Nobel C S, Aronson J K, van den Dobbelsteen D J, Slater A F
Institute of Environmental Medicine, Division of Toxicology, Karolinska Institutet, Box 210, S-171 77 Stockholm, Sweden.
Apoptosis. 2000 Apr;5(2):153-63. doi: 10.1023/a:1009684713784.
To investigate the involvement of K+ efflux in apoptotic cell shrinkage, we monitored efflux of the K+ congener, 86Rb+, and cell volume during CD95-mediated apoptosis in Jurkat cells. An anti-CD95 antibody caused apoptosis associated with intracellular GSH depletion, a significant increase in 86Rb+ efflux, and a decrease in cell volume compared with control cells. Preincubating Jurkat cells with Val-Ala-Asp-chloromethylketone (VAD-cmk), an inhibitor of caspase proteases, prevented the observed 86Rb+ efflux and cell shrinkage induced by the anti-CD95 antibody. A wide range of inhibitors against most types of K+ channels could not inhibit CD95-mediated efflux of 86Rb+, however, the uptake of 86Rb+ by Jurkat cells was severely compromised when treated with anti-CD95 antibody. Uptake of 86Rb+ in Jurkat cells was sensitive to ouabain (a specific Na+/K(+)-ATPase inhibitor), demonstrating Na+/K(+)-ATPase dependent K+ uptake. Ouabain induced significant 86Rb+ efflux in untreated cells, as well as it seemed to compete with 86Rb+ efflux induced by the anti-CD95 antibody, supporting a role for Na+/K(+)-ATPase in the CD95-mediated 86Rb+ efflux. Ouabain treatment of Jurkat cells did not cause a reduction in cell volume, although together with the anti-CD95 antibody, ouabain potentiated CD95-mediated cell shrinkage. This suggests that the observed inhibition of Na+/K(+)-ATPase during apoptosis may also facilitate apoptotic cell shrinkage.
为了研究钾离子外流在凋亡细胞皱缩中的作用，我们监测了钾离子类似物86Rb+的外流以及Jurkat细胞在CD95介导的凋亡过程中的细胞体积。与对照细胞相比，抗CD95抗体诱导的凋亡与细胞内谷胱甘肽耗竭、86Rb+外流显著增加以及细胞体积减小有关。用半胱天冬酶蛋白酶抑制剂缬氨酰-丙氨酰-天冬氨酰-氯甲基酮（VAD-cmk）预孵育Jurkat细胞，可防止抗CD95抗体诱导的86Rb+外流和细胞皱缩。多种针对大多数类型钾通道的抑制剂均不能抑制CD95介导的86Rb+外流，然而，用抗CD95抗体处理后，Jurkat细胞对86Rb+的摄取严重受损。Jurkat细胞对86Rb+的摄取对哇巴因（一种特异性钠钾ATP酶抑制剂）敏感，表明存在钠钾ATP酶依赖性钾摄取。哇巴因在未处理的细胞中诱导显著的86Rb+外流，并且似乎与抗CD95抗体诱导的86Rb+外流竞争，这支持了钠钾ATP酶在CD95介导的86Rb+外流中的作用。用哇巴因处理Jurkat细胞不会导致细胞体积减小，尽管与抗CD95抗体一起使用时，哇巴因会增强CD95介导的细胞皱缩。这表明在凋亡过程中观察到的钠钾ATP酶抑制也可能促进凋亡细胞皱缩。
