Time course of thrombin-induced increase in endothelial permeability: relationship to Ca2+i and inositol polyphosphates.
The temporal relationship between the alpha-thrombin-induced increase in transendothelial permeability and the alpha-thrombin-mediated changes in several key transmembrane signaling events was examined in confluent monolayers of bovine pulmonary artery endothelial cells (BPAEC). The time courses of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] generation, changes in cytosolic [Ca2+] ([Ca2+]i), and reorganization of cytoskeletal F-actin were determined to assess the relationship between these events and the onset of the alpha-thrombin-induced increase in endothelial permeability. alpha-Thrombin (10(-7) M) increased the transendothelial 125I-albumin clearance rate half-maximally by approximately 1 min and maximally by approximately 2 min (160% over control level). The increase in permeability occurred concomitantly with reorganization of F-actin cytoskeleton (i.e., loss of peripheral band and increased stress fiber density) and increased actin polymerization. Stimulation of fura-2-loaded BPAEC with 10(-7) M alpha-thrombin produced a typical biphasic rise in [Ca2+]i. The initial rapid increase in [Ca2+]i peaked by approximately 16 s after thrombin challenge and the [Ca2+]i response showed a slow decrease to half-maximal within 50 s. The alpha-thrombin-induced increase in permeability as well as the increase in [Ca2+]i were consistently preceded by increased Ins(1,4,5)P3 generation detectable within 10 s after thrombin challenge. These results indicate that alpha-thrombin triggers a cascade of events (i.e., Ins(1,4,5)P3 generation and the ensuing rise in [Ca2+]i), which may comprise the second messengers that mediate F-actin reorganization and the increase in endothelial permeability.
Lum
H
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Department of Physiology and Cell Biology, Albany Medical College, Union University, New York.
Aschner
J L
JL
Phillips
P G
PG
Fletcher
P W
PW
Malik
A B
AB
D000199:Actins / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002199:Capillary Permeability / Q000187:drug effects*; D002478:Cells, Cultured; D002850:Chromatography, Gel; D003599:Cytoskeleton / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D007295:Inositol Phosphates / Q000378:metabolism*; D012709:Serum Albumin / Q000493:pharmacokinetics; D013917:Thrombin / Q000494:pharmacology*; D013997:Time Factors; D014018:Tissue Distribution
D000199:Actins; D007295:Inositol Phosphates; D012709:Serum Albumin; D013917:Thrombin; D002118:Calcium