Effect of protease treatment and carboxypeptidase treatment on plasminogen-binding capacity induced by neutrophils, plasmin, leukocyte elastase, or cathepsin G.
(A) Effect of protease treatment on the plasminogen-binding capacity of neutrophils. Freshly isolated neutrophils (4 × 107/mL) were incubated for 30 to 60 minutes at 37°C with plasmin (100 nM), leukocyte elastase (100 nM), or leukocyte cathepsin G (50 nM), with or without an excess of a specific inhibitor of the enzyme used. The concentrations of the enzymes were based on dose titration experiments, and the concentrations of the inhibitors produced complete inhibition in chromogenic substrate assays. At the end of the incubation, cells were washed and plasminogen binding was assessed. The results are means ± SEM of triplicates, and the experiments were repeated at least 3 times. For 3 such experiments, the increase in plasminogen binding to cells was statistically significant for each enzyme (P < .05). (B) Effects of carboxypeptidase treatment on the up-regulation of plasminogen binding induced by plasmin, leukocyte elastase, or cathepsin G. Neutrophils, treated as in panel A, were washed and then incubated for 30 minutes with 5 U/mL CpB. Specific plasminogen binding was then measured, and the data are expressed as the inhibition of the increase in plasminogen binding induced by each of the enzymes. The means ± SEM from 3 experiments are shown.
