDGFαR gene, 5′-CCCCAGATTGCATAAGAGCAAAAAGCCA-3′. Another double-stranded oligodeoxynucleotide probe for the consensus sequence of nuclear factor-1, 5′-CCTTTGGCATGCTGCCAATAT G-3′, was purchased from Promega (Madison, WI) and used as an unrelated competitor. The C/EBP probe was end-labeled with [γ-32P]ATP using T4-polynucleotide kinase. Nuclear extracts (2 μg) were incubated with 2.0 × 104 cpm of the labeled C/EBP probe for 30 min at room temperature in a 10-μl binding buffer containing 12 mm Hepes-KOH, pH 7.9, 60 mm KCl, 4 mm MgCl2, 1 mm EDTA, 1 mm dithiothreitol, 10% glycerol, and 50 μg/ml of poly(dI-dC)(dI-dC) (Amersham Pharmacia Biotech). For competition experiments or supershift assay, a 100-fold molar excess of unlabeled probe or 1–2 μl of antibodies against each subtype of the C/EBP was to nuclear and was incubated for 30 min at room temperature of the labeled C/EBP all were by 5% and the was and as described previously (19Kitami Y. Inui H. Uno S. Inagami T. J. Clin. Invest. 1995; 96: 558-567Crossref PubMed Scopus (45) Google Scholar). PDGFαR which was as was prepared by of the promoter region spanning of the PDGFαR gene (19Kitami Y. Inui H. Uno S. Inagami T. J. Clin. Invest. 1995; 96: 558-567Crossref PubMed Scopus (45) Google Scholar) onto which was as was prepared by of the region of C/EBPδ cDNA from which can was used as an control to VSMC were seeded onto dishes × or × 104 for or cell proliferation assay, DNA was performed with cells at according to the of Inc., Japan). For assay, or was used for with EBPα, -β, -δ, or in to For cell proliferation assay, EBPδ or was used for of VSMC. activity was determined by the Assay as described previously Y. T. T. Y. M. M. K. Hiwada K. J. Hypertens. 1998; 16: PubMed Scopus Google Scholar, Y. Kitami Y. T. T. Hiwada K. PubMed Google Scholar). After for in to determined each promoter activity was as a activity in to the activity of with that was to proliferation Molecular was used for VSMC proliferation assay according to the after PDGF-AA or was to the and cells were incubated for an of was to each and cells were incubated for 30 min at 37 °C. the of each was by an assay and cell proliferation activity was as a activity in to the activity of VSMC after treatment with PDGF-AA that was to Western blotting was performed by the method described previously (17Inui H. Kitami Y. Kondo T. Inagami T. J. Biol. Chem. 1993; 268: 17045-17050Abstract Full Text PDF PubMed Google Scholar, Y. Inui H. Uno S. Inagami T. J. Clin. Invest. 1995; 96: 558-567Crossref PubMed Scopus (45) Google Scholar). nuclear extracts μg) prepared from VSMC were to Western blotting for C/EBPα, -β, and -δ. After with was performed using a according to Nature. PubMed Scopus Google Scholar), and in the were to a by for 1 at The was with primary antibodies against each C/EBP and were by using a Western blotting analysis Amersham Pharmacia Biotech). VSMC were cultured for in of at a density of 1 × 104 and then with for 30 were with containing for min at 37 and the were performed according to the for the Inc., CA). After for 30 min at 37 °C with normal were with primary antibodies for 1 at room After to a containing the were with a cells were with a of The were with with and with and then in medium Japan). of with analysis was used to two are expressed as and is defined as VSMC from Harlan Sprague-Dawley rats were incubated for in the of IL-1β at concentrations and mRNA levels of both PDGFαR and C/EBPδ were determined by Northern blotting. Although levels of both mRNA expression were very or in quiescent VSMC, they were markedly increased by the treatment with IL-1β at to in a not Therefore, we used the of IL-1β at To the C/EBP and PDGFαR expression treatment with mRNA levels of C/EBP and PDGFαR were for 48 h. shown in A and a high level of PDGFαR mRNA expression was by a induction of C/EBPδ mRNA in VSMC the of IL-1β. A rapid induction of C/EBPδ mRNA within 30 min was followed by slower emergence of PDGFαR mRNA, which reached the maximum level the time in 12 h, whereas C/EBPδ mRNA reached the maximum level the time at 3 h, at for 12 h, and then to a level within 48 h. The induction of PDGFαR mRNA expression to at 3 and at for h. This time a in which C/EBPδ induced PDGFαR gene In a high level of C/EBPα mRNA expression was in a quiescent and IL-1β not significantly Although the induction of