Published online before print June 3, 2004, doi: 10.1161/01.RES.0000134631.75684.4A
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© 2004 American Heart Association, Inc.
Molecular Medicine |
Myocyte Enhancer Factor 2 Mediates Vascular Inflammation via the p38-Dependent Pathway
From the Departments of Internal Medicine (E.S., H.S., S.O., R.T., M.O., T.F., R.N., Y.H.) and Urology (H.N.), Faculty of Medicine, University of Tokyo, Tokyo, Japan.
Correspondence to Etsu Suzuki, MD, PhD, Division of Nephrology and Endocrinology #202, The Department of Internal Medicine, Faculty of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan. E-mail suzuki-2im{at}h.u-tokyo.ac.jp
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Abstract |
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Although it has been established that myocyte enhancer factor 2 (MEF2) plays pivotal roles in the development of the cardiovascular system as well as skeletal muscle cells, little is known of its role in vascular inflammatory diseases such as atherosclerosis and restenosis after angioplasty. To investigate the role of MEF2 in vascular inflammation and that of p38 in the activation of MEF2, we infected cultured rat vascular smooth muscle cells (VSMCs) with an adenovirus construct expressing a dominant-negative mutant of MEF2A (MEF2ASA) or mitogen-activated protein kinase kinase 6 (MEK6AA), and examined their effects on the expression of monocyte chemoattractant protein-1 (MCP-1), which is known to play important roles in vascular inflammation. We also examined the role of MEF2 in vivo using a rat model of transluminal wire-induced injury of the femoral artery. Angiotensin II (Ang II)–induced expression of MCP-1 mRNA was significantly inhibited by infection with adenoviruses encoding MEF2ASA (AdMEF2ASA) or MEK6AA. Ang II–induced increase of MCP-1 promoter activity was also significantly suppressed by overexpression of MEF2ASA or MEK6AA. Ang II stimulated the transactivating function of MEF2A and this activation was inhibited by overexpression of MEK6AA. Infection with AdMEF2ASA suppressed MCP-1 expression in the femoral artery after the transluminal mechanical injury. AdMEF2ASA infection also inhibited macrophages infiltration and neointimal formation in the wire-injured femoral arteries. These results suggested that MEF2 activation via the p38-dependent pathway mediates vascular inflammation via stimulation of MCP-1 expression in VSMCs and macrophages infiltration.
Key Words: atherosclerosis • angioplasty • angiotensin • signal transduction • inflammation
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Introduction |
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It has recently become clear that inflammation mediated by chemokines plays pivotal roles in the initiation and maintenance of vascular diseases such as atherosclerosis and restenosis after angioplasty.1 Among a variety of chemokines, monocyte chemoattractant protein-1 (MCP-1) is reportedly critically implicated in the development of vascular diseases. MCP-1 is widely expressed in atherosclerotic lesions including vascular endothelial cells, smooth muscle cells, and macrophages.2 It has been reported that the additional disruption of the MCP-1 receptor markedly attenuated atherosclerotic lesions by inhibiting macrophage infiltration in apolipoprotein E null mice.3 It has also been shown that neutralization of MCP-1 is effective in preventing restenosis after angioplasty.4 Intracellular signaling pathways leading to the activation of the MCP-1 gene have been intensively studied. Protein kinases including extracellular signal-regulated kinase (ERK), p38, and janus kinase (JAK) appear to be involved in the induction of MCP-1 expression.5–8 A variety of nuclear factors such as nuclear factor-
B (NF-B), SP-1, AP-1, and signal transducers and activators of transcription (STAT) have been shown to be implicated in the transcriptional activation of the MCP-1 gene.9–12 However, the mechanisms by which those protein kinases activate the nuclear factors remain to be elucidated. Accumulated evidence suggests that the renin-angiotensin system is implicated in the pathogenesis of atherosclerosis.13,14 It also seems to be involved in the pathogenesis of restenosis after angioplasty.15,16 It has recently been shown that angiotensin II (Ang II) promotes the expression of MCP-1 in vascular smooth muscle cells (VSMCs), although the precise mechanism has not been clarified.17
Myocyte enhancer factor 2 (MEF2) (related to serum response factor protein) is a family of transcription factors that comprise four isoforms, that is, MEF2A, -B, -C, and -D.18–21 MEF2A-deficient mice show dilation of the right ventricle, mitochondrial disorganization, and cardiac sudden death.22 MEF2C-null mice do not form a normal heart and vascular system,23,24 suggesting that MEF2 transcription factors are required for the development of the cardiovascular system. It has also been shown that some isoforms of MEF2 transcription factors are expressed predominantly in the neointima after balloon injury of the rat carotid artery.25 However, little is known about the role of MEF2 in vascular diseases. We recently reported that MEF2A was a major isoform expressed in vascular VSMCs, and that MEF2A expression, its DNA binding activity, and its transactivating function increased when serum-starved VSMCs were restimulated with serum mitogen.26 We have also shown that Ang II–induced activation of the c-jun gene is mediated by a p38-dependent pathway as well as an MEF2-dependent pathway.27 Furthermore, it has been demonstrated that MEF2A and MEF2C are phosphorylated by p38.28,29 These results indicate that MEF2 transcription factors may mediate signals from p38 in VSMCs. Because p38 is a well-known molecule that is involved in the expression of the MCP-1 gene, we speculated that MEF2 transcription factors might be implicated in Ang II–induced expression of MCP-1.
