Bouguerra Ghada, Bissinger Rosi, Abbès Salem, Lang Florian
Cell Physiol Biochem. 2015;37(4):1537-46. doi: 10.1159/000438521. Epub 2015 Oct 30.
BACKGROUND/AIMS: The aldose reductase inhibitor zopolrestat has been shown to either decrease or increase apoptosis, the suicidal death of nucleated cells. Erythrocytes may similarly enter suicidal death or eryptosis, which is characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Triggers of eryptosis include oxidative stress, Ca2+ entry with increase of cytosolic Ca2+ activity ([Ca2+]i), and ceramide formation. The present study explored, whether and how zopolrestat induces eryptosis.
Phosphatidylserine exposure at the cell surface was estimated from annexin V binding, cell volume from forward scatter, oxidative stress from DCFDA dependent fluorescence, [Ca2+]i from Fluo3-fluorescence, and ceramide abundance utilizing specific antibodies.
A 48 hours exposure of human erythrocytes to zopolrestat (≥ 150 µg/ml) significantly increased the percentage of annexin-V-binding cells, significantly decreased forward scatter (≥ 125 µg/ml), significantly increased Fluo3-fluorescence (200 µg/ml), significantly increased ceramide abundance (150 µg/ml), but did not significantly modify DCFDA fluorescence. The effect of zopolrestat on annexin-V-binding was significantly blunted, but not abolished by removal of extracellular Ca2+.
Exposure of human erythrocytes to zopolrestat triggers cell shrinkage and cell membrane scrambling, an effect in part due to Ca2+ entry and ceramide.
背景/目的：醛糖还原酶抑制剂唑泊司他已被证明可减少或增加细胞凋亡，即有核细胞的自杀性死亡。红细胞可能同样会进入自杀性死亡或红细胞凋亡，其特征是细胞收缩和细胞膜紊乱，磷脂酰丝氨酸易位至红细胞表面。红细胞凋亡的触发因素包括氧化应激、Ca2+内流导致胞质Ca2+活性（[Ca2+]i）增加以及神经酰胺形成。本研究探讨了唑泊司他是否以及如何诱导红细胞凋亡。
通过膜联蛋白V结合评估细胞表面磷脂酰丝氨酸的暴露情况，通过前向散射评估细胞体积，通过DCFDA依赖性荧光评估氧化应激，通过Fluo3荧光评估[Ca2+]i，并使用特异性抗体评估神经酰胺丰度。
将人红细胞暴露于唑泊司他（≥150μg/ml）48小时后，膜联蛋白V结合细胞的百分比显著增加，前向散射显著降低（≥125μg/ml），Fluo3荧光显著增加（200μg/ml），神经酰胺丰度显著增加（150μg/ml），但DCFDA荧光没有显著改变。去除细胞外Ca2+后，唑泊司他对膜联蛋白V结合的影响显著减弱，但并未消除。
将人红细胞暴露于唑泊司他会引发细胞收缩和细胞膜紊乱，部分原因是Ca2+内流和神经酰胺的作用。