C/EBPβ mRNA expression was also detectable at 30 for an extended period to 48 h. To the induction of PDGFαR mRNA expression after treatment with IL-1β is to the of IL-1β on the mRNA the time of PDGFαR mRNA was determined in the of D VSMC were with IL-1β for 12 h, with and then to a prepared medium with or IL-1β in the of D. A time of the PDGFαR mRNA in cells incubated in the medium with IL-1β was h, and that IL-1β was h, indicating that IL-1β not significantly the PDGFαR mRNA mobility shift assay was performed by using a labeled C/EBP probe containing the consensus sequence of C/EBP of rat PDGFαR promoter region Nuclear extracts were prepared from either quiescent or VSMC. Although the C/EBP probe was not by nuclear extracts from quiescent VSMC, was by nuclear extracts from VSMC with IL-1β for 12 h, a of the DNA-protein The DNA-protein was markedly out by a 100-fold molar excess of unlabeled C/EBP probe but not by a 100-fold molar excess of unlabeled nuclear To the specific subtype of C/EBP that is by the probe and involved in the transcriptional of the PDGFαR gene, supershift assay was performed using antibodies against major of C/EBP C/EBPα, -β, and -δ 3 In VSMC, the was by antibodies against either C/EBPβ or -δ but not To the for C/EBP in the transcription of the rat PDGFαR gene, we the of the C/EBP to the promoter of PDGFαR in a The PDGFαR was with a or an expression for each C/EBP EBPα, -β, or -δ. expression of C/EBPδ specifically transactivated the promoter activity of the of on the of with that of with On the other hand, expression of other C/EBP not significantly PDGFαR gene promoter Furthermore, cell proliferation activity treatment with PDGF-AA or was determined in the VSMC with or EBPδ 4 activity treatment with PDGF-AA or was significantly enhanced in the cells with EBPδ with with the of on the of or To C/EBPδ gene expression is activated by IL-1β other protein the of IL-1β to induce C/EBPδ gene expression was determined in the of (10 Although induction of PDGFαR mRNA expression by IL-1β was markedly in the of that of C/EBPδ mRNA expression was in the of On the other hand, not the of C/EBPδ mRNA In protein levels of PDGFαR and C/EBPα, -β, and -δ expression were by immunohistochemistry in quiescent or VSMC. Although protein levels of PDGFαR were very or in quiescent VSMC, IL-1β drastically induced PDGFαR expression exclusively in the of cells. A high level of C/EBPα protein expression was in both quiescent and VSMC and was mainly in the of C/EBPδ protein was not in quiescent VSMC, whereas that of C/EBPβ protein was specifically in the of the Furthermore, either C/EBPβ or -δ protein expression was markedly induced by the treatment with IL-1β and was exclusively and in the To a of C/EBP as nuclear nuclear extracts from quiescent or VSMC were to Western blotting using specific antibodies against C/EBPα, -β, and -δ Although nuclear extracts from quiescent VSMC only a level of C/EBPβ protein but not C/EBPα or -δ either C/EBPβ or C/EBPδ protein was markedly induced in the nuclear extracts from VSMC for 12 h. The expression level of C/EBPα protein was in the nuclear extracts from either quiescent or VSMC. C/EBPδ has been in the as one of the of C/EBP that to the basic transcriptional factors C.A. 5: PubMed Scopus Google Scholar, R.M. 5: PubMed Scopus Google Scholar, R. Y. S. J. Biol. Chem. Full Text PDF PubMed Google Scholar). Previous studies revealed that C/EBPδ expression was at an or level in normal cells or and was induced by and inflammatory cytokines such as and tumor necrosis factor M.D. G.J. U. S. A. 1993; PubMed Scopus Google Scholar, S. S. T. U. S. A. PubMed Scopus Google Scholar, T. J. J. Biol. Chem. Full Text PDF PubMed Google Scholar). Therefore, C/EBPδ is thought to be an factor to the gene transcription of M.D. G.J. U. S. A. 1993; PubMed Scopus Google Scholar, H. J. Biol. Chem. Full Text PDF PubMed Google Scholar). Both and its specific receptor, are also known to be up-regulated by the treatment with IL-1β in several cells including VSMC and fibroblasts, causing the cell migration and/or proliferation in the (12Raines E.W. Dower S.K. Ross R. Science. 1989; 243: 393-396Crossref PubMed Scopus (518) Google Scholar, J.C. Lindroos P.M. Rice A.B. Moomaw C.R. Morgan D.L. Am. J. Physiol. 