Calcium plays a key role in the effects induced by a snake venom Lys49 phospholipase A2 homologue on a lymphoblastoid cell line

A catalytically-inactive Lys49 phospholipase A2 homologue from the venom of the snake Bothrops asper induces diverse effects (necrosis, apoptosis and proliferation) in a lymphoblastoid cell line, depending on the toxin concentration. The increments in cytosolic Ca2+ levels induced by this toxin in this cell line were assessed. At high toxin concentration (100 μg/mL) the toxin induces drastic disruption of the plasma membrane, associated with a prominent Ca2+ influx and necrosis. Previous incubation of the cells with the chelating agent EGTA or with ruthenium red, an inhibitor of the uniporter mitochondrial Ca2+ transport, greatly reduced necrosis. At a toxin concentration of 12.5 μg/mL, apoptosis is the predominant response, being associated with lower increments in cytosolic Ca2+. This effect was inhibited by preincubation with ruthenium red and the cytosolic Ca2+ chelator BAPTA-AM. The proliferative response, which occurs at a low toxin concentration (0.5 μg/mL), is associated with a small and oscillatory increment in cytosolic Ca2+. It was inhibited by EGTA, ruthenium red and BAPTA-AM, by inhibitors of the endoplasmic reticulum Ca2+-ATPase (SERCA) and by blockade of the ryanodine receptor. It is concluded that necrosis and apoptosis induced by this toxin are associated with increments in cytosolic Ca2+ levels following plasma membrane perturbation, together with the involvement of mitochondria. The cellular proliferative response depends on a limited Ca2+ influx through the plasma membrane, being associated with a concerted functional unit constituted by SERCA, the ryanodine receptor and mitochondria, which regulate the observed oscillations in cytosolic Ca2+ concentration.