Thrombin Stimulates Vascular Smooth Muscle Cell Polyamine Synthesis by Inducing Cationic Amino Acid Transporter and Ornithine Decarboxylase Gene Expression

From the Houston VA Medical Center (W.D., L.L., K.J.P., A.I.S.) and the Departments of Medicine (W.D., L.L., A.I.S.) and Pharmacology (W.D.), Baylor College of Medicine, Houston, Tex.

Correspondence to William Durante, PhD, Houston VA Medical Center, Building 109, Room 128, 2002 Holcombe Blvd, Houston, TX 77030.

Abstract—Thrombin, a serine protease, is a potent mitogen for vascular smooth muscle cells (SMCs), but its mechanism of action is not known. Since L-ornithine is metabolized to growth-stimulatory polyamines, we examined whether thrombin regulates the transcellular transport and metabolism of L-ornithine by vascular SMCs. Treatment of SMCs with thrombin initially (0 to 2 hours) decreased L-ornithine uptake, whereas longer exposures (6 to 24 hours) progressively increased transport. Kinetic studies indicated that thrombin-induced inhibition was associated with a decrease in affinity for L-ornithine, whereas stimulation was mediated by an increase in transport capacity. Thrombin induced the expression of both cationic amino acid transporter (CAT)-1 and CAT-2 mRNA. Furthermore, thrombin stimulated L-ornithine metabolism by inducing ornithine decarboxylase (ODC) mRNA expression and activity. The stimulatory effect of thrombin on both L-ornithine transport and ODC activity was reversed by hirudin, a thrombin inhibitor, and was mimicked by a 14–amino acid thrombin receptor–activating peptide. Thrombin also markedly increased the capacity of SMCs to generate putrescine, a polyamine, from extracellular L-ornithine. The thrombin-mediated increase in putrescine production was reversed by N^G-methyl-L-arginine, a competitive inhibitor of cationic amino acid transport, or by -difluoromethylornithine (DFMO), an ODC inhibitor. DFMO also inhibited thrombin-induced SMC proliferation. These results demonstrate that thrombin stimulates polyamine synthesis by inducing CAT and ODC gene expression and that thrombin-stimulated SMC proliferation is dependent on polyamine formation. The ability of thrombin to upregulate L-ornithine transport and direct its metabolism to growth-stimulatory polyamines may contribute to postangioplasty restenosis and atherosclerotic lesion formation.

Key Words: thrombin • L-ornithine • polyamine • proliferation

This article has been cited by other articles:

				K. J. Peyton, D. Ensenat, M. A. Azam, A. N. Keswani, S. Kannan, X.-m. Liu, H. Wang, D. A. Tulis, and W. Durante Arginase Promotes Neointima Formation in Rat Injured Carotid Arteries Arterioscler. Thromb. Vasc. Biol., April 1, 2009; 29(4): 488 - 494. [Abstract] [Full Text] [PDF]

				L. H. Wei, Y. Yang, G. Wu, and L. J. Ignarro IL-4 and IL-13 upregulate ornithine decarboxylase expression by PI3K and MAP kinase pathways in vascular smooth muscle cells Am J Physiol Cell Physiol, May 1, 2008; 294(5): C1198 - C1205. [Abstract] [Full Text] [PDF]

				J. Dumont, M. Zureik, C. Bauters, M.-C. Grupposo, D. Cottel, M. Montaye, M. Hamon, P. Ducimetiere, P. Amouyel, and T. Brousseau Association of OAZ1 Gene Polymorphisms With Subclinical and Clinical Vascular Events Arterioscler. Thromb. Vasc. Biol., October 1, 2007; 27(10): 2120 - 2126. [Abstract] [Full Text] [PDF]

				J. Dumont, M. Zureik, D. Cottel, M. Montaye, P. Ducimetiere, P. Amouyel, and T. Brousseau Association of arginase 1 gene polymorphisms with the risk of myocardial infarction and common carotid intima media thickness J. Med. Genet., August 1, 2007; 44(8): 526 - 531. [Abstract] [Full Text] [PDF]

				S. Kaneko, E. Okuda-Ashitaka, A. Ando, K. Nishimura, K. Igarashi, M. Maeda, K. Furuta, M. Suzuki, M. Matsumura, and S. Ito Polyamines upregulate the mRNA expression of cationic amino acid transporter-1 in human retinal pigment epithelial cells Am J Physiol Cell Physiol, August 1, 2007; 293(2): C729 - C737. [Abstract] [Full Text] [PDF]

				D. Teupser, R. Burkhardt, W. Wilfert, I. Haffner, K. Nebendahl, and J. Thiery Identification of Macrophage Arginase I as a New Candidate Gene of Atherosclerosis Resistance Arterioscler. Thromb. Vasc. Biol., February 1, 2006; 26(2): 365 - 371. [Abstract] [Full Text] [PDF]

				D. Teixeira, M. L. Santaolaria, V. Meneu, and E. Alonso Dietary Arginine Slightly and Variably Affects Tissue Polyamine Levels in Male Swiss Albino Mice J. Nutr., December 1, 2002; 132(12): 3715 - 3720. [Abstract] [Full Text] [PDF]

				W. Durante, L. Liao, S. V. Reyna, K. J. Peyton, and A. I. Schafer Transforming Growth Factor-{beta}1 Stimulates L-Arginine Transport and Metabolism in Vascular Smooth Muscle Cells : Role in Polyamine and Collagen Synthesis Circulation, February 27, 2001; 103(8): 1121 - 1127. [Abstract] [Full Text] [PDF]

				L. J. Van Winkle Amino Acid Transport Regulation and Early Embryo Development Biol Reprod, January 1, 2001; 64(1): 1 - 12. [Abstract] [Full Text]

				W. DURANTE, L. LIAO, S. V. REYNA, K. J. PEYTON, and A. I. SCHAFER Physiological cyclic stretch directs L-arginine transport and metabolism to collagen synthesis in vascular smooth muscle FASEB J, September 1, 2000; 14(12): 1775 - 1783. [Abstract] [Full Text]

				F. Wu, B. Cholewa, and D. L. Mattson Characterization of L-arginine transporters in rat renal inner medullary collecting duct Am J Physiol Regulatory Integrative Comp Physiol, June 1, 2000; 278(6): R1506 - R1512. [Abstract] [Full Text] [PDF]