TM5275 prolongs secreted tissue plasminogen activator retention and enhances fibrinolysis on vascular endothelial cells

Hideki Yasui, Yuko Suzuki, Hideto Sano, Takafumi Suda, Kingo Chida, Takashi Dan, Toshio Miyata, Tetsumei Urano

Research output: Contribution to journalArticlepeer-review

2 Citations (Scopus)

Abstract

Introduction Elevated plasminogen activator inhibitor-1 (PAI-1) reduces fibrinolytic potential in plasma, contributing to thrombotic disease. Thus, inhibiting PAI-1 activity is clinically desirable. We recently demonstrated that tissue plasminogen activator (tPA) remains on the surface of vascular endothelial cells (VECs) after secretion in a heavy-chain dependent manner, which is essential for high fibrinolytic activity on the surface of VECs, and that PAI-1 dissociates retained tPA from the cell surface as a result of high-molecular weight complex formation. Based on the model whereby amounts of tPA and its equilibrium with PAI-1 dynamically change after exocytosis, we examined how TM5275, a newly synthesized small molecule PAI-1 inhibitor, modulated tPA retention and VEC surface-derived fibrinolytic activity using microscopic techniques. Materials and methods The effects of TM5275 on the kinetics of the secretion and retention of green fluorescent protein (GFP)-tagged tPA (tPA-GFP) on VECs were analyzed using total internal reflection fluorescence microscopy. The effects of TM5275 on the generation of plasmin activity were evaluated by both plasminogen accumulation and fibrin clot lysis on tPA-GFP-expressing VECs using confocal laser scanning microscopy. Results TM5275 at concentrations of 20 and 100 μM significantly prolonged the retention of tPA-GFP on VECs by inhibiting tPA-GFP-PAI-1 high-molecular-weight complex formation. TM5275 enhanced the time-dependent accumulation of plasminogen as well as the dissolution of fibrin clots on and around the tPA-GFP-expressing cells. Conclusions The profibrinolytic effects of TM5275 were clearly demonstrated by the prolongation of tPA retention and enhancement of plasmin generation on the VEC surface as a result of PAI-1 inhibition.

Original languageEnglish
Pages (from-to)100-105
Number of pages6
JournalThrombosis Research
Volume132
Issue number1
DOIs
Publication statusPublished - 2013 Jul

Keywords

  • (PAI-1) inhibitor
  • (TIRF) microscopy
  • TM5275
  • fibrinolysis plasminogen activator inhibitor-1 (PAI-1)
  • plasminogen activator inhibitor-1
  • total internal reflection fluorescence

ASJC Scopus subject areas

  • Hematology

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