In the present study, we examined whether Ang II–induced expression of MCP-1 would be mediated by MEF2 and p38. We also examined the mechanisms by which MEF2 was activated by p38. Finally, we investigated the role of MEF2 in vivo using a rat model of wire-induced injury of the femoral artery.
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Materials and Methods |
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Reagents
Valsartan (Val) was kindly supplied by Novartis Pharma AG. Phosphospecific anti-p38 antibody, which recognizes catalytically active p38, was obtained from Promega. Anti-MEF2A antibody and anti-p38 antibody were obtained from Santa Cruz Biotechnology. Curcumin and Bay 11-7082 were purchased from Sigma-Aldrich Co. Human tumor necrosis factor-
(TNF-) was obtained from Wako Pure Chemical Industries.
Cell Culture
Rat VSMCs were cultured from rat thoracic aortas following the explant method, as previously described.30
RNA Extraction and Real-Time PCR
RNA extraction and real-time PCR analysis were performed as previously described.31 To confirm that no significant amounts of primer dimers were formed, dissociation curves were analyzed. PCR-amplified products were also electrophoresed on 2% agarose gels to confirm single bands were amplified.
Plasmids
Isolation of the promoter region of human MCP-1 gene (2644 bp, pGL2/–2644humanMCP-1promoter) was performed by PCR as previously described.31 A single putative MEF2 binding site (TTAAAAATAA) that is located at –287 to –278 upstream from the transcription start site was mutated by PCR (pGL2/–2644humanMCP-1promoter/MEF2mutant). The primer used to introduce the mutation was as follows: 5'-CTTCACAGAAAGCAGAATCCTTggcAATAACCCTCTTAGTTCACATC-3'. The lowercase letters in bold type indicate nucleotides substitutions to induce the mutation. The DNA sequence was determined by cycle sequence reaction using a CEQ8000 DNA sequencer (Beckman Coulter). Details of the cloning of human MEF2A wild-type (MEF2Awt) and MEF2AS453A (MEF2ASA) in which serine 453 was substituted with alanine were as previously described.27 Construction of human MEF2A/R24LS453A (MEF2A24L/SA) and MEF2A/K31LS453A (MEF2A31L/SA), in which leucine was substituted for arginine 24 or lysine 31 in addition to the alanine substitution for serine 453, was described elsewhere (see the expanded Materials and Methods section in the online data supplement available at http://circres.ahajournals.org). To make fusion proteins of the GAL4 DNA binding domain and MEF2A transactivation domain, the transactivation domain of MEF2A (from codon 87 up to stop codon) was PCR amplified using MEF2Awt and MEF2ASA as templates. These fragments were digested with XbaI and KpnI, and ligated in the pBIND vector (Promega) at XbaI and KpnI sites, which are located downstream of the GAL4 DNA binding domain (pBINDMEF2Awt and pBINDMEF2ASA). Construction of MEK6S207A/T211A (MEK6AA) and MEK6S207E/T211E (MEK6EE) in which serine 207 and threonine 211 were replaced with alanine and glutamic acid, respectively, was as previously described.27
Transient Transfection
pRL-TK, which encodes the SeaPansy luciferase gene, was purchased from Toyo Ink and used as the internal control for the luciferase assays. To examine the activity of MCP-1 promoter, rat VSMCs were transiently transfected with reporter plasmids including pGL2/–2644humanMCP-1promoter or pGL2/–2644humanMCP-1promoter/MEF2mutant, along with pRL-TK using lipofectAMINE (Life Technologies). Rat VSMCs were also cotransfected with expression plasmids encoding MEF2ASA (pcDNA3HA/MEF2ASA), MEF2A24L/SA (pcDNA3HA/MEF2A24L/SA), MEF2A31L/SA (pcDNA3HA/MEF2A31L/SA), or MEK6AA (pcDNA3HA/MEK6AA) in some experiments to examine the effects of these mutants on the activities of those promoters. Some cells were stimulated with 10–7 mol/L Ang II in the presence and absence of pretreatment with 100 nmol/L Val. To examine the transactivating function of MEF2, rat VSMCs were transiently transfected with pG5luc vector (Promega), which contains five consecutive GAL4 binding sites upstream of the luciferase gene, and pBINDMEF2Awt or pBINDMEF2ASA, along with pRL-TK using lipofectAMINE. Rat VSMCs were also cotransfected with expression plasmids encoding MEK6AA or MEK6EE (pcDNA3HA/MEK6EE) in some experiments. Some cells were stimulated with 10–7 mol/L Ang II in the presence and absence of pretreatment with 100 nmol/L Val. The total amounts of plasmid DNA transfected in VSMCs were adjusted using the expression vector pcDNA3. Dual luciferase assay was performed using a luminometer (Lumat LB 9507, Berthold). SeaPansy luciferase activity was used as the internal control to normalize the promoter activity.
