Radosević K, van Leeuwen A M, Segers-Nolten I M, Figdor C G, de Grooth B G, Greve J
Department of Applied Physics, University of Twente, Enschede, The Netherlands.
Cytometry. 1995 Aug 1;20(4):273-80. doi: 10.1002/cyto.990200402.
The cytotoxic interaction between cloned human Natural Killer (NK) cells and K562 target cells was studied using confocal laser scanning microscopy (CLSM) and conventional fluorescence microscopy. We observed, using fixed as well as living cells, the occurrence of (pseudo) emperipolesis during the interaction. About 30% of conjugated NK cells penetrated, partly or completely, into the target cells (in-conjugation). Virtually all in-conjugated target cells exhibited polymerized actin. Killer cells of in-conjugates were frequently seen approaching the target cell nucleus or aligning along it. If the cytotoxic process was inhibited by the absence of calcium neither actin polymerization nor in-conjugation were observed. A kinetic study showed that in-conjugation starts somewhat later than actin polymerization but still within a few minutes after addition of calcium to conjugates previously formed in the absence of calcium. The presence of cytochalasin D (an inhibitor of actin polymerization) completely inhibited in-conjugation and partly reduced the cytotoxic activity. Zinc ions (endonuclease inhibition) inhibited in-conjugation and decreased the total number of target cells with polymerized actin in a concentration dependent manner. Cytotoxic activity was also reduced but not as efficiently as in-conjugation. Our study demonstrates that in-conjugation represents a significant fraction of the cytotoxic interaction. The results indicate that it may be a consequence of an actin polymerization and endonuclease activity dependent part of a cytotoxic mechanism.
利用共聚焦激光扫描显微镜（CLSM）和传统荧光显微镜研究了克隆的人类自然杀伤（NK）细胞与K562靶细胞之间的细胞毒性相互作用。我们使用固定细胞和活细胞观察到，在相互作用过程中出现了（假）核周运动。约30%的结合NK细胞部分或完全穿透靶细胞（结合内）。几乎所有结合内的靶细胞都表现出聚合肌动蛋白。结合内的杀伤细胞经常靠近靶细胞核或沿其排列。如果由于缺乏钙而抑制细胞毒性过程，则既未观察到肌动蛋白聚合，也未观察到结合内现象。动力学研究表明，结合内现象比肌动蛋白聚合开始得稍晚，但仍在向先前在无钙条件下形成的结合物中添加钙后的几分钟内开始。细胞松弛素D（肌动蛋白聚合抑制剂）的存在完全抑制了结合内现象，并部分降低了细胞毒性活性。锌离子（核酸内切酶抑制）抑制结合内现象，并以浓度依赖的方式减少具有聚合肌动蛋白的靶细胞总数。细胞毒性活性也降低了，但不如结合内现象有效。我们的研究表明，结合内现象是细胞毒性相互作用的一个重要部分。结果表明，它可能是细胞毒性机制中依赖肌动蛋白聚合和核酸内切酶活性部分的结果。
