Complement proteins C5b-9 induce vesiculation of the endothelial plasma membrane and expose catalytic surface for assembly of the prothrombinase enzyme complex.
Assembly of the terminal complement proteins C5b-9 on human endothelial cells results in increased cytosolic calcium and nonlytic secretion of high molecular weight multimers of von Willebrand factor from intracellular storage granules. We now demonstrate that this C5b-9-induced secretory response is accompanied by vesiculation of membrane particles from the endothelial surface which express binding sites for factor Va and support prothrombinase activity. Exposure of factor Va binding sites after C5b-9 assembly was accompanied by greater than 2-fold increase in prothrombinase activity, which was not observed for cells exposed to C5b-8 (in the absence of C9). By contrast, only a 3-16% increase in prothrombinase activity was observed when these cells were maximally stimulated to secrete by either histamine, thrombin, or the Ca2+ ionophore A23187. Increased prothrombinase activity after C5b-9 was not accompanied by a change in thrombomodulin activity, and was unrelated to cell lysis, the complement-treated cells remaining greater than 99% viable. Endothelial prothrombinase activity was predominately associated with small membrane vesicles (less than 1 microns diameter) released from the cell monolayer. Analysis by fluorescence-gated flow cytometry revealed that these vesicles incorporate the C5b-9 proteins and express binding sites for factor Va. The capacity of the C5b-9 proteins to induce vesiculation of the endothelial plasma membrane and thereby expose catalytic surface for the prothrombinase enzyme complex may contribute to fibrin deposition associated with immune endothelial injury.
Hamilton
K K
KK
Department of Medicine, University of Oklahoma Health Sciences Center, Oklahoma City.
Hattori
R
R
Esmon
C T
CT
Sims
P J
PJ
D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D002462:Cell Membrane / Q000201:enzymology; D002478:Cells, Cultured; D015938:Complement Membrane Attack Complex / Q000502:physiology*; D004730:Endothelium, Vascular / Q000201:enzymology*; D005165:Factor V / Q000378:metabolism*; D015943:Factor Va / Q000378:metabolism; D005170:Factor X / Q000378:metabolism*; D015951:Factor Xa / Q000378:metabolism; D005260:Female; D005434:Flow Cytometry; D006632:Histamine / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D011247:Pregnancy; D013917:Thrombin / Q000096:biosynthesis; D014471:Umbilical Veins
D015938:Complement Membrane Attack Complex; C022475:prothrombinase complex; D000001:Calcimycin; D015943:Factor Va; D006632:Histamine; D005165:Factor V; D005170:Factor X; D013917:Thrombin; D015951:Factor Xa; D008274:Magnesium; D002118:Calcium