Serum-induced vascular smooth muscle cell elastolytic activity through tyrosine kinase intracellular signalling.
In previous studies, we related increased elastolytic activity in pulmonary arteries (PA) with endothelial injury to the later development of PA hypertension in rats. As the mechanism causing the increased PA elastase was unknown, we hypothesized that serum factors which are accessible to vascular smooth muscle cells (SMC) following endothelial injury stimulate their elastolytic activity. To test this, we developed an in vitro assay in which we added [3H]-elastin to cultured vascular SMC after 24 h serum starvation and monitored elastolysis following a further 24 h incubation with fetal bovine serum (FBS). We observed that serum induced increased elastolytic activity in both PA and aorta-derived SMC but not in endothelial cells or SMC with low basal levels of elastolytic activity. Maximum stimulation of SMC elastolytic activity occurred with a concentration as low as 1% FBS and despite elastase inhibitors in serum, suggesting that the activity is confined to the immediate pericellular region where enzyme concentration is high. Serum-stimulated elastolytic activity was not reproduced by growth factors or cytokines known to be associated with vascular disease or to induce release of elastases in other cells. The serum inducing elastolytic activity was heat and acid labile. It was associated with increased elastin adhesion to the 67 kD elastin binding protein on SMC surfaces and was prevented by tyrosine kinase inhibitors but not protein kinase C or A inhibitors. Our studies therefore suggest a mechanism whereby serum induction of SMC elastase requires signalling through the elastin binding protein and activation of tyrosine kinase.
Division of Cardiovascular Research, Research Institute, Hospital for Sick Children, Ontario, Canada.
J
J
Kobayashi
D
D
Wigle
T
T
Childs
L
L
Zhu
F W
FW
Keeley
M
M
Rabinovitch
D000818:Animals; D001798:Blood Proteins / Q000494:pharmacology*; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D004549:Elastin / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007375:Interleukin-1 / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000201:enzymology* / Q000378:metabolism; D010196:Pancreatic Elastase / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000032:analysis* / Q000502:physiology*; D012756:Sheep; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013552:Swine; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D001798:Blood Proteins; D007375:Interleukin-1; D010982:Platelet-Derived Growth Factor; D014409:Tumor Necrosis Factor-alpha; D004549:Elastin; D011505:Protein-Tyrosine Kinases; D010196:Pancreatic Elastase