Construction of a Replication-Defective Adenovirus
Replication-defective adenoviruses that express MEK6AA (AdMEK6AA) or MEK6EE (AdMEK6EE) were constructed according to the method previously described using an AdMax kit (Microbix Biosystems Inc).27 Construction of a recombinant adenovirus that expresses MEF2ASA (AdMEF2ASA) was previously described.27 A recombinant adenovirus expressing green fluorescence protein (AdGFP) was obtained from Quantum Biotechnologies.
Western Blot Analysis
Protein extraction and Western blot analyses were performed as previously described.32 Antibodies were used at a dilution of 1:200 except for anti-phospho p38 antibody, which was used at a dilution of 1:500.
Rat Femoral Artery Injury and Gene Transfer
Male Wistar rats (8 to 10 weeks old) were obtained from Charles River (Wilmington, Mass.). Transluminal mechanical injury of the rat femoral artery was induced as previously described.31 After the mechanical injury, a 27-gauge needle was inserted in a muscular branch of the femoral artery proximally to it and clamped together with the artery. The femoral artery was also clamped with a plastic forceps at the inguinal portion. Adenoviruses (4x108 plaque forming units) or saline was injected and the femoral artery was incubated with the virus suspension for 30 minutes. The femoral artery was harvested 3 days after the injury to extract RNA, or 14 days after the injury for histochemical analysis.
Histochemical Analysis
The histochemical analysis of the femoral arteries was performed as previously described.31
Statistical Analysis
The values are the mean±SEM. Statistical analyses were performed using analysis of variance followed by the Student-Neumann-Keul test. Differences with a value of P<0.05 were considered statistically significant.
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Ang II Promotes MCP-1 Expression Through the MEF2 and p38-Mediated Pathways in Vascular Myocytes
To examine the role of MEF2 and p38 in the expression of MCP-1, cultured rat VSMCs were stimulated with Ang II and the effects of dominant-negative mutants of MEF2A and MEK6 were examined using real-time PCR. Ang II (10–7 mol/L) significantly stimulated the expression of MCP-1 mRNA 4 hours after stimulation (Figure 1A). This increase was significantly suppressed by pretreatment with 100 nmol/L valsartan (Val), a type I receptor antagonist for Ang II, and by infection with adenoviruses expressing a dominant-negative mutant of MEF2A (AdMEF2ASA) or MEK6 (AdMEK6AA). Infection with an adenovirus expressing a constitutively active mutant of MEK6 (AdMEK6EE) significantly stimulated MCP-1 expression in the absence of Ang II. We also analyzed PCR products by ethidium bromide staining and confirmed that single bands were amplified and that the amounts of the amplified products showed the same tendency as the results of real-time PCR (Figure 1B). To examine the role of the AP-1– and NF-
B–dependent pathways in Ang II stimulation of MCP-1 expression, we used several pharmacological inhibitors of those pathways. When rat VSMCs were pretreated for 1 hour with the AP-1 inhibitor curcumin (10 µmol/L) or the NF-B inhibitor Bay 11–7082 (5 µmol/L), these inhibitors did not significantly suppress Ang II induction of MCP-1 expression (Figure 2A). In marked contrast, these inhibitors significantly suppressed TNF- induction of MCP-1 expression (Figure 2B). These results suggested that Ang II stimulated MCP-1 expression via the p38- and MEF2-dependent pathways, but not via the AP-1– or NF-B–dependent pathway in VSMCs.
