Single-Chain Urokinase Alone or Complexed to Its Receptor in Tetracycline-induced Pleuritis in Rabbits

Abstract

Intrapleural loculation can increase morbidity in hemothoraces or parapneumonic effusions. Intrapleural fibrin precedes visceral-parietal pleural adhesions. We speculated that single-chain urokinase plasminogen activator alone or bound to its receptor could prevent these adhesions by their relative resistance to local inhibition by plasminogen activator inhibitors. We found that recombinant human single-chain urokinase-bound rabbit pleural mesothelial cells or lung fibroblasts with kinetics similar to that reported for human cells (kD of approximately 5 nM). The receptor-bound fibrinolysin maintained in vitro fibrinolytic activity in the presence of pleural fluids from rabbits with tetracycline-induced pleural injury over 24 hours. In rabbits given intrapleural single-chain urokinase 24 and 48 hours after intrapleural tetracycline (n = 10 animals), adhesions were prevented, whereas the receptor-complexed form (n = 12) attenuated adhesions versus vehicle/tetracycline-treated rabbits (n = 22, p <or= 0.005 in both cases). There were more adhesions in the complex than the single-chain urokinase group (p = 0.02). Residual antigenic but not functional evidence of the interventional agents remained in pleural fluids at 72 hours after tetracycline. No local or systemic bleeding occurred because of either interventional agent. The data demonstrate that single-chain urokinase inhibits, whereas lysin-receptor complexes attenuate, adhesion formation in tetracycline-induced pleural injury in rabbits.

Thrombin–thrombomodulin inhibits prourokinase-mediated pleural mesothelial cell-dependent fibrinolysis

“…Pericellular proteolysis, including degradation of provisional fibrin matrix by serine-and metallo-proteinases, has been implicated as a common feature of the pathogenesis of several diseases, including lung and pleural injury [10,[14][15][16]. During the last decade, evidence for the involvement of the uPA system in pleural injury and repair has steadily increased [2,16,17].…”

“…Thrombin-cleaved uPA has previously been detected in plasma of patients with sepsis [18] and in synovial fluids of patients with rheumatoid arthritis [18], leading us to infer that thrombin-TM-mediated cleavage of scuPA contributes to intrapleural regulation of fibrinolysis during evolving pleurodesis or other pleural diseases. We also inferred that this mechanism may be involved in cleavage of exogenous scuPA, which prevents intrapleural loculations and lung trapping induced by tetracycline and is being evaluated as a candidate for future clinical use [16].…”

“…During the last decade, evidence for the involvement of the uPA system in pleural injury and repair has steadily increased [2,16,17]. Supporting this view, intrapleural administration of uPA or heparin reduces pleural adhesions and fibrosis [17].…”

“…Thrombin-cleaved uPA has previously been detected in plasma of patients with sepsis [18] and in synovial fluids of patients with rheumatoid arthritis [18], leading us to infer that thrombin-TM-mediated cleavage of scuPA contributes to intrapleural regulation of fibrinolysis during evolving pleurodesis or other pleural diseases. We also inferred that this mechanism may be involved in cleavage of exogenous scuPA, which prevents intrapleural loculations and lung trapping induced by tetracycline and is being evaluated as a candidate for future clinical use [16].To address these gaps in current knowledge of the regulation of fibrinolysis in the pleural space, we established a new model of cell-dependent fibrinolysis to determine how thrombin regulates the fibrinolytic capacity of pleural mesothelial cells in vitro. We provide new evidence that thrombin-dependent cleavage of scuPA contributes to the regulation of local fibrinolysis in diverse forms of pleural disease and in the processing of exogenous scuPA administered in a therapeutic context.…”

“…Naïve MeT5A cells, which expressed relatively smaller amounts of uPA, were used as comparator controls. Primary rabbit pleural mesothelial cells (RPMC) were isolated and cultured as previously described [16]. …”