A new evidence of a relationship between store-operated calcium entry and the regulation of procoagulant phosphatidylserine exposure at the surface of stimulated cells
Abstract number: P1076
Proulle* V., Capiod T., Galitzine* M., Martinez* C., Freyssinet* J. M., Le Deist F., Kerbiriou-Nabias* D.
*INSERM U143, France; INSERM U422, France; INSERM U429, France
The mechanism underlying phosphatidylserine (PS) externalization that promotes the formation of enzyme complexes of the coagulation cascade at the surface of activated blood cells remains largely unknown. The study of B lymphoblasts originating from a patient with Scott syndrome, a rare inherited hemorrhagic disease, led to better understand the intracellular calcium mobilization mechanisms involved in PS externalization. In Scott cells, defective PS externalization and shedding of membrane microparticles was associated with a defect of calcium influx across the plasma membrane induced by depletion of the endoplasmic reticulum calcium stores, a mechanism also known as store-operated calcium entry (SOCE) (Martinez et al. Biochemistry 1999; 38: 10092). In order to further illustrate the relationship between SOCE and PS externalization, B lymphoblasts originating from a patient with a primary immunodeficiency (PI) related to a deficiency of calcium influx in many cell types including T and B lymphocytes, platelets, leukocytes and fibroblasts (Le Deist et al. Blood 1995; 85: 1053) were studied. SOCE was monitored after the successive additions of 1 mM thapsigargin and 2 mM calcium on fura2-loaded cells using fluorescence spectroscopy. PS exposure was assessed by flow cytometry through annexin V-FITC binding before and after 10 min stimulation with 5 mM calcium ionophore A23187. The PS-dependent procoagulant phenotype was determined by prothrombinase assay. The A23187-induced increase in fluorescence intensity was 16.52 ± 1.24% in Scott and 11.9 ± 3.48% in PI lymphoblasts as compared to 42.4 ± 11.32% in control cells. Prothrombinase assay showed that A23187 stimulation induced an increase of PS externalization of 1.4-fold in Scott and 1.5-fold in PI lymphoblasts as compared to 1.9-fold in control cells. Release of microparticles was also markedly diminished after A23187 stimulation of Scott and of PI lymphoblasts with an increase in PS exposure of 1-fold and 1.4-fold, respectively, as compared to 4.2-fold for control cells. This study performed on B lymphoblasts of unrelated patients with different pathologies, is a new demonstration suggesting that SOCE is coupled with PS externalization and shedding of microparticles. The characterization of the as yet unknown genes involved in PS transmembrane movements in stimulated cells should therefore take into account the genes playing a role in calcium influx controlled by the depletion of intracellular calcium stores.
To cite this abstract use the following format:
Journal of Thrombosis and Haemostasis 2003; 1 Supplement 1 July: abstract number
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