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We next examined whether Ang II–induced increase of MCP-1 expression was mediated at the level of transcription. We used the human MCP-1 promoter (pGL2/–2644humanMCP-1promoter). Ang II (10–7 mol/L) significantly enhanced this promoter activity, and pretreatment with Val or cotransfection with expression plasmids encoding MEF2ASA or MEK6AA significantly suppressed Ang II–induced increase of the promoter activity (Figure 3A). These results suggested that Ang II–induced MCP-1 expression and the effects of MEF2ASA and MEK6AA on MCP-1 expression were, at least partly, mediated at the level of transcription.
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The promoter region of the MCP-1 gene we used in this study contains a single putative MEF2 binding site. We, therefore, mutated the MEF2 binding site (pGL2/–2644humanMCP-1promoter/MEF2mutant) and examined whether Ang II–induced increase of the promoter activity depended on the MEF2 binding site (Figure 3B). Surprisingly, Ang II significantly stimulated the activity of the MEF2 binding site-mutated promoter to the same extent as that of the wild-type promoter. Furthermore, cotransfection with the expression plasmid encoding MEF2ASA significantly suppressed Ang II–induced activation of the MEF2 binding site-mutated promoter to the same level as that observed in the wild-type promoter. These results suggested that although MEF2 was implicated in Ang II–induced transcriptional activation of the MCP-1 gene, the MEF2 binding site in the MCP-1 promoter region was not required for the MEF2-dependent expression of the MCP-1 gene. To further examine this mechanism, we mutated the DNA binding domain of MEF2ASA to construct mutants that do not bind consensus MEF2 binding sites and also lack an active transactivation domain. It has been reported that when leucine was substituted for arginine 24 or lysine 31 of human MEF2C it resulted in mutants that did not bind MEF2 binding sites.33 Because the DNA binding domain of MEF2 isoforms is highly conserved, we inserted the same mutation in the DNA binding domain of MEF2ASA (MEF2A24L/SA and MEF2A31L/SA) and subcloned these mutants in an expression vector (pcDNA3HA/MEF2A24L/SA and pcDNA3HA/MEF2A31L/SA). The expression of these mutants was confirmed by Western blot analysis (See online Figure 1 in the online data supplement). Incapability of these mutants to bind to MEF2 binding sites was confirmed by electrophoretic mobility shift assay (data not shown). When these mutants were cotransfected into VSMCs with the MCP-1 promoter, Ang II induction of MCP-1 promoter activity was significantly suppressed by these mutants as well as by expression plasmid encoding MEF2ASA (Figure 3C). These results suggested that MEF2 potentially transactivated MCP-1 gene without binding to the MEF2 binding site.
Ang II Promotes the Transactivating Function of MEF2 via the p38-Dependent Pathway in Vascular Myocytes
It has been reported that Ang II activates the p38-dependent pathway in VSMCs.34 To confirm the activation of p38 by Ang II and that our MEK6AA and MEK6EE mutants functioned as dominant-negative and constitutively active mutants, respectively, in our system, we stimulated cultured rat VSMCs with Ang II and examined the phosphorylation of p38. Ang II (10–7 mol/L) stimulated p38 phosphorylation, which peaked 15 minutes after stimulation with Ang II (Figure 4A). We, therefore, stimulated rat VSMCs with Ang II for 15 minutes and examined the effects of the MEK6 mutants. Ang II–induced phosphorylation of p38 was remarkably inhibited by pretreatment with Val and infection with AdMEK6AA. Infection with AdMEK6EE increased p38 phosphorylation in the absence of Ang II (Figure 4B), suggesting that MEK6AA and MEK6EE functioned as dominant-negative and constitutively active mutants, respectively, for p38 activation in our system.
