Vascular Smooth Muscle Cells Inhibit the Plasminogen Activators Secreted by Endothelial Cells

Summary

TPure cultures of bovine endothelial cells (EC) produce and secrete large amounts of plasminogen activators (PA). Cocultivation of EC with vascular smooth muscle cells (SMC) resulted in a significant decrease of PA activities secreted by the EC, whereas the cellular PA activities remained unaffected. Secreted PA activities were absent in the growth medium as long as the SMC to EC ratio was 2:1 or higher. The PA inhibitory activity of the SMC was rapid and cell-to-cell contact was not necessary.

The PA inhibitory activity was present in homogenates of SMC as well as in the medium conditioned by them but not in the extracellular matrix elaborated by these cells. Serum free medium conditioned by SMC neutralized both tissue type (t-PA) and urokinase like (u-PA) plasminogen activators. Gel electrophoretic analysis of SMC conditioned medium followed by reverse fibrin autography demonstrated PA inhibitory activities in the molecular weight (Mr) range of 50,000 to 52,000 similar to those present in media conditioned by bovine endothelial cells or fibroblasts. Regular fibrin zymography of SMC conditioned medium incubated with u-PA or t-PA revealed the presence of a component with a calculated approximate Mr of 45,000 to 50,000 which formed SDS resistant complexes with both types of PA.

These data demonstrate that vascular SMC produce and secrete (a) inhibitor(s) of PAs which may influence the fibrinolytic potential of EC.

Keywords

Endothelial cells - Vascular smooth muscle cells - Inhibitors of plasminogen activators

• References

• 1 Laug WE, Tokes ZA, Benedict WF, Sorgente N. Anchorage independent growth and plasminogen activator production by bovine endothelial cells. J Cell Biol 1980; 84: 281-293
• 2 Levin EG, Loskutoff DJ. Cultured bovine endothelial cells produce both urokinase and tissue-type plasminogen activators. J Cell Biol 1982; 94: 631-636
• 3 Laug WE. Glucocorticoids inhibit plasminogen activator production by endothelial cells. Thromb Haemostas 1983; 50: 888-892
• 4 Pandolfi M, Isaacson S, Nilsson IM. Low fibrinolytic activity in the walls of veins of patients with thrombosis. Acta Med Scand 1969; 1861-1865
• 5 Ross R, Glomset JA. The pathogenesis of atherosclerosis. N Engl J Med 1976; 295: 369-377
• 6 Laug WE. Secretion of plasminogen activators by cultural bovine endothelial cells: Partial purification, characterization and evidence for multiple forms. Thromb Haemostas 1981; 45: 219-224
• 7 Jones PA, DeClerck YA. Destruction of extracellular matrices containing glycoproteins, elastin, and collagen by metastatic human tumor cells. Cancer Res 1980; 40: 3222-3227
• 8 Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 1976; 72: 248-254
• 9 Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 1970; 227: 680-685
• 10 Granelli-Piperno A, Reich E. A study of proteases and proteaseinhibitor complexes in biological fluids. J Exp Med 1978; 143: 223-234
• 11 Loskutoff DJ, Van Mourik JA, Erickson LA, Lawrence D. Detection of an unusually stable fibrinolytic inhibitor produced by bovine endothelial cells. Proc Natl Acad Sci USA 1983; 80: 2956-2960
• 12 Baker JB, Low DA, Simmer RL, Cunningham DD. Protease-nexin: A cellular component that links thrombin and plasminogen-activator and mediates their binding to cells. Cell 1980; 21: 37-45
• 13 Deutsch DG, Mertz ET. Plasminogen: Purification from human plasma by affinity chromatography. Science 1970; 170: 1095-1096
• 14 Laki K. The polymerization of proteins: The action of thrombin on fibrinogen. Arch Biochem Biophys 1951; 32: 317-324
• 15 Mussoni L, Lawrence D, Loskutoff DJ. A direct, plasmin-independent assay for plasminogen activator. Thromb Res 1984; 34: 241-254
• 16 Loskutoff DJ. The fibrinolytic system of cultured endothelial cells: Deciphering the balance between plasminogen activation and inhibition. Haemostasis 1984; 14: 96 (a)
• 17 Scott RW, Baker JB. Purification of human protease nexin. J Biol Chem 1983; 258: 10439-10444
• 18 Eaton L, Baker JB. Evidence that a variety of cultured cells secrete protease nexin and produce a distinct cytoplasmic serine proteasebinding factor. J Cell Physiol 1983; 117: 175-182
• 19 Chmielewska J, Ranby M, Wiman B. Evidence for a rapid inhibitor to tissue plasminogen activator in plasma. Thromb Res 1983; 31: 417-435
• 20 Juhan-Vague I, Moerman B, DeCock F, Ailland MF, Collen D. Plasma levels of a specific inhibitor of tissue-type plasminogen activator (and urokinase) in normal and pathological conditions. Thromb Res 1984; 33: 523-530
• 21 Levin EG. Latent tissue plasminogen activator produced by human endothelial cells in culture: Evidence for an enzyme-inhibitor complex. Proc Natl Acad Sci USA 1983; 80: 6804-6808
• 22 Laug WE, DeClerck YA, Jones PA. Degradation of the subendothelial matrix by tumor cells. Cancer Res 1983; 43: 1827-1834