- Email: [email protected]
Angio-Associated Migratory Cell Protein and Smooth Muscle Cell Migration in Development of Restenosis and Atherosclerosis⁎
Journal of the American College of Cardiology © 2008 by the American College of Cardiology Foundation Published by Elsevier Inc.
EDITORIAL COMMENT
Angio-Associated Migratory Cell Protein and Smooth Muscle Cell Migration in Development of Restenosis and Atherosclerosis* Paul Holvoet, PHD,† Peter Sinnaeve, MD, PHD‡ Leuven, Belgium
Migration of smooth muscle cells (SMCs) into the developing neointima is a pivotal step during atherosclerosis development in native vessels as well as in restenosis after vascular intervention. Before migration can occur, a switch of the SMC phenotype from the quiescent to the secretory phenotype and production of extracellular matrix are required (1–3). See page 302
Previously, Blindt et al. (1) analyzed differences in gene expression pattern between the quiescent and the invasive SMC phenotype in vitro. They identified 9 clusters of differentially regulated genes—the most important being cluster 3 containing 17 of the 47 deregulated genes, among them genes that are involved in cell communication, matrix organization, and cell adhesion and migration. One of them was the angio-associated migratory cell protein (AAMP) gene. It was first identified as a member of the immunoglobulin superfamily present in endothelial cells of all human tissues and shown to mediate cell– cell and cell– substrate interactions (4). In this issue of the Journal, Vogt et al. (5) demonstrated an important functional role of AAMP for migration of SMC in vitro and for the development of neointimal hyperplasia in vivo. The AAMP overexpression resulted in increased migration, whereas antibody- or silencing ribonucleic acid (RNA)–mediated inhibition of AAMP decreased SMC migration. Intraperitoneal administration of an anti-
*Editorials published in the Journal of the American College of Cardiology reflect the views of the authors and do not necessarily represent the views of JACC or the American College of Cardiology. From the †Atherosclerosis and Metabolism Unit and ‡Division of Cardiology, Department of Cardiovascular Diseases, Katholieke Universiteit Leuven, Leuven, Belgium.
Vol. 52, No. 4, 2008 ISSN 0735-1097/08/$34.00 doi:10.1016/j.jacc.2008.04.024
AAMP antibody reduced neointimal SMC density and neointima formation in apolipoprotein E– deficient mice. Their data show for the first time that AAMP inhibition offers a unique way of downstream targeting cytosolic RhoA pathways resulting in inhibition of SMC migration. Studies in experimental models demonstrated that mechanical stretch and interaction with extracellular matrix leads to AAMP-mediated activation of RhoA. Activated RhoA then interacts with its effector rho-kinase (ROCK), generating the driving force for SMCs to migrate and to divide, leading to restenosis and atherosclerosis (Fig. 1). Direct inhibition of ROCK by fasudil and Y-27632 inhibited vascular SMC migration (6). Similarly, 3-hydroxy-3methylglutaryl coenzyme A reductase inhibitors or statins blocked RhoA pathways (7). However, it remains to be determined whether statins do this by impairing RhoA translocation and consequent activation of RhoA or by binding to ROCK (Fig. 1). In addition, activation of RhoA pathways induced endothelial cell migration (8) and endothelial progenitor cell differentiation (9,10). Furthermore, RhoA/ROCK activation in endothelial cells decreased endothelial nitric oxide (NO) synthase (eNOS) expression by reducing eNOS messenger RNA stability. Consequently, ROCK inhibitors or statins upregulated eNOS expression (11,12) (Fig. 1). In aggregate, animal studies suggested that RhoA pathways might contribute to several cardiovascular pathologies, including arterial and pulmonary hypertension (13–15), myocardial hypertrophy and remodeling (16), coronary artery spasm (17), atherosclerosis (18), and restenosis (19,20). Recent data obtained in patients, although sparse, support in part ROCK’s involvement in these conditions. Fasudil attenuated coronary vasoconstrictor responses to acetylcholine in patients with angina (21) and ameliorated myocardial ischemia in patients having coronary microvascular spasm as well (22). It also prolonged symptom-limited exercise duration of patients with stable angina more than placebo without affecting heart rate or blood pressure and was well-tolerated (23). Fasudil improved endothelium-dependent vasodilation and decreased vascular