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We next examined whether the transactivating function of MEF2 was mediated by the p38-dependent pathway in VSMCs. We used fusion proteins of the GAL4 DNA binding domain and transactivation domain of human MEF2Awt (pBINDMEF2Awt) or MEF2ASA (pBINDMEF2ASA). Cultured rat VSMCs were transfected with these plasmids and pG5luc, which contains five consecutive GAL4 DNA binding sites upstream of the luciferase gene. Therefore, activation of the luciferase reporter gene depended on the DNA binding of pBINDMEF2Awt or pBINDMEF2ASA via the GAL4 DNA binding domain and the transactivating function of the transactivation domain of MEF2A (Figure 5A). When pBINDMEF2Awt was used, Ang II significantly stimulated the activity of the luciferase gene, and this increase was significantly inhibited by pretreatment with Val or cotransfection with an expression plasmid encoding MEK6AA (Figure 5B). Cotransfection with an expression plasmid encoding MEK6EE significantly stimulated luciferase activity in the absence of Ang II. In marked contrast, when pBINDMEF2ASA was used, neither Ang II nor cotransfection with the expression plasmid encoding MEK6EE significantly enhanced luciferase activity. These results indicated that Ang II stimulated the transactivating function of MEF2A via the p38-dependent pathway and that serine 453 of MEF2A was critically implicated in the p38-dependent stimulation of the transactivating function of MEF2A in VSMCs.
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MEF2 Is Implicated in Neointimal Formation and Macrophages Infiltration in Blood Vessels
It has been reported that MCP-1 is required for macrophages infiltration and neointimal formation after balloon injury.4,35 To study the significance of the MEF2- and p38-dependent upregulation of MCP-1 expression in vivo, we infected adenoviruses expressing a dominant-negative mutant of MEF2A (AdMEF2ASA) or MEK6 (AdMEK6AA) and examined the effect of these mutants on macrophages infiltration and neointimal formation in the rat femoral artery after transluminal wire-induced injury. We first checked whether infection of these mutants inhibited MCP-1 expression in this transluminal wire-induced injury model. MCP-1 mRNA expression in the femoral artery was significantly enhanced three days after the injury, and this increase was significantly suppressed by infection with AdMEF2ASA or AdMEK6AA, suggesting that these mutants also inhibited MCP-1 mRNA expression in vivo (Figure 6). Neointimal formation [the ratio of intimal to medial area (I/M ratio)] was significantly inhibited by infection with AdMEF2ASA or AdMEK6AA compared with AdGFP infection (Figure 7A and 7C). Infiltration of macrophages was observed mainly in the intima. The number of macrophages in the intima was significantly attenuated by infection with AdMEF2ASA or AdMEK6AA (Figure 7B and 7D). Finally, to study whether suppression of the MEF2-dependent pathway inhibited neointimal formation via its direct effect on the proliferation of VSMCs, we examined the effect of AdMEF2ASA infection on endothelin (ET)-1–induced 3H-thymidine uptake in cultured rat VSMCs. AdMEF2ASA infection did not significantly inhibit the ET-1–induced increase of 3H-thymidine incorporation (online Figure 2).
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Discussion |
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Accumulated evidence has suggested that a variety of cis-elements in the promoter region of the MCP-1 gene such as distal NF-
B sites, a STAT binding site, AP-1 sites, SP-1 sites, and a proximal NF-B site are implicated in the transcriptional activation of the gene.9–12 It has also been reported that mitogen-activated protein kinases such as ERK and p38, and JAK are involved in the activation of the MCP-1 gene.5–8 However, the mechanisms by which these protein kinases activate those transcription factors to induce the activation of the MCP-1 gene remain to be determined. Furthermore, it also remains unclear whether other transcription factors participate in the activation of the MCP-1 gene. In this study, we showed that Ang II–induced expression of the MCP-1 gene is mediated by the p38- and MEF2-dependent pathways in VSMCs. Surprisingly, mutation in a single putative MEF2 binding site in the MCP-1 promoter region did not affect Ang II–induced increase of the promoter activity. Furthermore, forced expression of MEF2A mutants, which do not bind a consensus MEF2 binding sequence and lack transactivation capacity, effectively suppressed the activity of the MCP-1 promoter. These results suggested that MEF2 transcription factors did not need to bind directly to DNA elements located in the promoter region of the MCP-1 gene. MEF2 might transactivate the MCP-1 gene via a protein-protein interaction with some other DNA-bound transcription factors without by itself binding to DNA. It has been reported that MEF2 induces the expression of muscle-specific genes without directly binding to DNA. MEF2 seems to associate with the MyoD family of transcription factors, which are bound to DNA, via a protein-protein interaction and transactivate genes.33 Thus, it appears that MEF2 potentially transactivates genes without binding to DNA.
