Circulation Research, Vol 74, 214-224, Copyright © 1994 by American Heart Association

ARTICLES

Osteopontin promotes vascular cell adhesion and spreading and is chemotactic for smooth muscle cells in vitro

L Liaw, M Almeida, CE Hart, SM Schwartz and CM Giachelli
Department of Pathology, University of Washington, Seattle 98195.

Osteopontin is an Arg-Gly-Asp-containing acidic phosphoprotein recently shown to be upregulated in vascular smooth muscle during rat arterial neointima formation and in human atherosclerotic plaques. Functional studies showed that osteopontin promoted adhesion of both cultured aortic endothelial cells and aortic smooth muscle cells. Adhesion of vascular cells to osteopontin was dose dependent and half maximal when solutions containing 7 and 30 nmol/L osteopontin were used to coat wells for endothelial and smooth muscle cells, respectively. Smooth muscle cells adherent to osteopontin were spread after 60 minutes, whereas endothelial cells remained round, although flattened, at this time point but were spread at 90 minutes. Cell spreading on osteopontin was accompanied by the formation of focal adhesion plaques. A newly developed anti-osteopontin antibody completely inhibited adhesion of both cell types to osteopontin but not to fibronectin or vitronectin. In addition, the peptide GRGDSP blocked adhesion to osteopontin, suggesting that integrins mediate Arg-Gly-Asp-dependent adhesion. Indeed, an antibody against the alpha v beta 3 integrin neutralized adhesion of both endothelium and smooth muscle cells to osteopontin by approximately 50%, demonstrating that alpha v beta 3 is one osteopontin receptor on vascular cells. Osteopontin also promoted the migration of smooth muscle cells in a Boyden-type chamber, with half-maximal effects observed at 77 nmol/L osteopontin. Checkerboard analysis demonstrated that this stimulus was chemotactic in nature. Our findings suggest that osteopontin may be functionally important as an adhesive and chemotactic molecule for vascular cells, particularly when levels of osteopontin are dramatically increased, as is the case after arterial angioplasty and in atherosclerotic plaques.

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				S.S. Srivatsa, L. A Fitzpatrick, P. W Tsao, T. M Reilly, D. R Holmes Jr, R. S Schwartz, and S. A Mousa Selective {alpha}v{beta}3 integrin blockade potently limits neointimal hyperplasia and lumen stenosis following deep coronary arterial stent injury:: Evidence for the functional importance of integrin {alpha}v{beta}3 and osteopontin expression during neointima formation Cardiovasc Res, December 1, 1997; 36(3): 408 - 428. [Abstract] [Full Text] [PDF]

				D. Panda, G. C. Kundu, B. I. Lee, A. Peri, D. Fohl, I. Chackalaparampil, B. B. Mukherjee, X. D. Li, D. C. Mukherjee, S. Seides, et al. Potential roles of osteopontin and alpha Vbeta 3 integrin in the development of coronary artery restenosis after angioplasty PNAS, August 19, 1997; 94(17): 9308 - 9313. [Abstract] [Full Text] [PDF]

				A. Hultgardh-Nilsson, C. Lovdahl, K. Blomgren, Bengt Kallin, and J. Thyberg Expression of phenotype- and proliferation-related genes in rat aortic smooth muscle cells in primary culture Cardiovasc Res, May 1, 1997; 34(2): 418 - 430. [Abstract] [Full Text] [PDF]

				E. R. Mohler III, L. P. Adam, P. McClelland, L. Graham, and D. R. Hathaway Detection of Osteopontin in Calcified Human Aortic Valves Arterioscler Thromb Vasc Biol, March 1, 1997; 17(3): 547 - 552. [Abstract] [Full Text]

				L. Liaw, D. M. Lombardi, M. M. Almeida, S. M. Schwartz, D. deBlois, and C. M. Giachelli Neutralizing Antibodies Directed Against Osteopontin Inhibit Rat Carotid Neointimal Thickening After Endothelial Denudation Arterioscler Thromb Vasc Biol, January 1, 1997; 17(1): 188 - 193. [Abstract] [Full Text]

				D. deBlois, D. M. Lombardi, E. J. Su, A. W. Clowes, S. M. Schwartz, and C. M. Giachelli Angiotensin II Induction of Osteopontin Expression and DNA Replication in Rat Arteries Hypertension, December 1, 1996; 28(6): 1055 - 1063. [Abstract] [Full Text]

				L. L. Smith, H.-K. Cheung, L. E. Ling, J. Chen, D. Sheppard, R. Pytela, and C. M. Giachelli Osteopontin N-terminal Domain Contains a Cryptic Adhesive Sequence Recognized by alpha 9beta 1 Integrin J. Biol. Chem., November 8, 1996; 271(45): 28485 - 28491. [Abstract] [Full Text] [PDF]

				X. Wang, C. Louden, E. H. Ohlstein, J. M. Stadel, J.-L. Gu, and T.-L. Yue Osteopontin Expression in Platelet-Derived Growth Factor–Stimulated Vascular Smooth Muscle Cells and Carotid Artery After Balloon Angioplasty Arterioscler Thromb Vasc Biol, November 1, 1996; 16(11): 1365 - 1372. [Abstract] [Full Text]

				M. P. Bendeck, S. Regenass, W. D. Tom, C. M. Giachelli, S. M. Schwartz, C. Hart, and M. A. Reidy Differential Expression of {alpha}1 Type VIII Collagen in Injured Platelet-Derived Growth Factor-BB–Stimulated Rat Carotid Arteries Circ. Res., September 1, 1996; 79(3): 524 - 531. [Abstract] [Full Text]