resistance in patients with coronary heart disease as well (24,25). Indeed, oral fasudil during 1 month was shown in an elegant study to significantly improve endotheliumdependent brachial artery dilation in coronary artery disease patients (n ⫽ 13) but not in healthy subjects (n ⫽ 16). Because no effect of fasudil on endothelium-independent vasodilatation was observed in either groups these findings suggest that ROCK inhibition at least in part restores decreased NO availability associated with atherosclerosis (25). To our knowledge, proof of the effect of RhoA/ROCK inhibition on angioplasty- or stent-associated vascular SMC migration and proliferation in patients in vivo is lacking to date. However, inhibition of the RhoA/ROCK pathway does inhibit pulsatile stretch-induced proliferation of human saphenous vein SMCs (26), indicating that clinical studies with inhibitors of this
JACC Vol. 52, No. 4, 2008 July 22, 2008:312–4
Figure 1
Holvoet and Sinnaeve Editorial Comment
313
Schematic Presentation of the Role of AAMP in Rho Activation That Contributes to Cardiovascular Pathologies
Angio-associated migratory cell protein (AAMP) causes the translocation and activation of RhoA in smooth muscle cells (SMCs) and endothelial cells (ECs). The activated RhoA interacts with its effector rho-kinase (ROCK), inducing migration of SMCs and ECs and attachment of endothelial progenitor cells. Inhibition of AAMP in SMCs leads to inhibition of atherosclerosis and restenosis. Inhibition of AAMP in ECs and progenitor cells might impair re-endothelialization at the site of arterial injury, possibly leading to thrombosis. In contrast, AAMP inhibition might have an anti-thrombotic effect by restricting (impairing) endothelial nitric oxide (NO) synthase (eNOS) activity and NO production. Fasudil and Y-27432 are ROCK inhibitors (indicated by blunted arrows). Inhibition of ROCK is one of the putative (possible; not proven in humans) pleiotropic effects of statins; they might also impair translocation of RhoA. GDP ⫽ guanosine 5=-diphosphate; GTP ⫽ guanosine 5=-triphosphate. Figure illustration done by Rob Flewell.
signal transduction system for the prevention of vein graft disease or restenosis should be considered. One important limitation of fasudil and Y-27632 is that they are not specific for ROCK. They inhibit several other serine-threonine protein kinases, such as protein kinase A and C (27), which are also involved in several signaling pathways in the cardiovascular system, particularly in the regulation of myocardial contraction, hypertrophy, and remodeling (28). Therefore, the effects of currently available ROCK inhibitors might at least in part be due to blockage of other than RhoA pathways. However, there are no assays to evaluate how effective these agents are for inhibiting these specific pathways. The structure of ROCK dimers has been determined, in search of the mechanism for selectivity of ROCK. This structural analysis showed that subtle differences exist between the ROCK- and protein kinase A– bound conformations of the inhibitors. These structural differences are expected to determine the relative selectivity of these compounds. In in vitro binding studies, hydroxyfasudil—a metabolite of fasudil—was indeed selective for ROCK over protein kinase A (29). Its higher selectivity remains, however, to be confirmed in vivo.
The current study shows that blocking AAMP offers a possibility to downstream inhibition of RhoA-dependent pathways and therefore the development of restenosis and atherosclerosis. In addition, it offers the advantage of a well-defined target allowing more specific inhibition with high-affinity antibodies, silencing RNA, or synthetic inhibitors (30). The AAMP blockage might also improve endothelial function and increase NO availability, therefore enhancing its inhibitory effect on atherosclerosis and restenosis development. Some caution, however, is warranted. Indeed, AAMP also regulates the motility of endothelial cells (8) and is involved in endothelial progenitor cell differentiation (9). Thus, blocking AAMP in endothelial (progenitor) cells might impair re-endothelialization, leading to delayed arterial healing and promoting thrombosis (31), and therefore prevent adequate prevention of restenosis. Finally, further studies on the role of AAMP in regulating monocyte infiltration into the subendothelial space are needed. Indeed, ROCK inhibitors were shown to inhibit atherosclerosis by preventing macrophage migration (32) and inflammation (33).
