Bradykinin antagonizes the effects of alpha-thrombin.
alpha-Thrombin (AT) and bradykinin (BK) are endogenous mediators that are released during an inflammatory response, and could have a synergistic effect on endothelial permeability. Human umbilical vein endothelial cells (HUVEC) were grown on Transwell membranes and then tested for alterations in permeability to fluorescein isothiocyanate-labeled human serum albumin. Addition of 1 microM AT produced a significant increase in the permeability coefficient at 30 minutes from control levels of 1.59 x 10(-6) cm/sec to 4.92 x 10(-6) cm/sec. BK (1 microM) produced a similar increase to 4.46 x 10(-6) cm/sec. For both compounds, permeability remained elevated for 90 minutes. Pre-treatment of the HUVEC with the bradykinin receptor antagonist, Na-adamantaneacetyl-bradykinin (NA-BK) (1 microM), prior to addition of AT, reduced the AT permeability coefficient to 2.69 x 10(-6) cm/sec. Addition of NA-BK (1 microM) for 5 minutes, then BK (1 microM) for 5 minutes, inhibited the effect of BK and of AT (1 microM on permeability, decreasing the permeability coefficient of the endothelial monolayer to control levels (1.62 x 10(-6) cm/sec). AT (1 microM) increased HUVEC intracellular calcium mobilization, as monitored by FURA-2, to 245 nM from control (70 nM), however, pre-treatment with either BK or the bradykinin receptor antagonist decreased the AT induced intracellular calcium mobilization compared to AT alone. Pre-treatment of the HUVEC with bradykinin (1 microM) for 2 minutes also inhibited the effects of alpha-thrombin (1 microM) on f-actin distribution examined by BODIPY-phallodin staining and increased the clotting times for an alpha-thrombin dependent fibrinogen to fibrin clotting assay. However, incubation of bradykinin (1 microM) with alpha-thrombin (1 microM) for either 10 minutes or 100 minutes produced no detectable hydrolysis products. These data strongly suggest that the inflammatory mediators alpha-thrombin and bradykinin when released together, rather than being synergistic, are antagonistic.
Ehringer
W D
WD
Center for Applied Microcirculatory Research, University of Louisville School of Medicine, Kentucky 40292, USA.
Edwards
M J
MJ
Gray
R D
RD
Miller
F N
FN
D000199:Actins / Q000378:metabolism; D000218:Adamantane / Q000031:analogs & derivatives / Q000494:pharmacology; D000595:Amino Acid Sequence; D001777:Blood Coagulation / Q000187:drug effects; D001920:Bradykinin / Q000031:analogs & derivatives / Q000737:chemistry / Q000494:pharmacology*; D065168:Bradykinin Receptor Antagonists; D002118:Calcium / Q000378:metabolism; D002199:Capillary Permeability / Q000187:drug effects; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism; D004730:Endothelium, Vascular / Q000187:drug effects / Q000502:physiology; D016650:Fluorescein-5-isothiocyanate; D006801:Humans; D018836:Inflammation Mediators / Q000037:antagonists & inhibitors / Q000494:pharmacology; D012709:Serum Albumin; D013917:Thrombin / Q000037:antagonists & inhibitors* / Q000494:pharmacology
D000199:Actins; D065168:Bradykinin Receptor Antagonists; D018836:Inflammation Mediators; D012709:Serum Albumin; C074544:bradykinin, 1-adamantanecarboxylic acid-Arg(0)-Hyp(3)-Thi(5,8)-Phe(7)-; C046722:bradykinin, des-Arg(1)-; D013917:Thrombin; D016650:Fluorescein-5-isothiocyanate; D000218:Adamantane; D001920:Bradykinin; D002118:Calcium