Dual effect of hydrogen peroxide on store-mediated calcium entry in human platelets

Abstract

Redox regulation is important for the modulation of cytosolic Ca 2+ concentration. Hence, we have investigated the effect of H 2O 2 on store-mediated Ca 2+ entry (SMCE). In fura-2-loaded human platelets treatment with H 2O 2 resulted in a concentration-dependent increase in Ca 2+ release from intracellular stores, while the effect on Ca 2+ entry was biphasic. In addition, 1 mM H 2O 2 reduced SMCE induced by agonists. The inhibitory effect of 1 mM H 2O 2 was prevented by inhibition of actin polymerization with cytochalasin D. Consistent with this, we found that 10 μM H 2O 2 and store depletion by treatment with thapsigargin plus ionomycin induced a similar temporal sequence of actin reorganization, while exposure to 1 mM H 2O 2 shifted the dynamics between polymerization and depolymerization in favor of the former. One millimolar H 2O 2-induced polymerization was reduced by treatment with methyl 2,5-dihydroxycinnamate and farnesylthioacetic acid, inhibitors of tyrosine kinases and Ras superfamily proteins, respectively. Finally, exposure to 1 mM H 2O 2 significantly increased store depletion-induced p60 src activation. We conclude that H 2O 2 exerted a biphasic effect on SMCE. The inhibitory role of high H 2O 2 concentrations is mediated by an abnormal actin reorganization pattern involving both Ras- and tyrosine kinases-dependent pathways.