314
Holvoet and Sinnaeve Editorial Comment
In conclusion, this study is important because it draws our attention to the functional role of AAMP as a regulator of SMC migration. Further studies of the effect of AAMP blockage on SMC migration and endothelial (progenitor) cell and macrophage migration and inflammation, which are important processes in the development of atherosclerosis and restenosis, are warranted. Reprint requests and correspondence: Dr. Paul Holvoet, Atherosclerosis and Metabolism Unit, Department of Cardiovascular Diseases, Katholieke Universiteit Leuven, Herestraat 49, PB 705, B-3000 Leuven, Belgium. E-mail: [email protected]. REFERENCES
1. Blindt R, Krott N, Hanrath P, vom Dahl J, van Eys G, Bosserhoff AK. Expression patterns of integrins on quiescent and invasive smooth muscle cells and impact on cell locomotion. J Mol Cell Cardiol 2002;34:1633– 44. 2. Blindt R, Vogt F, Lamby D, et al. Characterization of differential gene expression in quiescent and invasive human arterial smooth muscle cells. J Vasc Res 2002;39:340 –52. 3. Ross R. Atherosclerosis is an inflammatory disease. Am Heart J 1999;138:S419 –20. 4. Beckner ME, Krutzsch HC, Stracke ML, Williams ST, Gallardo JA, Liotta LA. Identification of a new immunoglobulin superfamily protein expressed in blood vessels with a heparin-binding consensus sequence. Cancer Res 1995;55:2140 –9. 5. Vogt F, Zernecke A, Beckner M, et al. Blockade of angio-associated migratory cell protein inhibits smooth muscle cell migration and neointima formation in accelerated atherosclerosis. J Am Coll Cardiol 2008;52:302–11. 6. Loirand G, Guerin P, Pacaud P. Rho kinases in cardiovascular physiology and pathophysiology. Circ Res 2006;98:322–334. 7. Cordle A, Koenigsknecht-Talboo J, Wilkinson B, Limpert A, Landreth G. Mechanisms of statin-mediated inhibition of small G-protein function. J Biol Chem 2005;280:34202–9. 8. Beckner ME, Jagannathan S, Peterson VA. Extracellular angioassociated migratory cell protein plays a positive role in angiogenesis and is regulated by astrocytes in coculture. Microvasc Res 2002;63: 259 – 69. 9. Martin CB, Mahon GM, Klinger MB, et al. The thrombin receptor, PAR-1, causes transformation by activation of Rho-mediated signaling pathways. Oncogene 2001;20:1953– 63. 10. Yin L, Morishige K, Takahashi T, et al. Fasudil inhibits vascular endothelial growth factor-induced angiogenesis in vitro and in vivo. Mol Cancer Ther 2007;6:1517–25. 11. Laufs U, La Fata V, Plutzky J, Liao JK. Upregulation of endothelial nitric oxide synthase by HMG CoA reductase inhibitors. Circulation 1998;97:1129 –35. 12. Laufs U, Liao JK. Post-transcriptional regulation of endothelial nitric oxide synthase mRNA stability by Rho GTPase. J Biol Chem 1998;273:24266 –71. 13. Abe K, Shimokawa H, Morikawa K, et al. Long-term treatment with a Rho-kinase inhibitor improves monocrotaline-induced fatal pulmonary hypertension in rats. Circ Res 2004;94:385–93. 14. Seko T, Ito M, Kureishi Y, et al. Activation of RhoA and inhibition of myosin phosphatase as important components in hypertension in vascular smooth muscle. Circ Res 2003;92:411– 8.
