Induction of Eryptosis in Red Blood Cells Using a Calcium Ionophore
Induction of Eryptosis in Red Blood Cells Using a Calcium Ionophore
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DOI：
10.3791/60659
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发表时间：
2020-01-01
影响因子：
1.2
通讯作者：
Farnoud, Amir M.
中科院分区：
文献类型：
--
作者：
Bigdelou, Parnian;Farnoud, Amir M.
红细胞凋亡，红细胞程序性细胞死亡，发生在许多血液学疾病和红细胞损伤期间。萎缩细胞的一个特征是细胞膜组成不对称的丧失，导致磷脂酰丝氨酸向膜外小叶移位。这一过程是由细胞内Ca2+浓度的增加触发的，Ca2+激活了超燃酶，一种促进磷脂在膜小叶之间双向运动的酶。鉴于紫癜在各种疾病条件下的重要性，已经努力在体外诱导紫癜。这些努力通常依赖于钙离子载体，离子霉素，以提高细胞内Ca2+浓度和诱导凋亡。然而，关于使用离子霉素诱导脓毒症的程序，文献中报道了许多差异。在这里，我们报告一步一步的方案离子霉素诱导的红细胞凋亡。重点介绍了离子载体浓度、培养时间、葡萄糖消耗等过程中的重要步骤，并给出了具有代表性的结果。该方案可用于在实验室中可重复地诱导脓毒症。
Eryptosis, erythrocyte programmed cell death, occurs in a number of hematological diseases and during injury to erythrocytes. A hallmark of eryptotic cells is the loss of compositional asymmetry of the cell membrane, leading to the translocation of phosphatidylserine to the membrane outer leaflet. This process is triggered by increased intracellular concentration of Ca2+, which activates scramblase, an enzyme that facilitates bidirectional movement of phospholipids between membrane leaflets. Given the importance of eryptosis in various diseased conditions, there have been efforts to induce eryptosis in vitro. Such efforts have generally relied on the calcium ionophore, ionomycin, to enhance intracellular Ca2+ concentration and induce eryptosis. However, many discrepancies have been reported in the literature regarding the procedure for inducing eryptosis using ionomycin. Herein, we report a step-by-step protocol for ionomycin-induced eryptosis in human erythrocytes. We focus on important steps in the procedure including the ionophore concentration, incubation time, and glucose depletion, and provide representative result. This protocol can be used to reproducibly induce eryptosis in the laboratory.