				A. Hultgardh-Nilsson, B. Cercek, J.-W. Wang, S. Naito, C. Lovdahl, B. Sharifi, J. S. Forrester, and J. A. Fagin Regulated Expression of the Ets-1 Transcription Factor in Vascular Smooth Muscle Cells In Vivo and In Vitro Circ. Res., April 1, 1996; 78(4): 589 - 595. [Abstract] [Full Text]

				J. Seki, N. Koyama, N. L. Kovach, T. Yednock, A. W. Clowes, and J. M. Harlan Regulation of ß1-Integrin Function in Cultured Human Vascular Smooth Muscle Cells Circ. Res., April 1, 1996; 78(4): 596 - 605. [Abstract] [Full Text]

				M. Hoshiga, C. E. Alpers, L. L. Smith, C. M. Giachelli, and S. M. Schwartz {alpha}vß3 Integrin Expression in Normal and Atherosclerotic Artery Circ. Res., December 1, 1995; 77(6): 1129 - 1135. [Abstract] [Full Text]

				D. D. Hu, E. C. K. Lin, N. L. Kovach, J. R. Hoyer, and J. W. Smith A Biochemical Characterization of the Binding of Osteopontin to Integrins alpha(v)beta(1) and alpha(v)beta(5) J. Biol. Chem., November 3, 1995; 270(44): 26232 - 26238. [Abstract] [Full Text] [PDF]

				K. D. O'Brien, J. Kuusisto, D. D. Reichenbach, M. Ferguson, C. Giachelli, C. E. Alpers, and C. M. Otto Osteopontin Is Expressed in Human Aortic Valvular Lesions Circulation, October 15, 1995; 92(8): 2163 - 2168. [Abstract] [Full Text]

				L. Liaw, V. Lindner, S. M. Schwartz, A. F. Chambers, and C. M. Giachelli Osteopontin and ß3 Integrin Are Coordinately Expressed in Regenerating Endothelium In Vivo and Stimulate Arg-Gly-Asp–Dependent Endothelial Migration In Vitro Circ. Res., October 1, 1995; 77(4): 665 - 672. [Abstract] [Full Text]

				H. Hao, S. Hirota, Y. Tsukamoto, M. Imakita, H. Ishibashi-Ueda, and C. Yutani Alterations of Bone Matrix Protein mRNA Expression in Rat Aorta In Vitro Arterioscler Thromb Vasc Biol, September 1, 1995; 15(9): 1474 - 1480. [Abstract] [Full Text]

				E. B. Williams, I. Halpert, S. Wickline, G. Davison, W. C. Parks, and J. N. Rottman Osteopontin Expression Is Increased in the Heritable Cardiomyopathy of Syrian Hamsters Circulation, August 15, 1995; 92(4): 705 - 709. [Abstract] [Full Text]

				S. L. Stanley Jr., K. Tian, J. P. Koester, and E. Li The Serine-rich Entamoeba histolytica Protein Is a Phosphorylated Membrane Protein Containing O-Linked Terminal N-Acetylglucosamine Residues J. Biol. Chem., February 24, 1995; 270(8): 4121 - 4126. [Abstract] [Full Text] [PDF]

				M. Gilbert, W. J. Shaw, J. R. Long, K. Nelson, G. P. Drobny, C. M. Giachelli, and P. S. Stayton Chimeric Peptides of Statherin and Osteopontin That Bind Hydroxyapatite and Mediate Cell Adhesion J. Biol. Chem., May 19, 2000; 275(21): 16213 - 16218. [Abstract] [Full Text] [PDF]

				K. J. Bayless and G. E. Davis Identification of Dual alpha 4beta 1 Integrin Binding Sites within a 38 Amino Acid Domain in the N-terminal Thrombin Fragment of Human Osteopontin J. Biol. Chem., April 13, 2001; 276(16): 13483 - 13489. [Abstract] [Full Text] [PDF]

				R. Agnihotri, H. C. Crawford, H. Haro, L. M. Matrisian, M. C. Havrda, and L. Liaw Osteopontin, a Novel Substrate for Matrix Metalloproteinase-3 (Stromelysin-1) and Matrix Metalloproteinase-7 (Matrilysin) J. Biol. Chem., July 20, 2001; 276(30): 28261 - 28267. [Abstract] [Full Text] [PDF]

				S. Jono, C. Peinado, and C. M. Giachelli Phosphorylation of Osteopontin Is Required for Inhibition of Vascular Smooth Muscle Cell Calcification J. Biol. Chem., June 23, 2000; 275(26): 20197 - 20203. [Abstract] [Full Text] [PDF]

				M. Rezaee, K. Penta, and T. Quertermous Del1 mediates VSMC adhesion, migration, and proliferation through interaction with integrin alpha vbeta 3 Am J Physiol Heart Circ Physiol, May 1, 2002; 282(5): H1924 - H1932. [Abstract] [Full Text] [PDF]

				Y. Oyama, N. Akuzawa, R. Nagai, and M. Kurabayashi PPAR{gamma} Ligand Inhibits Osteopontin Gene Expression Through Interference With Binding of Nuclear Factors to A/T-Rich Sequence in THP-1 Cells Circ. Res., February 22, 2002; 90(3): 348 - 355. [Abstract] [Full Text] [PDF]