JACC Vol. 52, No. 4, 2008 July 22, 2008:312–4 15. Uehata M, Ishizaki T, Satoh H, et al. Calcium sensitization of smooth muscle mediated by a Rho-associated protein kinase in hypertension. Nature 1997;389:990 – 4. 16. Hattori T, Shimokawa H, Higashi M, et al. Long-term treatment with a specific Rho-kinase inhibitor suppresses cardiac allograft vasculopathy in mice. Circ Res 2004;94:46 –52. 17. Kandabashi T, Shimokawa H, Miyata K, et al. Inhibition of myosin phosphatase by upregulated rho-kinase plays a key role for coronary artery spasm in a porcine model with interleukin-1beta. Circulation 2000;101:1319 –23. 18. Mallat Z, Gojova A, Sauzeau V, et al. Rho-associated protein kinase contributes to early atherosclerotic lesion formation in mice. Circ Res 2003;93:884 – 8. 19. Matsumoto Y, Uwatoku T, Oi K, et al. Long-term inhibition of Rho-kinase suppresses neointimal formation after stent implantation in porcine coronary arteries: involvement of multiple mechanisms. Arterioscler Thromb Vasc Biol 2004;24:181– 6. 20. Sawada N, Itoh H, Ueyama K, et al. Inhibition of rho-associated kinase results in suppression of neointimal formation of ballooninjured arteries. Circulation 2000;101:2030 –3. 21. Masumoto A, Mohri M, Shimokawa H, Urakami L, Usui M, Takeshita A. Suppression of coronary artery spasm by the Rho-kinase inhibitor fasudil in patients with vasospastic angina. Circulation 2002;105:1545–7. 22. Mohri M, Shimokawa H, Hirakawa Y, Masumoto A, Takeshita A. Rho-kinase inhibition with intracoronary fasudil prevents myocardial ischemia in patients with coronary microvascular spasm. J Am Coll Cardiol 2003;41:15–9. 23. Vicari RM, Chaitman B, Keefe D, et al. Efficacy and safety of fasudil in patients with stable angina: a double-blind, placebo-controlled, phase 2 trial. J Am Coll Cardiol 2005;46:1803–11. 24. Kishi T, Hirooka Y, Masumoto A, et al. Rho-kinase inhibitor improves increased vascular resistance and impaired vasodilation of the forearm in patients with heart failure. Circulation 2005;111:2741–7. 25. Nohria A, Grunert ME, Rikitake Y, et al. Rho kinase inhibition improves endothelial function in human subjects with coronary artery disease. Circ Res 2006;99:1426 –32. 26. Kozai T, Eto M, Yang Z, Shimokawa H, Luscher TF. Statins prevent pulsatile stretch-induced proliferation of human saphenous vein smooth muscle cells via inhibition of Rho/Rho-kinase pathway. Cardiovasc Res 2005;68:475– 82. 27. Noma K, Oyama N, Liao JK. Physiological role of ROCKs in the cardiovascular system. Am J Physiol Cell Physiol 2006;290:C661– 8. 28. Avkiran M, Rowland AJ, Cuello F, Haworth RS. Protein kinase d in the cardiovascular system: emerging roles in health and disease. Circ Res 2008;102:157– 63. 29. Jacobs M, Hayakawa K, Swenson L, et al. The structure of dimeric ROCK I reveals the mechanism for ligand selectivity. J Biol Chem 2006;281:260 – 8. 30. Beckner ME, Krutzsch HC, Klipstein S, et al. AAMP, a newly identified protein, shares a common epitope with alpha-actinin and a fast skeletal muscle fiber protein. Exp Cell Res 1996;225:306 –14. 31. Luscher TF, Steffel J, Eberli FR, et al. Drug-eluting stent and coronary thrombosis: biological mechanisms and clinical implications. Circulation 2007;115:1051– 8. 32. Kim JS, Kim JG, Moon MY, et al. Transforming growth factor-beta1 regulates macrophage migration via RhoA. Blood 2006;108:1821–9. 33. Cui R, Tieu B, Recinos A, Tilton RG, Brasier AR. RhoA mediates angiotensin II-induced phospho-Ser536 nuclear factor kappaB/RelA subunit exchange on the interleukin-6 promoter in VSMCs. Circ Res 2006;99:723–30. Key Words: atherosclerosis y restenosis y kinases y migration y smooth muscle cell.