1998; 274: L72-L80Crossref PubMed Google Scholar, 16Lindroos P.M. Coin P.G. Badgett A. Morgan D.L. Bonner J.C. Am. J. Respir. Cell. Mol. Biol. 1998; 16: 283-292Crossref Scopus (59) Google Scholar). little has been known about molecular of its gene et al. T. Science. PubMed Scopus Google Scholar) have demonstrated that major vascular such as transforming growth factor β1, and factor are transactivated by the of two specific regulatory nuclear and that a mechanism may in the transcriptional of we have demonstrated that the PDGF β-receptor gene is mainly by the at of its promoter region in VSMC Y. T. T. Y. M. M. K. Hiwada K. J. Hypertens. 1998; 16: PubMed Scopus Google Scholar, Y. Kitami Y. T. T. Hiwada K. PubMed Google Scholar), that a mechanism via the also in the transcriptional of vascular such as PDGFαR and In the present we have demonstrated that an in the level of PDGFαR mRNA in VSMC treatment with IL-1β is mainly to the transcriptional of the gene but not to of mRNA Northern blotting revealed that C/EBPδ mRNA expression was drastically induced in the VSMC in with results obtained from the supershift assay 3 immunohistochemistry and Western blotting shown in mechanism is by a rapid C/EBPδ induction by IL-1β on within 30 min and at 3 that was followed by a slower emergence at 12 of PDGFαR mRNA, indicating that C/EBPδ expression is to the of the PDGFαR gene in VSMC. Furthermore, we have determined the of IL-1β to induce C/EBPδ gene expression in the of to C/EBPδ gene expression is activated by IL-1β other protein Although either or with IL-1β not induce PDGFαR mRNA, with IL-1β a C/EBPδ mRNA This that C/EBPδ the endogenous PDGFαR gene expression in VSMC of other we have that a C/EBP binding in PDGFαR gene promoter region as a major regulatory for its expression in a Y. T. Hiwada K. Inagami T. Circ. Res. 84: PubMed Scopus Google Scholar). et al. C.A. Am. J. Physiol. 1995; PubMed Google Scholar) have that the C/EBP is involved in the transcriptional of the gene in et al. S. Mol. Cell. Biol. 1995; PubMed Google Scholar) have that the gene, which to the with PDGFαR and also has a C/EBP binding in the promoter and either or C/EBPα mainly the cell gene expression in cells. the that the C/EBP especially is a major of PDGFαR gene transcription in VSMC. assay and Western blotting that IL-1β markedly induced specific which are as C/EBP and C/EBP with the C/EBP binding of PDGFαR promoter are C/EBPβ and -δ 3 and Although C/EBPβ was induced by was also in quiescent VSMC and On the other hand, C/EBPδ was exclusively in the nuclei after treatment with IL-1β and was of with the C/EBP binding of the PDGFαR Furthermore, overexpression studies have demonstrated that C/EBPδ but not C/EBPα or specifically transactivated PDGFαR promoter activity in VSMC 4 and that cell proliferation activity treatment with PDGF-AA or was significantly enhanced in the VSMC with EBPδ with with can to not only but also enhanced on the cell proliferation treatment with is mediated by the action the PDGFαR not up-regulated by C/EBPδ of cell proliferation was significantly in the VSMC after treatment with with with PDGF-AA. This was in with (17Inui H. Kitami Y. Kondo T. Inagami T. J. Biol. Chem. 1993; 268: 17045-17050Abstract Full Text PDF PubMed Google Scholar) in which we the mitogenic activity after treatment with PDGF-AA or by of and that was significantly in the cells after treatment with with with PDGF-AA. we have isolated and the promoter region of the rat C/EBPδ gene to understand the regulatory mechanism of C/EBPδ gene transcription by IL-1β in VSMC T. Kitami Y. K. Hiwada K. Res. PubMed Scopus Google Scholar). A with to the molecular mechanism of rat C/EBPδ gene transcription in human cell has demonstrated that the C/EBPδ gene is activated by the regulatory which is by response and of transcription 3 T. K. S. T. M. J. PubMed Scopus Google Scholar). of response and of transcription 3 by increased its DNA binding activity and an induction of C/EBPδ gene that a mechanism may on the of the C/EBPδ gene by IL-1β and to the that of other is not for its In the present is at the molecular mechanism in the gene expression of PDGFαR in VSMC. The results obtained a for of the C/EBP especially on vascular growth and and also to understand the mechanism pathogenesis of vascular and atherosclerosis or restenosis. We are to Dr. Steven L. of of for the generous of EBPα, -β, and -δ We are also to Dr. Inagami of of for critical of the