We also showed in this study that Ang II–induced increase of the transactivating function of MEF2A was mediated by the p38-dependent pathway in VSMCs. Previous reports have shown that MEF2 transcription factors are activated via the p38-dependent pathway.28,36 Our results were compatible with those findings. We also confirmed that serine 453 of MEF2A was critically important for the p38-dependent stimulation of the transactivating function of MEF2A in VSMCs. Because the dominant-negative mutant of MEF2A we used in this study, in which serine 453 was replaced with alanine, inhibited Ang II–induced expression of MCP-1, it seemed highly likely that Ang II stimulated MCP-1 expression, at least partly, by activating MEF2 transcription factors via the p38-dependent pathway.
To study the significance of the p38- and MEF2-dependent expression of MCP-1 in vivo, we transferred genes encoding a dominant-negative mutant of MEK6 and MEF2 using adenoviruses and examined their effects on the expression of MCP-1 and neointimal formation of the rat femoral artery after transluminal mechanical injury. We found that infection of AdMEK6AA and AdMEF2ASA significantly inhibited MCP-1 expression in the rat femoral artery. We also found that the infection of these viruses significantly suppressed both macrophages infiltration and neointimal formation. It has been shown that macrophages infiltrating the blood vessel wall play critical roles in neointimal formation. Inactivation of macrophages by an anti-CD4 antibody or bisphosphonate-containing liposomes, which kill macrophages after phagocytosis, resulted in significant suppression of neointimal formation after transluminal endothelial injury.37,38 Furthermore, inactivation of MCP-1 function using a neutralizing antibody for MCP-1 or by expressing a dominant-negative mutant of MCP-1, has been demonstrated to result in decreases of macrophages infiltration and neointimal formation after balloon injury.4,35 Thus, the p38- and MEF2-dependent increase of MCP-1 expression in vascular myocytes potentially stimulated macrophages infiltration and neointimal formation. Because it has been shown that MEF2 is activated via the p38-dependent pathway in monocytes,28 it is also possible that infection with AdMEK6AA and AdMEF2ASA inhibited neointimal formation, at least partly, by suppressing macrophages activation. Although it is also possible that MEF2 stimulated neointimal formation by directly stimulating the proliferation of VSMCs, we did not find any suppressive effects of AdMEF2ASA on the ET-1–induced proliferation of VSMCs as assessed by 3H-thymidine uptake. Thus, MEF2 did not seem to have a potent effect on the proliferation of VSMCs. Collectively, it is probable that the MEF2-dependent pathway was implicated in macrophages infiltration and neointimal formation by stimulating MCP-1 production in the vessel wall. Interestingly, although the p38-dependent pathway potentially activated other transcription factors as well as MEF2, blockade of the MEF2-dependent pathway was sufficient to inhibit macrophages infiltration and neointimal formation, suggesting critical roles of MEF2 in macrophages infiltration and neointimal formation after transluminal mechanical injury.
We have recently reported that the calcineurin-dependent pathway promotes the expression of MCP-1 in VSMCs and is involved in macrophages infiltration and neointimal formation after transluminal mechanical injury. The calcineurin-dependent pathway seems to be implicated in the stabilization of MCP-1 mRNA rather than in the transcriptional activation of the MCP-1 gene.31 In contrast, the p38- and MEF2-dependent pathways seemed to participate in the transcriptional activation of the MCP-1 gene. Thus, the calcineurin-dependent pathway and the MEF2-dependent pathway appear to cooperate to induce the expression of MCP-1 and stimulate vascular inflammation.
In summary, MEF2 seems to be activated via the p38-dependent pathway and to be implicated in the expression of MCP-1 in VSMCs. This function of MEF2 appears to mediate vascular inflammation, which is observed in conditions such as atherosclerosis and restenosis after angioplasty. MEF2 can be a novel molecular target to modulate vascular inflammation.
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Acknowledgments |
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This study was supported in part by Grants-in-Aid 13670695 (to E.S.), 15590725 (to E.S.), 13470141 (to Y.H.), and 10218202 (to Y.H.) and by the Advanced and Innovational Research program in Life Sciences (to Y.H.) from the Ministry of Education, Culture, Sports, Science and Technology of Japan. We thank Etsuko Taira, Marie Morita, and Reiko Sato for technical assistance. We also thank Novartis Pharma AG for supplying valsartan.
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Footnotes |
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Original received January 9, 2004; revision received May 19, 2004; accepted May 21, 2004.
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References |
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