EDITORIAL COMMENT
Angio-Associated Migratory Cell Protein and Smooth Muscle Cell Migration in Development of Restenosis and Atherosclerosis* Paul Holvoet, PHD,† Peter Sinnaeve, MD, PHD‡ Leuven, Belgium
Migration of smooth muscle cells (SMCs) into the developing neointima is a pivotal step during atherosclerosis development in native vessels as well as in restenosis after vascular intervention. Before migration can occur, a switch of the SMC phenotype from the quiescent to the secretory phenotype and production of extracellular matrix are required (1–3). See page 302
Previously, Blindt et al. (1) analyzed differences in gene expression pattern between the quiescent and the invasive SMC phenotype in vitro. They identified 9 clusters of differentially regulated genes—the most important being cluster 3 containing 17 of the 47 deregulated genes, among them genes that are involved in cell communication, matrix organization, and cell adhesion and migration. One of them was the angio-associated migratory cell protein (AAMP) gene. It was first identified as a member of the immunoglobulin superfamily present in endothelial cells of all human tissues and shown to mediate cell– cell and cell– substrate interactions (4). In this issue of the Journal, Vogt et al. (5) demonstrated an important functional role of AAMP for migration of SMC in vitro and for the development of neointimal hyperplasia in vivo. The AAMP overexpression resulted in increased migration, whereas antibody- or silencing ribonucleic acid (RNA)–mediated inhibition of AAMP decreased SMC migration. Intraperitoneal administration of an anti-
*Editorials published in the Journal of the American College of Cardiology reflect the views of the authors and do not necessarily represent the views of JACC or the American College of Cardiology. From the †Atherosclerosis and Metabolism Unit and ‡Division of Cardiology, Department of Cardiovascular Diseases, Katholieke Universiteit Leuven, Leuven, Belgium.
Vol. 52, No. 4, 2008 ISSN 0735-1097/08/$34.00 doi:10.1016/j.jacc.2008.04.024
AAMP antibody reduced neointimal SMC density and neointima formation in apolipoprotein E– deficient mice. Their data show for the first time that AAMP inhibition offers a unique way of downstream targeting cytosolic RhoA pathways resulting in inhibition of SMC migration. Studies in experimental models demonstrated that mechanical stretch and interaction with extracellular matrix leads to AAMP-mediated activation of RhoA. Activated RhoA then interacts with its effector rho-kinase (ROCK), generating the driving force for SMCs to migrate and to divide, leading to restenosis and atherosclerosis (Fig. 1). Direct inhibition of ROCK by fasudil and Y-27632 inhibited vascular SMC migration (6). Similarly, 3-hydroxy-3methylglutaryl coenzyme A reductase inhibitors or statins blocked RhoA pathways (7). However, it remains to be determined whether statins do this by impairing RhoA translocation and consequent activation of RhoA or by binding to ROCK (Fig. 1). In addition, activation of RhoA pathways induced endothelial cell migration (8) and endothelial progenitor cell differentiation (9,10). Furthermore, RhoA/ROCK activation in endothelial cells decreased endothelial nitric oxide (NO) synthase (eNOS) expression by reducing eNOS messenger RNA stability. Consequently, ROCK inhibitors or statins upregulated eNOS expression (11,12) (Fig. 1). In aggregate, animal studies suggested that RhoA pathways might contribute to several cardiovascular pathologies, including arterial and pulmonary hypertension (13–15), myocardial hypertrophy and remodeling (16), coronary artery spasm (17), atherosclerosis (18), and restenosis (19,20). Recent data obtained in patients, although sparse, support in part ROCK’s involvement in these conditions. Fasudil attenuated coronary vasoconstrictor responses to acetylcholine in patients with angina (21) and ameliorated myocardial ischemia in patients having coronary microvascular spasm as well (22). It also prolonged symptom-limited exercise duration of patients with stable angina more than placebo without affecting heart rate or blood pressure and was well-tolerated (23). Fasudil improved endothelium-dependent vasodilation and decreased vascular resistance in patients with coronary heart disease as well (24,25). Indeed, oral fasudil during 1 month was shown in an elegant study to