Abstract
Phenotypic modulation of vascular smooth muscle cells (VSMC) and reactive oxygen species (ROS) is important in vascular pathogenesis. Understanding how these factors relate to cell migration can improve design of therapeutic interventions to control vascular disease. We compared the proliferation, protein content and migration of cultured aortic VSMC from wild type (WT) versus transgenic mice (Tgp22phox), in which overexpression of p22phox was targeted to VSMC. Also, we compared H2O2 generation and expression of specific phenotypic markers of non-migrating with migrating WT versus Tgp22phox VSMC in an in vitro wound scratch model. Enhanced H2O2 production in Tgp22phox versus WT VSMC (p < 0.005) significantly correlated with increased protein content, proliferation, and migration. VSMC migrating across the wound edge produced more H2O2 than non-migrating VSMC (p < 0.05). The expression of synthetic phenotypic markers, tropomyosin 4 and myosin heavy chain embryonic (SMemb), was enhanced significantly, while the expression of contractile marker, smooth muscle α-actin, was reduced significantly in migrating versus non-migrating cells, and also in Tgp22phox versus WT (p < 0.005) VSMC. These results are consistent with increased production of ROS accelerating the switch from the contractile to the synthetic phenotype, characterized by increases in proliferation, migration, and expression of TM4 and SMemb and decreased α-actin.
Similar content being viewed by others
References
Abouhamed, M., S. Reichenberg, H. Robenek, and G. Plenz. Tropomyosin 4 expression is enhanced in dedifferentiating smooth muscle cells in vitro and during atherogenesis. Eur. J. Cell. Biol. 82:473–482, 2003.
Aikawa, M., E. Rabkin, S. J. Voglic, H. Shing, R. Nagai, F. J. Schoen, and P. Libby. Lipid lowering promotes accumulation of mature smooth muscle cells expressing smooth muscle myosin heavy chain isoforms in rabbit atheroma. Circ. Res. 83:1015–1026, 1998.
Arnold, R. S., J. Shi, E. Murad, A. M. Whalen, C. Q. Sun, R. Polavarapu, S. Parthasarathy, J. A. Petros, and J. D. Lambeth. Hydrogen peroxide mediates the cell growth and transformation caused by the mitogenic oxidase Nox1. Proc. Natl. Acad. Sci. U.S.A. 98:5550–5555, 2001.
Azumi, H., N. Inoue, Y. Ohashi, M. Terashima, T. Mori, H. Fujita, K. Awano, K. Kobayashi, K. Maeda, K. Hata, T. Shinke, S. Kobayashi, K. Hirata, S. Kawashima, H. Itabe, Y. Hayashi, S. Imajoh-Ohmi, H. Itoh, and M. Yokoyama. Superoxide generation in directional coronary atherectomy specimens of patients with angina pectoris: Important role of NAD(P)H oxidase. Arterioscler. Thromb. Vasc. Biol. 22:1838–1844, 2002.
Azumi, H., N. Inoue, S. Takeshita, Y. Rikitake, S. Kawashima, Y. Hayashi, H. Itoh, and M. Yokoyama. Expression of NADH/NADPH oxidase p22phox in human coronary arteries. Circulation 100:1494–1498, 1999.
Bai, H., J. Masuda, Y. Sawa, S. Nakano, R. Shirakura, Y. Shimazaki, J. Ogata, and H. Matsuda. Neointima formation after vascular stent implantation. Spatial and chronological distribution of smooth muscle cell proliferation and phenotypic modulation. Arterioscler. Thromb. Vasc. Biol. 14:1846–1853, 1994.
Bayraktutan, U. Free radicals, diabetes and endothelial dysfunction. Diabetes Obes. Metab. 4:224–238, 2002.
Cahilly, C., C. M. Ballantyne, D. S. Lim, A. Gotto, and A. J. Marian. A variant of p22(phox), involved in generation of reactive oxygen species in the vessel wall, is associated with progression of coronary atherosclerosis. Circ. Res. 86:391–395, 2000.
Dalle-Donne, I., R. Rossi, A. Milzani, P. Di Simplicio, and R. Colombo. The actin cytoskeleton response to oxidants: From small heat shock protein phosphorylation to changes in the redox state of actin itself. Free Radic. Biol. Med. 31:1624–1632, 2001.
Fukui, T., N. Ishizaka, S. Rajagopalan, J. B. Laursen, Q. T. Capers, W. R. Taylor, D. G. Harrison, H. de Leon, J. N. Wilcox, and K. K. Griendling. p22phox mRNA expression and NADPH oxidase activity are increased in aortas from hypertensive rats. Circ. Res. 80:45–51, 1997.
Galis, Z. S., C. Johnson, D. Godin, R. Magid, J. M. Shipley, R. M. Senior, and E. Ivan. Targeted disruption of the matrix metalloproteinase-9 gene impairs smooth muscle cell migration and geometrical arterial remodeling. Circ. Res. 91:852–859, 2002.
Godin, D., E. Ivan, C. Johnson, R. Magid, and Z. S. Galis. Remodeling of carotid artery is associated with increased expression of matrix metalloproteinases in mouse blood flow cessation model. Circulation 102:2861–2866, 2000.
Gorlach, A., I. Diebold, V. B. Schini-Kerth, U. Berchner-Pfannschmidt, U. Roth, R. P. Brandes, T. Kietzmann, and R. Busse. Thrombin activates the hypoxia-inducible factor-1 signaling pathway in vascular smooth muscle cells: Role of the p22(phox)-containing NADPH oxidase. Circ. Res. 89:47–54, 2001.
