Rapid Procoagulant Phosphatidylserine Exposure Relies on High Cytosolic Calcium Rather Than on Mitochondrial Depolarization

Abstract

Relationships between intracellular Ca(2+) concentration ([Ca(2+)](cyt)) and apoptotic events, such as mitochondrial depolarization (DeltaPsim loss) and Bcl-2 and Bad phosphorylation, were analyzed in platelets and Jurkat cells in relation to rapid procoagulant phosphatidylserine (PS) exposure. Platelets were stimulated with A23187, thapsigargin (TG) and thrombin plus convulxin (Thr/Cvx), and Jurkat cells with ionomycin, in the presence or absence of cyclosporin A (CsA), a mitochondrial permeability transition pore inhibitor. DeltaPsim loss occurred when platelets were stimulated in Ca(2+) medium in conditions exposing PS, but also in EGTA medium. CsA inhibited PS exposure, [Ca(2+)](cyt) increase, and DeltaPsim loss in platelets stimulated with TG and Thr/Cvx, but had no inhibitory effect on A23187 stimulation. CsA reduced TG-induced Ca(2+) release from the endoplasmic reticulum and, consequently, external Ca(2+) influx. In ionomycin-stimulated Jurkat cells, rapid PS exposure was evidenced but not DeltaPsim loss, and CsA did not inhibit the process. The status of phosphorylated Bad and Bcl-2 in both cell types remained unchanged on stimulation. Whether DeltaPsim loss occurs or not, PS exposure is triggered by a high [Ca(2+)](cyt) increase. Data further demonstrate that CsA prevents membrane scrambling by inhibiting the high [Ca(2+)](cyt) increase, independently of its effect on mitochondrial permeability transition pore.