Rapid inactivation of the plasminogen-activator inhibitor upon secretion from cultured human endothelial cells

Abstract

In conditioned medium (CM) from cultured human endothelial cells, two forms of plasminogen-activator inhibitor (PA-inhibitor) can be demonstrated: a fast-acting active form and an immunologically related, inactive form. Evidence is presented that endothelial cells produce active PA-inhibitor which is rapidly inactivated upon secretion into the medium. This inactivation can, at least partly, be prevented by culturing cells with excess of tissue-type plasminogen activator (t-PA). This results in the formation of large amounts of t-PA-PA-inhibitor complex at the cost of accumulation of inactive PA-inhibitor. No complex was detectable when inactive PA-inhibitor preparations were incubated with t-PA either in the absence or in the presence of cells. Furthermore, in cell extracts, predominantly functionally active PA-inhibitor was present. PA-inhibitor derived from the t-PA-PA-inhibitor complex showed an Mr approx. 4000 lower by polyacrylamide-gel electrophoresis than that of the inactive form. The rapid inactivation seems to be confined to newly synthesized molecules, since PA-inhibitor molecules in CM are inactivated much more slowly (even with cells or cell homogenates) than necessary to explain the excessive production of inactivated PA-inhibitor by cells. It could not be prevented by inhibitors of oxidative processes, like butylated hydroxytoluene, dithiothreitol, superoxide dismutase and catalase.