Hao, H., P. Ropraz, V. Verin, E. Camenzind, A. Geinoz, M. S. Pepper, G. Gabbiani, and M. L. Bochaton-Piallat. Heterogeneity of smooth muscle cell populations cultured from pig coronary artery. Arterioscler. Thromb. Vasc. Biol. 22:1093–1099, 2002.
Huang, C., Z. Rajfur, C. Borchers, M. D. Schaller, and K. Jacobson. JNK phosphorylates paxillin and regulates cell migration. Nature 424:219–223, 2003.
Itoh, S., S. Umemoto, M. Hiromoto, Y. Toma, Y. Tomochika, S. Aoyagi, M. Tanaka, T. Fujii, and M. Matsuzaki. Importance of NAD(P)H oxidase-mediated oxidative stress and contractile type smooth muscle myosin heavy chain SM2 at the early stage of atherosclerosis. Circulation 105:2288–2295, 2002.
Johnson, C., and Z. S. Galis. Matrix metalloproteinase-2 and -9 differentially regulate smooth muscle cell migration and cell-mediated collagen organization. Arterioscler. Thromb. Vasc. Biol. 24:54–60, 2004.
Kataoka, N., S. Ujita, and M. Sato. Effect of flow direction on the morphological responses of cultured bovine aortic endothelial cells. Med. Biol. Eng. Comput. 36:122–128, 1998.
Khatri, J. J., C. Johnson, R. Magid, S. M. Lessner, K. M. Laude, S. I. Dikalov, D. G. Harrison, H. J. Sung, Y. Rong, and Z. S. Galis. Vascular oxidant stress enhances progression and angiogenesis of experimental atheroma. Circulation 109:520–525, 2004.
Kuro-o, M., R. Nagai, K. Nakahara, H. Katoh, R. C. Tsai, H. Tsuchimochi, Y. Yazaki, A. Ohkubo, and F. Takaku. cDNA cloning of a myosin heavy chain isoform in embryonic smooth muscle and its expression during vascular development and in arteriosclerosis. J. Biol. Chem. 266:3768–3773, 1991.
Lampugnani, M. G. Cell migration into a wounded area in vitro. Methods Mol. Biol. 96:177–182, 1999.
Lee, S. L., W. W. Wang, and B. L. Fanburg. Superoxide as an intermediate signal for serotonin-induced mitogenesis. Free Radic. Biol. Med. 24:855–858, 1998.
Lemire, J. M., M. J. Merrilees, K. R. Braun, and T. N. Wight. Overexpression of the V3 variant of versican alters arterial smooth muscle cell adhesion, migration, and proliferation in vitro. J. Cell. Physiol. 190:38–45, 2002.
Li, P. F., R. Dietz, and R. von Harsdorf. Differential effect of hydrogen peroxide and superoxide anion on apoptosis and proliferation of vascular smooth muscle cells. Circulation 96:3602–3609, 1997.
Li, S., S. Sims, Y. Jiao, L. H. Chow, and J. G. Pickering. Evidence from a novel human cell clone that adult vascular smooth muscle cells can convert reversibly between noncontractile and contractile phenotypes. Circ. Res. 85:338–348, 1999.
Miller, F. J., D. D. Jr. Gutterman, C. D. Rios, D. D. Heistad, and B. L. Davidson. Superoxide production in vascular smooth muscle contributes to oxidative stress and impaired relaxation in atherosclerosis. Circ. Res. 82:1298–1305, 1998.
Nakamura, Y., Q. Feng, T. Kumagai, K. Torikai, H. Ohigashi, T. Osawa, N. Noguchi, E. Niki, and K. Uchida. Ebselen, a glutathione peroxidase mimetic seleno-organic compound, as a multifunctional antioxidant. Implication for inflammation-associated carcinogenesis. J. Biol. Chem. 277:2687–2694, 2002.
Newby, A. C., and A. B. Zaltsman. Molecular mechanisms in intimal hyperplasia. J. Pathol. 190:300–309, 2000.
Owens, G. K., M. S. Kumar, and B. R. Wamhoff. Molecular regulation of vascular smooth muscle cell differentiation in development and disease. Physiol. Rev. 84:767–801, 2004.
Rovner, A. S., R. A. Murphy, and G. K. Owens. Expression of smooth muscle and nonmuscle myosin heavy chains in cultured vascular smooth muscle cells. J. Biol. Chem. 261:14740–14745, 1986.
Stocker, R., and J. F. Jr. Keaney. Role of oxidative modifications in atherosclerosis. Physiol. Rev. 84:1381–1478, 2004.
Thyberg, J. Phenotypic modulation of smooth muscle cells during formation of neointimal thickenings following vascular injury. Histol. Histopathol. 13:871–891, 1998.
West, N., T. Guzik, E. Black, and K. Channon. Enhanced superoxide production in experimental venous bypass graft intimal hyperplasia: Role of NAD(P)H oxidase. Arteriosclr. Thromb. Vasc. Biol. 21:189–194, 2001.
Worth, N. F., B. E. Rolfe, J. Song, and G. R. Campbell. Vascular smooth muscle cell phenotypic modulation in culture is associated with reorganisation of contractile and cytoskeletal proteins. Cell. Motil. Cytoskeleton 49:130–145, 2001.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Sung, HJ., Eskin, S.G., Sakurai, Y. et al. Oxidative Stress Produced with Cell Migration Increases Synthetic Phenotype of Vascular Smooth Muscle Cells. Ann Biomed Eng 33, 1546–1554 (2005). https://doi.org/10.1007/s10439-005-7545-2
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/s10439-005-7545-2