Ferrini S, Miescher S, Zocchi M R, von Fliedner V, Moretta A
J Immunol. 1987 Feb 15;138(4):1297-302.
In these studies we investigated the phenotypic and functional characteristics of human rIL 2-activated killer cells (LAK). By FACS sorting we separated PBL into Leu-11- and Leu-11+ cell fractions and cultured them for 4 days in 100 U/ml rIL 2. Under these culture conditions, cells of the Leu-11+ fraction acquired a stronger LAK activity against fresh autologous or allogeneic melanoma cells as compared with Leu-11- cells or unfractionated PBL. To better characterize the cells responsible for this cytolytic activity, we directly cloned Leu-11+ and Leu-11- FACS-sorted cells in the presence of 1% PHA, irradiated spleen feeder cells, and rIL 2. From 6 to 10% of the Leu-11+ cells and from 42 to 66% of the Leu-11- cells plated gave rise to clonal progenies that were tested simultaneously for cytolytic activity against fresh melanoma cells and NK-sensitive K562 target cells in a 4-hr 51Cr-release assay. Most of the Leu-11+ microcultures lysed fresh melanoma target cells (35 out of 38 and 26 out of 34 in two separate experiments), whereas only a few clones derived from the Leu-11- cell fraction had this capability (four out of 45 and one out of 41). All the clones lysing fresh melanoma cells also efficiently killed K562 target cells, whereas other clones lysing only K562 could be found among Leu-11+ and Leu-11- clones. Nine clones expressing LAK activity were tested for their reactivity against a panel of different tumor target cells. All clones were able to lyse a broad panel of target cells including NK-sensitive and NK-resistant cultured or noncultured human tumor target cells, as well as mouse tumor cell lines. Surface marker analysis of 14 clones displaying LAK activity, all derived from Leu-11+ cells, showed that they were all T3 (CD3)-, whereas 10 out of 14 expressed the T11 (CD2) antigen and only four were weakly stained by an anti-T8 (CD8) mAb. All 14 clones expressed the T40 (CD7) T cell marker and DR and LFA-1 antigens. Cytolysis inhibition experiments performed on a rIL 2-activated Leu-11+ population and on two LAK cell clones, both expressing T11 antigen, showed that anti-LFA-1 but not anti-T11 mAb could inhibit cytolysis of freshly derived tumor target cells.
在这些研究中，我们调查了人重组白细胞介素2激活的杀伤细胞（LAK）的表型和功能特性。通过荧光激活细胞分选术（FACS），我们将外周血淋巴细胞（PBL）分离为Leu-11-和Leu-11+细胞组分，并在100 U/ml重组白细胞介素2中培养4天。在这些培养条件下，与Leu-11-细胞或未分离的PBL相比，Leu-11+组分的细胞对新鲜的自体或同种异体黑色素瘤细胞具有更强的LAK活性。为了更好地表征负责这种细胞溶解活性的细胞，我们在存在1%植物血凝素（PHA）、经辐照的脾饲养细胞和重组白细胞介素2的情况下，直接克隆FACS分选的Leu-11+和Leu-11-细胞。接种的Leu-11+细胞中有6%至10%以及Leu-11-细胞中有42%至66%产生了克隆后代，这些克隆后代在4小时51铬释放试验中同时针对新鲜黑色素瘤细胞和自然杀伤（NK）敏感的K562靶细胞进行细胞溶解活性测试。大多数Leu-11+微培养物裂解新鲜黑色素瘤靶细胞（在两个独立实验中分别为38个中的35个和34个中的26个），而来自Leu-11-细胞组分的克隆中只有少数具有这种能力（45个中的4个和41个中的1个）。所有裂解新鲜黑色素瘤细胞的克隆也能有效杀伤K562靶细胞，而在Leu-11+和Leu-11-克隆中可以找到其他仅裂解K562的克隆。对9个具有LAK活性的克隆进行了针对一组不同肿瘤靶细胞的反应性测试。所有克隆都能够裂解广泛的靶细胞，包括NK敏感和NK抗性的培养或未培养的人肿瘤靶细胞以及小鼠肿瘤细胞系。对14个均来自Leu-11+细胞且具有LAK活性的克隆进行表面标志物分析，结果显示它们均为T3（CD3）阴性，而14个中有10个表达T11（CD2）抗原，只有4个被抗T8（CD8）单克隆抗体弱阳性染色。所有14个克隆均表达T40（CD7）T细胞标志物以及DR和淋巴细胞功能相关抗原-1（LFA-1）抗原。对一个重组白细胞介素2激活的Leu-11+群体以及两个均表达T11抗原的LAK细胞克隆进行的细胞溶解抑制实验表明，抗LFA-1单克隆抗体而非抗T11单克隆抗体能够抑制对新鲜来源肿瘤靶细胞的细胞溶解。
