Novel mechanism to enhance tPA-induced fibrinolysis: effect of limited proteolysis of PAI-1 by neutrophil elastase.
The effect of the proteolytic cleavage of plasminogen activator inhibitor type 1 (PAI-1) by human neutrophil elastase (HNE) on fibrinolysis was investigated. HNE cleaved active recombinant prokaryotic PAI-1 (rpPAI-1) resulting in the formation of low molecular weight forms of rpPAI-1 as previously reported. The latent form of rpPAI-1 was resistant to HNE. NH2-terminal sequence analysis indicated that the cleavage site was Val355-Ser356 (P4-P3). The fact that the strained loop of the latent form of PAI-1 is buried inside the molecule most likely accounts for its resistance to HNE. After the cleavage by HNE, active rpPAI-1 lost its specific activity toward plasminogen activators. The cleavage was both enzyme concentration and time dependent, and the almost complete inactivation of rpPAI-1 (2.9 microM) activity was obtained by a HNE (83 nM) treatment for 30 min at 37 degrees C. Vitroectin partially protected active rpPAI-1 from the HNE digestion. The effect of PAI-1 cleavage by HNE on tissue type PA (tPA) induced clot lysis was studied in a purified system. Clot lysis time without rpPAI-1 was 20.0 +/- 5.0 min, and was prolonged to 86.7 +/- 2.9 min by 68 nM of rpPAI-1. It was shortened when HNE (from 0.6 nM to 80 nM) was added and returned to the value obtained without rpPAI-1 when 80 nM of HNE was present (20.0 +/- 5.8 min). In the absence of PAI-1, however, HNE did not enhance clot lysis at all. The cleavage and inactivation of PAI-1 by HNE was shown to be a novel pathway to enhance fibrinolysis.
Urano
T
T
Department of Physiology, Hamamatsu University School of Medicine, Shizuoka, Japan.
Wu
K
K
Ihara
H
H
Takada
Y
Y
Takada
A
A
D000595:Amino Acid Sequence; D015180:Electrophoresis, Gel, Two-Dimensional; D005342:Fibrinolysis / Q000187:drug effects*; D006801:Humans; D019272:Leukocyte Elastase / Q000378:metabolism*; D008969:Molecular Sequence Data; D017395:Plasminogen Activator Inhibitor 1 / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D010959:Tissue Plasminogen Activator / Q000494:pharmacology*
D017395:Plasminogen Activator Inhibitor 1; D011994:Recombinant Proteins; D019272:Leukocyte Elastase; D010959:Tissue Plasminogen Activator