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Blood, Vol. 94 No. 4 (August 15), 1999:
pp. 1300-1312
Synthesis and Ultrastructural Localization of Protein C Inhibitor in
Human Platelets and Megakaryocytes
Maria J. Prendes,
Edith Bielek,
Margareta Zechmeister-Machhart,
Erika Vanyek-Zavadil,
Veronica A. Carroll,
Johannes Breuss,
Bernd
R. Binder, and
Margarethe Geiger
From the Department of Vascular Biology and Thrombosis Research, and
the Department of Histology and Embryology, University of Vienna,
Vienna, Austria.
The occurrence of protein C inhibitor (PCI) in human platelets and
megakaryocytes was analyzed. As judged from enzyme-linked immunosorbent
assays (ELISAs), PCI was present in platelets at a concentration of 160 ng/2 × 109 cells. Its specific activity was 5 times
higher than that of plasma PCI. Consistently, mainly the 57-kD form
(active PCI) and some high molecular weight
(Mr) forms, but no bands corresponding to
cleaved PCI, were detected when platelet lysates were
immunoprecipitated with monoclonal anti-PCI-IgG and analyzed by sodium
dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and
Western blotting. The localization of PCI in platelets was studied by
immunofluorescence histochemistry and immunotransmission electron
microscopy: PCI was detected in  granules, in the open
canalicular system, and on the plasma membrane. At these sites,
colocalization with plasminogen activator inhibitor-1 was seen. Studies
were performed to clarify whether platelet PCI is endogenously
synthesized or taken up from plasma. Internalization of
biotinylated-PCI was analyzed using platelets in suspension and
gold-labeled streptavidin for visualization of incorporated biotin.
Dose- and time-dependent uptake of PCI was found. PCI mRNA was detected
in platelets by reverse transcriptase-polymerase chain reaction
(RT-PCR) and Southern blotting, as well as in
megakaryocytes by in situ hybridization of human bone marrow
cryosections. We therefore conclude that platelets contain a
functionally active PCI pool that is derived from both endogenous
synthesis as well as internalization.
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