Chow S C, Jondal M
Department of Immunology, Karolinska Institute, Stockholm, Sweden.
Immunology. 1990 May;70(1):106-10.
Using neomycin, which inhibits phosphoinositide breakdown, cytotoxicity mediated by natural killer (NK) cells was suppressed in a dose-dependent manner, with complete inhibition at 16 mM. Generation of inositol phosphates in effector cells after target cell binding was inhibited in the presence of neomycin. The formation of effector to target cell conjugates was not affected. Neomycin-induced inhibition of NK killing was abolished when TPA was added to the cytotoxic assays. This reconstitution was dependent upon extracellular Ca2+. When the intracellular free Ca2+ level in effector cells was reduced from 73 +/- 11 nM to 43 +/- 3 nM (n = 4) using the Ca2+ indicator dye, Quin 2, NK killing was markedly reduced. Inhibiting the enzyme diacylglycerol (DG) kinase in effector cells with 10 microM R59022 (DG kinase inhibitor) potentiates NK killing, suggesting an increase in protein kinase C (PKC) activity due to accumulation of DG. The PKC inhibitor, H-7, suppressed NK killing in a concentration-dependent manner. These results demonstrate that phosphoinositide metabolism is an early event and its derived second messengers play a central role in activating the lytic mechanism of NK cells.
使用抑制磷酸肌醇分解的新霉素，自然杀伤（NK）细胞介导的细胞毒性以剂量依赖方式受到抑制，在16 mM时完全抑制。在新霉素存在下，靶细胞结合后效应细胞中肌醇磷酸的生成受到抑制。效应细胞与靶细胞结合物的形成不受影响。当将佛波酯（TPA）添加到细胞毒性试验中时，新霉素诱导的NK杀伤抑制作用被消除。这种恢复依赖于细胞外钙离子（Ca2+）。当使用钙离子指示剂染料喹啉2将效应细胞内的游离钙离子水平从73±11 nM降低到43±3 nM（n = 4）时，NK杀伤明显降低。用10 microM R59022（二酰基甘油激酶抑制剂）抑制效应细胞中的二酰基甘油（DG）激酶可增强NK杀伤，表明由于DG积累导致蛋白激酶C（PKC）活性增加。PKC抑制剂H-7以浓度依赖方式抑制NK杀伤。这些结果表明，磷酸肌醇代谢是早期事件，其衍生的第二信使在激活NK细胞的裂解机制中起核心作用。
