Plasminogen activator inhibitor-1 (PAI-1) is the most important inhibitor of tissue-type plasminogen activator (t-PA) in plasma and plays a major role in the regulation of fibrinolysis. Plasma t-PA/PAI-1 complexes are cleared via a receptor-dependent mechanism in hepatocytes, while the fate of complexes formed in the extracellular matrix and in thrombi is less well understood. In this study, the degradation of t-PA/PAI-1 complexes by monocytes was examined. THP-1 monocytoid cells and freshly isolated human monocytes internalize and degrade [125I]t-PA/PAI-1 complexes at rates of 11.4 ± 5.9 (mean ± S.D.) and 44.6 ± 6.3 ng/106 cells/h, respectively. Degradation is blocked by receptor-associated protein (RAP), indicating a member of the low density lipoprotein (LDL) receptor family is involved in the uptake /degradation of t-PA/PAI-1 complexes by monocytes. Degradation of t-PA/PAI-1 complexes is also inhibited by chloroquine and by pepstatin A, suggesting that a lysosomal aspartyl protease is likely involved. SDS-PAGE and Western blotting demonstrated that the purifed. lysosomal aspartyl protease, cathepsin D, is capable of digesting t-PA (t 1 2 15 min), active PAI-1 (t 1 2 2 h), and t-PA/PAI-1 complex (t 1 2 30 min). Cathepsin D sequentially cleaves PAI-1 after hydrophobic amino acids, yielding lower molecular weight fragments. PAI-1 conformation influences the degradative efficiency of cathepsin D, with vitronectin-bound PAI-1 and latent PAM exhibiting resistance to proteolysis and > 10-fold prolongation in t 1 2. These data provide evidence that t-PA/PAI-1 complexes are internalized by human monocytes via a member of the low density lipoprotein (LDL) receptor family, and identifies cathepsin D-like aspartyl protease activity as largely responsible for the degradation of these complexes. Furthermore, vitronectin-bound PAM and latent PAI-1 are relatively resistant to degradation by cathepsin D, which may be of importance in complex physiological environments.
- Cathepsin D
- Plasminogen activator inhibitor-1
- Tissue-type plasminogen activator
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology