Brahmi Z, Bray R A, Abrams S I
J Immunol. 1985 Dec;135(6):4108-13.
In this study, we examined the functional status of human natural killer (NK) cells after their direct interaction with the NK-sensitive tumor target cell (TC), K562. Human peripheral blood lymphocytes depleted of adherent cells were incubated for 4 hr with unlabeled K562 cells at an effector cell (EC) to TC ratio of 2:1. After incubation, the EC were separated from the TC via centrifugation over a single-step Percoll gradient. K562-treated and separated EC were subsequently shown to be unable to lyse fresh K562 TC when retested in the standard chromium-release assay. Kinetic studies revealed that greater than 90% inactivation of NK cell-mediated cytotoxicity (CMC) could be achieved within 2 hr. Inactivation of NK-CMC by K562 was not caused by a specific loss of NK cells, as detected by changes in the expression of two NK cell-associated markers, Leu-7 and Leu-11, or to alterations in EC viability and target binding cell capacity. Interestingly, NK inactivation also occurred in medium devoid of extracellular calcium, although parallel testing of NK-CMC in the same medium resulted in no chromium release. NK inactivation, however, was significantly prevented when the EC and TC were co-incubated at 4 degrees C, or in medium without magnesium. Additional studies revealed that inactivation of NK-CMC could be achieved with another NK-sensitive, but not with an NK-resistant TC. Overall, we demonstrated that NK cells rapidly lost their lytic potential after direct interaction with a sensitive TC, although the cells remained viable, expressed the same percentage of Leu-7 and Leu-11, and could still bind the TC; and NK inactivation occurred in the absence of extracellular calcium, but not when EC and TC were incubated in medium without magnesium. These latter results provide evidence for an early event in the activation of human NK cells that is binding dependent, temperature sensitive, and independent of extracellular calcium.
在本研究中，我们检测了人类自然杀伤（NK）细胞与NK敏感的肿瘤靶细胞（TC）K562直接相互作用后的功能状态。将去除贴壁细胞的人外周血淋巴细胞与未标记的K562细胞以效应细胞（EC）与TC 2:1的比例孵育4小时。孵育后，通过在单步Percoll梯度上离心将EC与TC分离。随后，在标准的铬释放试验中重新检测时，经K562处理并分离的EC无法裂解新鲜的K562 TC。动力学研究表明，在2小时内可实现超过90%的NK细胞介导的细胞毒性（CMC）失活。K562导致的NK-CMC失活并非由NK细胞的特异性损失引起，这可通过两种NK细胞相关标志物Leu-7和Leu-11表达的变化检测到，也不是由EC活力和靶细胞结合能力的改变引起。有趣的是，在无细胞外钙的培养基中也发生了NK失活，尽管在相同培养基中对NK-CMC进行平行检测未导致铬释放。然而，当EC和TC在4℃下共孵育或在无镁培养基中时，NK失活被显著阻止。进一步的研究表明，另一种NK敏感但非NK抗性的TC也能实现NK-CMC的失活。总体而言，我们证明NK细胞在与敏感的TC直接相互作用后迅速丧失其裂解潜能，尽管细胞仍存活，Leu-7和Leu-11的表达百分比相同，且仍能结合TC；并且NK失活发生在无细胞外钙的情况下，但当EC和TC在无镁培养基中孵育时则不会发生。后一结果为人类NK细胞激活过程中的一个早期事件提供了证据，该事件依赖于结合、对温度敏感且与细胞外钙无关。
