Generation of cytotoxic T lymphocytes in vitro. IV. Functional activation of memory cells in the absence of DNA synthesis.
Generation of cytotoxic T lymphocytes in vitro. IV. Functional activation of memory cells in the absence of DNA synthesis.
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DOI：
10.1084/jem.142.3.622
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发表时间：
1975-09-01
影响因子：
15.3
通讯作者：
Brunner, K T
中科院分区：
文献类型：
--
作者：
MacDonald, H R;Sordat, B;Cerottini, J C;Brunner, K T
第14天的混合白细胞培养物（MLC）细胞再暴露于原始刺激性同种异体抗原（二次应答）先前已显示导致显著增殖和在接下来的4天内快速再现高水平的溶细胞性T淋巴细胞（CTL）活性。此外，有证据表明，在第14天MLC群体中的CTL前体细胞，虽然它们来源于在初次应答高峰（第4天）时较大的细胞，但它们本身是小淋巴细胞，在再刺激后发展成大CTL。在这项研究中，DNA合成的抑制阿糖胞苷（ARA-C）被用来调查是否可以从二次反应过程中的增殖分离的CTL形成。发现在再刺激后的前24小时内（a）CTL活性增加6-20倍，（B）60-70%的小T淋巴细胞变成中等至大尺寸的细胞，和（c）这两个事件都不依赖于DNA合成。通过在ARA-C存在或不存在下刺激第14天MLC细胞24小时之前和之后，在单位重力下通过速度沉降进行两次连续的细胞分离，获得了小CTL前体细胞发育成大CTL的直接证据，而与DNA合成无关。ARA-C的存在时间长于24小时抑制任何进一步增加的CTL活性，相反，平行增加的裂解活性和细胞数量从第1天到第4天在控制再刺激培养。与对照再刺激培养物中90%的中型和大型淋巴样细胞在24小时内进行DNA合成的发现一起，这些结果因此表明，在二次MLC反应期间，最初存在导致CTL形成的分化步骤，尽管其可以明显地与DNA合成分离，但在正常条件下，随后是这些效应细胞的增殖。
Re-exposure of day 14 mixed leukocyte culture (MLC) cells to the original stimulating alloantigens (secondary response) has previously been shown to result in significant proliferation and in rapid reappearance of high levels of cytolytic T-lymphocyte (CTL) activity within the next 4 days. Moreover, evidence has been presented that CTL precursor cells in day 14 MLC populations, while they derived from cells were large at peak of the primary response (day 4) were themselves small lymphocytes which developed into large CTL after restimulation. In this study, inhibition of DNA synthesis by cytosine arabinoside (ARA-C) was used to investigate whether CTL formation could be dissociated from proliferation during the secondary response. It was found that within the first 24 h after restimulation (a) CTL activity increased 6-to-20-fold, (b) 60-70% of the small T lymphocytes became medium- to large-sized cells, and (c) both events were independent of DNA synthesis. By using two successive cell separations by velocity sedimentation at unit gravity, before and after stimulation of day 14 MLC cells for 24 h in the presence or absence of ARA-C, direct evidence was obtained that small CTL precursor cells developed into large CTL, irrespective of DNA synthesis. The presence of ARA-C for periods longer than 24 h inhibited any further increase in CTL activity, in contrast to a parallel increase in lytic activity and cell number from day 1 to day 4 in control restimulated cultures. Taken together with the finding that 90% of the medium- and large-sized lymphoid cells in control restimulated cultures underwent DNA synthesis within 24 h, these results thus suggest that during a secondary MLC response there is initially a differentiation step leading to the formation of CTL which, although it can be clearly dissociated from DNA synthesis, is under normal conditions followed by proliferation of these effector cells.