significantly improve endotheliumdependent brachial artery dilation in coronary artery disease patients (n ⫽ 13) but not in healthy subjects (n ⫽ 16). Because no effect of fasudil on endothelium-independent vasodilatation was observed in either groups these findings suggest that ROCK inhibition at least in part restores decreased NO availability associated with atherosclerosis (25). To our knowledge, proof of the effect of RhoA/ROCK inhibition on angioplasty- or stent-associated vascular SMC migration and proliferation in patients in vivo is lacking to date. However, inhibition of the RhoA/ROCK pathway does inhibit pulsatile stretch-induced proliferation of human saphenous vein SMCs (26), indicating that clinical studies with inhibitors of this
JACC Vol. 52, No. 4, 2008 July 22, 2008:312–4
Figure 1
Holvoet and Sinnaeve Editorial Comment
313
Schematic Presentation of the Role of AAMP in Rho Activation That Contributes to Cardiovascular Pathologies
Angio-associated migratory cell protein (AAMP) causes the translocation and activation of RhoA in smooth muscle cells (SMCs) and endothelial cells (ECs). The activated RhoA interacts with its effector rho-kinase (ROCK), inducing migration of SMCs and ECs and attachment of endothelial progenitor cells. Inhibition of AAMP in SMCs leads to inhibition of atherosclerosis and restenosis. Inhibition of AAMP in ECs and progenitor cells might impair re-endothelialization at the site of arterial injury, possibly leading to thrombosis. In contrast, AAMP inhibition might have an anti-thrombotic effect by restricting (impairing) endothelial nitric oxide (NO) synthase (eNOS) activity and NO production. Fasudil and Y-27432 are ROCK inhibitors (indicated by blunted arrows). Inhibition of ROCK is one of the putative (possible; not proven in humans) pleiotropic effects of statins; they might also impair translocation of RhoA. GDP ⫽ guanosine 5=-diphosphate; GTP ⫽ guanosine 5=-triphosphate. Figure illustration done by Rob Flewell.
signal transduction system for the prevention of vein graft disease or restenosis should be considered. One important limitation of fasudil and Y-27632 is that they are not specific for ROCK. They inhibit several other serine-threonine protein kinases, such as protein kinase A and C (27), which are also involved in several signaling pathways in the cardiovascular system, particularly in the regulation of myocardial contraction, hypertrophy, and remodeling (28). Therefore, the effects of currently available ROCK inhibitors might at least in part be due to blockage of other than RhoA pathways. However, there are no assays to evaluate how effective these agents are for inhibiting these specific pathways. The structure of ROCK dimers has been determined, in search of the mechanism for selectivity of ROCK. This structural analysis showed that subtle differences exist between the ROCK- and protein kinase A– bound conformations of the inhibitors. These structural differences are expected to determine the relative selectivity of these compounds. In in vitro binding studies, hydroxyfasudil—a metabolite of fasudil—was indeed selective for ROCK over protein kinase A (29). Its higher selectivity remains, however, to be confirmed in vivo.
The current study shows that blocking AAMP offers a possibility to downstream inhibition of RhoA-dependent pathways and therefore the development of restenosis and atherosclerosis. In addition, it offers the advantage of a well-defined target allowing more specific inhibition with high-affinity antibodies, silencing RNA, or synthetic inhibitors (30). The AAMP blockage might also improve endothelial function and increase NO availability, therefore enhancing its inhibitory effect on atherosclerosis and restenosis development. Some caution, however, is warranted. Indeed, AAMP also regulates the motility of endothelial cells (8) and is involved in endothelial progenitor cell differentiation (9). Thus, blocking AAMP in endothelial (progenitor) cells might impair re-endothelialization, leading to delayed arterial healing and promoting thrombosis (31), and therefore prevent adequate prevention of restenosis. Finally, further studies on the role of AAMP in regulating monocyte infiltration into the subendothelial space are needed. Indeed, ROCK inhibitors were shown to inhibit atherosclerosis by preventing macrophage migration (32) and inflammation (33).
