title: Cathepsin D-like aspartyl protease activity mediates the degradation of tissue-type plasminogen activator/plasminogen activator inhibitor-1 complexes in human monocytes.
abstract: 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/10(6) 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 purified lysosomal aspartyl protease, cathepsin D, is capable of digesting t-PA (t1/2 15 min), active PAI-1 (t1/2 2 h), and t-PA/PAI-1 complex (t1/2 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 PAI-1 exhibiting resistance to proteolysis and > 10-fold prolongation in t1/2. 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 PAI-1 and latent PAI-1 are relatively resistant to degradation by cathepsin D, which may be of importance in complex physiological environments.
authors.affiliation: Department of Medicine, Brigham and Women's Hospital, Boston, MA, USA.
authors.forename: D I
authors.initials: DI
authors.lastname: Simon
authors.forename: H
authors.initials: H
authors.lastname: Xu
authors.forename: D E
authors.initials: DE
authors.lastname: Vaughan
mesh_terms: D002402:Cathepsin D / Q000494:pharmacology*; D006801:Humans; D009000:Monocytes / Q000187:drug effects* / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D017395:Plasminogen Activator Inhibitor 1 / Q000378:metabolism*; D010959:Tissue Plasminogen Activator / Q000378:metabolism*
chemical_list: D010446:Peptide Fragments; D017395:Plasminogen Activator Inhibitor 1; D010959:Tissue Plasminogen Activator; D002402:Cathepsin D