Fibrin Gel Induces the Migration of Smooth Muscle Cells from Rabbit Aortic Explants

Hideki Nomura; Michitaka Naito; Akihisa Iguchi; W. Douglas Thompson; Elspeth B. Smith

doi:10.1055/s-0037-1614388

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 1999; 82(04): 1347-1352
DOI: 10.1055/s-0037-1614388

Review Article

Schattauer GmbH

Fibrin Gel Induces the Migration of Smooth Muscle Cells from Rabbit Aortic Explants

Authors

Hideki Nomura

¹Departments of Pathology and Clinical Biochemistry, University of Aberdeen, Aberdeen, UK
Michitaka Naito

¹Departments of Pathology and Clinical Biochemistry, University of Aberdeen, Aberdeen, UK
Akihisa Iguchi

¹Departments of Pathology and Clinical Biochemistry, University of Aberdeen, Aberdeen, UK
W. Douglas Thompson

¹Departments of Pathology and Clinical Biochemistry, University of Aberdeen, Aberdeen, UK
Elspeth B. Smith

¹Departments of Pathology and Clinical Biochemistry, University of Aberdeen, Aberdeen, UK

Abstract

Summary

A major step in the pathogenesis of atherosclerosis is the vectorial migration of smooth muscle cells (SMCs) from the arterial media into the intima. Although subcultured SMCs usually show synthetic phenotype, the behaviour of contractile SMCs may be crucial for the subsequent migration of the cells. In the present study, we utilized an in vitro assay system to evaluate the effects of fibrin gels on the migration of SMCs from explants taken from rabbit aorta. After cultured for 5-7 days in a serum-free condition, SMCs appeared from explants covered with fibrin gel. The cells were positive on immunostaining for SMC specific α-actin. No migration of SMCs from the control explants without fibrin gel was observed. Then the percentage of explants showing cell migration and the number of migrating cells increased with time. The migration of SMCs into fibrin gels was not dependent on the concentration of fibrinogen used for the preparation of fibrin gel in the range of 1.5-3 mg/ml. Variations of thrombin concentration in the range of 0.25-1.25 U/ml had no significant effect. However, there was less migration of SMCs with higher concentrations of thrombin. Thrombin inhibitors, hirudin and PPACK had no significant effect on the migration of SMCs. An RGD-containing peptide, GRGDS inhibited the migration of SMCs although a control peptide GRGES at the same concentration had no significant effect. A monoclonal antibody to α_vβ₃, LM609, completely inhibited the migration of SMCs from the explants, suggesting that α_vβ₃ integrin is involved in the migration of SMCs into fibrin gels. SMCs which migrated from the explants showed the positive staining with the monoclonal antibodies against SMC myosin heavy chain isoforms, SMemb, SM1 and SM2, suggesting that they are in an intermediate state changing from contractile to synthetic state. In conclusion, the present study showed that fibrin gel induces the migration of SMCs from explants into itself and the process may not need other growth factors or cytokines.

Key Words

Fibrin gel, smooth muscle cell, migration, phenotype, integrin α_vβ₃

Full Text

References

References
1 Smith EB, Slater RS, Hunter JA. Quantitative studies on fibrinogen and low density lipoprotein in human aortic intima. Atherosclerosis 1973; 18: 479-87.
2 Smith EB, Thompson WD. Fibrin as a factor in atherogenesis. Thromb Res 1994; 73: 1-19.
3 Shekhonin BV, Tararak EM, Samokhin GP, Mitkevich OV, Mazurov AV, Vinogradov DV, Vlasik TN, Kalantarov GF, Koteliansky VE. Visualization of apo B, fibrinogen/fibrin, and fibronectin in the intima of normal human aorta and large arteries and during atherosclerosis. Atherosclerosis 1990; 213-26.
4 Naito M, Hayashi T, Iguchi A. New approaches to the prevention of atherosclerosis. Drugs 1995; 50: 440-53.
5 Bini A, Fenoglio JJ, Sobel J, Owen J, Fejgi M, Kaplan KL. Immunochemical characterization of fibrinogen, fibrin I and fibrin II in human thrombi and atherosclerotic lesions. Blood 1987; 69: 1038-45.
6 Bini A, Fenoglio JJ, Mesa-Tejada R, Kudryk B, Kaplan KL. Identification and distribution of fibrinogen, fibrin and fibrin(ogen) degradation products in atherosclerosis. Arteriosclerosis 1989; 9: 109-21.
7 Smith EB, Keen GA, Grant A, Stirk C. Fate of fibrinogen in human arterial intima. Arteriosclerosis 1990; 10: 263-75.
8 Smith EB, Alexander KM, Massie IB. Insoluble “fibrin” in human aortic intima. Quantitative studies on the relationship between insoluble “fibrin”, soluble fibrinogen and low density lipoprotein. Atherosclerosis 1976; 23: 19-39.
9 Ross R. The pathogenesis of atherosclerosis: a perspective for the 1990s. Nature 1993; 362: 801-9.
10 Naito M, Hayashi T, Kuzuya M, Funaki C, Asai K, Kuzuya F. Fibrinogen is chemotactic for vascular smooth muscle cells. FEBS Lett 1989; 247: 358-60.
11 Naito M, Hayashi T, Kuzuya M, Funaki C, Asai K, Kuzuya F. Effects of fibrinogen and fibrin on the migration of vascular smooth muscle cells in vitro. Atherosclerosis 1990; 83: 9-14.
12 Naito M, Funaki C, Hayashi T, Yamada K, Asai K, Yoshimine N, Kuzuya F. Substrate-bound fibrinogen, fibrin and other cell attachment-promoting proteins as a scaffold for cultured vascular smooth muscle cells. Atherosclerosis 1992; 96: 227-34.
13 Naito M, Nomura H, Iguchi A. Migration of cultured vascular smooth muscle cells into non-crosslinked fibrin gels. Thromb Res 1996; 84: 129-36.
14 Naito M, Nomura H, Iguchi A, Thompson WD, Smith EB. Effect of cross-linking by factor XIIIa on the migration of vascular smooth muscle cells into fibrin gels. Thromb Res 1998; 90: 111-6.
15 Chamley-Campbell J, Campbell GR, Ross R. The smooth muscle cell in culture. Physiol Rev 1979; 59: 1-61.
16 Thyberg J, Hedin U, Sjolund M, Palmberg L, Bottger BA. Regulation of differentiated properties and proliferation of arterial smooth muscle cells. Arteriosclerosis 1990; 10: 966-90.
17 Rekhter MD, Andreeva AA, Mironov AA, Orekov AN. Three-dimensional cytoarchitecture of normal and atherosclerotic intima of human aorta. Am J Pathol 1991; 138: 569-80.
18 Kuro-o M, Nagai R, Nakahara K, Katoh H, Tsai RC, Tsuchimochi H, Yazaki Y, Ohkubo A, Takaku F. cDNA cloning of a myosin heavy chain isoform in embryonic smooth muscle and its expression during vascular development and in arteriosclerosis. J Biol Chem 1991; 266: 3768-73.
19 Thompson WD, Smith EB, Stirk CM, Marshall FI, Stout AJ, Kocchar A. Angiogenic activity of fibrin degradation products is located in fibrin fragment E. J Pathol 1992; 168: 47-53.
20 Stirk CM, Kocchar A, Smith EB, Thompson WD. Presence of growth-stimulating fibrin degradation products containing fragment E in human atherosclerotic plaques. Atherosclerosis 1993; 103: 159-69.
21 Naito M, Sabally K, Thompson WD, Stirk CM, Smith EB, Benjamin N. Stimulation of proliferation of smooth muscle cells in culture by fibrin degradation products. Fibrinolysis 1996; 10 (Suppl. 04) 12.
22 Freedman JE, Fabian A, Loscalzo J. Impaired EDRF production by endothelial cells exposed to fibrin monomer and FDP. Am J Physiol 1995; 268: C520-6.
23 Ge M, Ryan TJ, Lum H, Malik AB. Fibrinogen degradation product fragment D increases endothelial monolayer permeability. Am J Physiol 1991; 261: L283-9.
24 Scrutton MC, Rossmurphy SB, Bennett GM. et al. Changes in clot deformability – a possible explanation for the epidemiological association between plasma fibrinogen concentration and myocardial infarction. Blood Coag Fibrinol 1994; 5: 719-23.
25 Fatah K, Beving H, Albage A, Ivert T, Blomback M. Acetylsalicylic acid may protect the patient by increasing fibrin gel porosity. Is withdrawing of treatment harmful to the patient?. Eur Heart J 1996; 17: 1362-6.
26 Naito M, Hayashi T, Kuzuya M, Funaki C, Asai K, Kuzuya F. Thrombin stimulates the migration of fetal bovine aortic smooth muscle cells in culture. Igakunoayumi 1988; 144: 699-700. (In Japanese, Abstract in English)
27 Farrell DH, Al-Mondhiry A. Human fibroblast adhesion to fibrinogen. Biochemistry 1997; 36: 1123-8.
28 Gailit J, Clarke C, Newman D, Tonnesen MG, Mosesson MW, Clark RAF. Human fibroblasts bind directly to fibrinogen at RGD sites through integrin α_vβ₃ . Exp Cell Res 1997; 232: 118-26.
29 Katagiri Y, Hiroyama T, Akamatsu N, Suzuki H, Yamazaki H, Tanoue K. Involvement of α_vβ₃ integrin in mediating fibrin gel retraction. J Biol Chem 1995; 270: 1785-90.
30 Chen Y-P, O’Toole TE, Leong L, Liu B-Q, Diaz-Gonzalez F, Ginsberg MH. β₃ integrin-mediated fibrin clot retraction by nucleated cells: differing behavior of α_IIbβ₃ . Blood 1995; 86: 2606-15.
31 Hoshiga M, Alpers CE, Smith LL, Giachelli CM, Schwartz SM. α_vβ₃ integrin expression in normal and atherosclerotic artery. Circ Res 1995; 77: 1129-35.