314
Holvoet and Sinnaeve Editorial Comment
In conclusion, this study is important because it draws our attention to the functional role of AAMP as a regulator of SMC migration. Further studies of the effect of AAMP blockage on SMC migration and endothelial (progenitor) cell and macrophage migration and inflammation, which are important processes in the development of atherosclerosis and restenosis, are warranted. Reprint requests and correspondence: Dr. Paul Holvoet, Atherosclerosis and Metabolism Unit, Department of Cardiovascular Diseases, Katholieke Universiteit Leuven, Herestraat 49, PB 705, B-3000 Leuven, Belgium. E-mail: [email protected]. REFERENCES
1. Blindt R, Krott N, Hanrath P, vom Dahl J, van Eys G, Bosserhoff AK. Expression patterns of integrins on quiescent and invasive smooth muscle cells and impact on cell locomotion. J Mol Cell Cardiol 2002;34:1633– 44. 2. Blindt R, Vogt F, Lamby D, et al. Characterization of differential gene expression in quiescent and invasive human arterial smooth muscle cells. J Vasc Res 2002;39:340 –52. 3. Ross R. Atherosclerosis is an inflammatory disease. Am Heart J 1999;138:S419 –20. 4. Beckner ME, Krutzsch HC, Stracke ML, Williams ST, Gallardo JA, Liotta LA. Identification of a new immunoglobulin superfamily protein expressed in blood vessels with a heparin-binding consensus sequence. Cancer Res 1995;55:2140 –9. 5. Vogt F, Zernecke A, Beckner M, et al. Blockade of angio-associated migratory cell protein inhibits smooth muscle cell migration and neointima formation in accelerated atherosclerosis. J Am Coll Cardiol 2008;52:302–11. 6. Loirand G, Guerin P, Pacaud P. Rho kinases in cardiovascular physiology and pathophysiology. Circ Res 2006;98:322–334. 7. Cordle A, Koenigsknecht-Talboo J, Wilkinson B, Limpert A, Landreth G. Mechanisms of statin-mediated inhibition of small G-protein function. J Biol Chem 2005;280:34202–9. 8. Beckner ME, Jagannathan S, Peterson VA. Extracellular angioassociated migratory cell protein plays a positive role in angiogenesis and is regulated by astrocytes in coculture. Microvasc Res 2002;63: 259 – 69. 9. Martin CB, Mahon GM, Klinger MB, et al. The thrombin receptor, PAR-1, causes transformation by activation of Rho-mediated signaling pathways. Oncogene 2001;20:1953– 63. 10. Yin L, Morishige K, Takahashi T, et al. Fasudil inhibits vascular endothelial growth factor-induced angiogenesis in vitro and in vivo. Mol Cancer Ther 2007;6:1517–25. 11. Laufs U, La Fata V, Plutzky J, Liao JK. Upregulation of endothelial nitric oxide synthase by HMG CoA reductase inhibitors. Circulation 1998;97:1129 –35. 12. Laufs U, Liao JK. Post-transcriptional regulation of endothelial nitric oxide synthase mRNA stability by Rho GTPase. J Biol Chem 1998;273:24266 –71. 13. Abe K, Shimokawa H, Morikawa K, et al. Long-term treatment with a Rho-kinase inhibitor improves monocrotaline-induced fatal pulmonary hypertension in rats. Circ Res 2004;94:385–93. 14. Seko T, Ito M, Kureishi Y, et al. Activation of RhoA and inhibition of myosin phosphatase as important components in hypertension in vascular smooth muscle. Circ Res 2003;92:411– 8.
