Sulindac sulfide--induced stimulation of eryptosis

Abstract

Background: Sulindac sulfide, a non-steroidal anti-inflammatory drug (NSAID), stimulates apoptosis of tumor cells and is thus effective against malignancy. In analogy to apoptosis of nucleated cells, erythrocytes may undergo eryptosis, an apoptosis-like suicidal erythrocyte death, characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine-exposure at the cell surface. Stimulators of eryptosis include increase of cytosolic Ca(2+)-activity ([Ca(2+)](i)) and ceramide formation. The present study explored, whether sulindac sulfide stimulates eryptosis.

Methods: [Ca(2+)](i) was estimated from Fluo-3 fluorescence, cell volume from forward scatter, phosphatidylserine-exposure from binding of fluorescent annexin-V, hemolysis from hemoglobin release, and ceramide abundance utilizing fluorescent antibodies.

Results: A 48 h exposure to sulindac sulfide (≤ 20 µM) was followed by significant increase of [Ca(2+)](i), enhanced ceramide abundance, decreased forward scatter and increased percentage of annexin-V-binding erythrocytes. Sulindac sulfide triggered slight but significant hemolysis. Removal of extracellular Ca(2+) significantly blunted, but did not abrogate the effect of sulindac sulfide (20 µM) on annexin-V-binding.

Conclusion: Sulindac sulfide stimulates the suicidal death of erythrocytes or eryptosis, an effect paralleled by Ca(2+)-entry, ceramide formation, cell shrinkage and phosphatidylserine-exposure.