Exocytic responses of single leukaemic human cytotoxic T lymphocytes stimulated by agents that bypass the T cell receptor
Exocytic responses of single leukaemic human cytotoxic T lymphocytes stimulated by agents that bypass the T cell receptor
复制标题
DOI：
10.1113/jphysiol.2005.089565
复制
发表时间：
2005-09-15
影响因子：
5.5
通讯作者：
Zweifach, A
中科院分区：
文献类型：
--
作者：
Pores-Fernando, AT;Bauer, RA;Zweifach, A
细胞毒性T淋巴细胞通过分泌包括穿孔素和颗粒酶在内的溶解剂杀死目标。近年来，新的方法已经发展监测细胞毒性T淋巴细胞脱颗粒。这些包括检测细胞表面溶酶体相关膜蛋白的外观，以及监测细胞穿孔素含量的减少。我们将这些方法与显微镜和流式细胞术相结合，首次分析了单个细胞毒性T细胞如何脱颗粒。我们使用TALL-104人白血病细胞毒性T细胞作为模型系统，并用thapsigargin和PMA刺激它们，这两种可溶性药物模拟T细胞受体交联激活的两种主要信号通路。我们的结果表明，基本上每个TALL-104细胞对最大刺激的反应是释放大约一半的溶解颗粒补体。这反映了一半细胞颗粒内容物的完全释放，而不是所有颗粒内容物的部分释放。亚最大刺激减少了反应细胞的比例和单个细胞反应的幅度。我们发现单个细胞对最大刺激的反应具有可变的潜伏期，并提供证据表明，一旦开始，脱颗粒是一个缓慢的过程，需要数十分钟。
Cytotoxic T lymphocytes kill targets via secretion of lytic agents including perforin and granzymes. Recently, new methods have been developed to monitor cytotoxic T lymphocyte degranulation. These include detecting the appearance of lysosome-associated membrane protein on the cell's surface, and monitoring decreases in cellular perforin content. We have combined these methods with microscopy and flow cytometry to provide the first analysis of how single cytotoxic T cells degranulate. We used TALL-104 human leukaemic cytotoxic T cells as a model system, and stimulated them with thapsigargin and PMA, soluble agents that mimic the two major signalling pathways activated by T cell receptor cross-linking. Our results indicate that essentially every TALL-104 cell responds to maximal stimulation by releasing about half of its lytic granule complement. This reflects complete release of the contents of half the cell's granules, rather than partial release of the contents of all of the granules. Sub-maximal stimulation reduces both the fraction of cells that respond and the magnitude of single cell responses. We find that individual cells respond to maximal stimulation with a variable latency, and provide evidence that, once it starts, degranulation is a slow process taking tens of minutes.