Stimulation of Erythrocyte Cell Membrane Scrambling by Nystatin

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The antifungal ionophore nystatin dissipates the Na+ and K+ gradients across the cell membrane, leading to cellular gain of Na+ and cellular loss of K+. The increase of cellular Na+ concentration may result in Ca2+ accumulation in exchange for Na+. Increase of cytosolic Ca2+ activity ([Ca2+]i) and loss of cellular K+ foster apoptosis-like suicidal erythrocyte death or eryptosis, which is characterised by cell shrinkage and cell membrane scrambling leading to phosphatidylserine exposure at the erythrocyte surface. The present study explored whether nystatin stimulates eryptosis. Cell volume was estimated from forward scatter (FSC), phosphatidylserine exposure from annexin V binding and [Ca2+]i from Fluo3-fluorescence in flow cytometry. A 48-hr exposure to nystatin (15 μg/ml) was followed by a significant increase of [Ca2+]i, a significant increase of annexin V binding and a significant decrease of FSC. The annexin V binding after nystatin treatment was significantly blunted in the nominal absence of extracellular Ca2+. Partial replacement of extracellular Na+ with extracellular K+ blunted the nystatin-induced erythrocyte shrinkage but increased [Ca2+]i and annexin V binding. Nystatin triggers cell membrane scrambling, an effect at least partially due to entry of extracellular Ca2+.

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