JACC Vol. 52, No. 4, 2008 July 22, 2008:312–4 15. Uehata M, Ishizaki T, Satoh H, et al. Calcium sensitization of smooth muscle mediated by a Rho-associated protein kinase in hypertension. Nature 1997;389:990 – 4. 16. Hattori T, Shimokawa H, Higashi M, et al. Long-term treatment with a specific Rho-kinase inhibitor suppresses cardiac allograft vasculopathy in mice. Circ Res 2004;94:46 –52. 17. Kandabashi T, Shimokawa H, Miyata K, et al. Inhibition of myosin phosphatase by upregulated rho-kinase plays a key role for coronary artery spasm in a porcine model with interleukin-1beta. Circulation 2000;101:1319 –23. 18. Mallat Z, Gojova A, Sauzeau V, et al. Rho-associated protein kinase contributes to early atherosclerotic lesion formation in mice. Circ Res 2003;93:884 – 8. 19. Matsumoto Y, Uwatoku T, Oi K, et al. Long-term inhibition of Rho-kinase suppresses neointimal formation after stent implantation in porcine coronary arteries: involvement of multiple mechanisms. Arterioscler Thromb Vasc Biol 2004;24:181– 6. 20. Sawada N, Itoh H, Ueyama K, et al. Inhibition of rho-associated kinase results in suppression of neointimal formation of ballooninjured arteries. Circulation 2000;101:2030 –3. 21. Masumoto A, Mohri M, Shimokawa H, Urakami L, Usui M, Takeshita A. Suppression of coronary artery spasm by the Rho-kinase inhibitor fasudil in patients with vasospastic angina. Circulation 2002;105:1545–7. 22. Mohri M, Shimokawa H, Hirakawa Y, Masumoto A, Takeshita A. Rho-kinase inhibition with intracoronary fasudil prevents myocardial ischemia in patients with coronary microvascular spasm. J Am Coll Cardiol 2003;41:15–9. 23. Vicari RM, Chaitman B, Keefe D, et al. Efficacy and safety of fasudil in patients with stable angina: a double-blind, placebo-controlled, phase 2 trial. J Am Coll Cardiol 2005;46:1803–11. 24. Kishi T, Hirooka Y, Masumoto A, et al. Rho-kinase inhibitor improves increased vascular resistance and impaired vasodilation of the forearm in patients with heart failure. Circulation 2005;111:2741–7. 25. Nohria A, Grunert ME, Rikitake Y, et al. Rho kinase inhibition improves endothelial function in human subjects with coronary artery disease. Circ Res 2006;99:1426 –32. 26. Kozai T, Eto M, Yang Z, Shimokawa H, Luscher TF. Statins prevent pulsatile stretch-induced proliferation of human saphenous vein smooth muscle cells via inhibition of Rho/Rho-kinase pathway. Cardiovasc Res 2005;68:475– 82. 27. Noma K, Oyama N, Liao JK. Physiological role of ROCKs in the cardiovascular system. Am J Physiol Cell Physiol 2006;290:C661– 8. 28. Avkiran M, Rowland AJ, Cuello F, Haworth RS. Protein kinase d in the cardiovascular system: emerging roles in health and disease. Circ Res 2008;102:157– 63. 29. Jacobs M, Hayakawa K, Swenson L, et al. The structure of dimeric ROCK I reveals the mechanism for ligand selectivity. J Biol Chem 2006;281:260 – 8. 30. Beckner ME, Krutzsch HC, Klipstein S, et al. AAMP, a newly identified protein, shares a common epitope with alpha-actinin and a fast skeletal muscle fiber protein. Exp Cell Res 1996;225:306 –14. 31. Luscher TF, Steffel J, Eberli FR, et al. Drug-eluting stent and coronary thrombosis: biological mechanisms and clinical implications. Circulation 2007;115:1051– 8. 32. Kim JS, Kim JG, Moon MY, et al. Transforming growth factor-beta1 regulates macrophage migration via RhoA. Blood 2006;108:1821–9. 33. Cui R, Tieu B, Recinos A, Tilton RG, Brasier AR. RhoA mediates angiotensin II-induced phospho-Ser536 nuclear factor kappaB/RelA subunit exchange on the interleukin-6 promoter in VSMCs. Circ Res 2006;99:723–30. Key Words: atherosclerosis y restenosis y kinases y migration y smooth muscle cell.