Stimulation of Eryptosis by Cadmium Ions

Abstract

Cadmium ions are known to trigger apoptosis. Erythrocytes may similarly undergo suicidal death or eryptosis, which is characterized by exposure of phosphatidylserine at the erythrocyte surface. As macrophages are equipped with phosphatidylserine receptors, they bind, engulf and degrade phosphatidylserine exposing cells. Cellular mechanisms known to trigger cell membrane phospholipid scrambling include increased cytosolic Ca²⁺ activity and activation of a sphingomyelinase with formation of ceramide. The present experiments were performed to explore whether cadmium ions (Cd²⁺) trigger phosphatidylserine exposure of erythrocytes and to possibly identify underlying mechanisms. Phosphatidylserine exposure was estimated from annexin V-binding as determined in fluorescence activated cell sorting (FACS) analysis. Exposure to Cd²⁺ (≧ 5.5 µM Cd²⁺) indeed significantly increased annexin V-binding. This effect was paralleled by erythrocyte shrinkage as apparent from the decrease of forward scatter in FACS analysis. According to Fluo3 fluorescence, Cd²⁺ increased the entry of Ca²⁺ into erythrocytes. According to antibody binding, Cd²⁺ did not stimulate the formation of ceramide. In the nominal absence of extracellular Ca²⁺ and in the presence of cation channel inhibitor amiloride the effects of Cd²⁺ on erythrocyte phosphatidylserine exposure and forward scatter were blunted. In conclusion, in human erythrocytes Cd²⁺ stimulates entry of Ca²⁺, which activates Ca²⁺-sensitive K⁺ channels leading to erythrocyte shrinkage and triggers Ca²⁺-sensitive erythrocyte membrane scrambling leading to phosphatidylserine exposure.