Ciccone E, Viale O, Bottino C, Pende D, Migone N, Casorati G, Tambussi G, Moretta A, Moretta L
Istituto Nazionale per la Ricerca sul Cancro, University of Genova, Italy.
J Exp Med. 1988 Apr 1;167(4):1517-22. doi: 10.1084/jem.167.4.1517.
These experiments were designed to define the ability of human TCR-gamma+ cells to recognize allogeneic cells. TCR-gamma+-enriched populations were obtained by treating peripheral blood E-rosetting cells with anti-CD4 and anti-CD8 mAbs. The resulting populations were CD2+4-8- expressed variable proportions of CD3+ cells (40-90%), and did not react with the WT31 mAb, which is specific for a framework determinant of the alpha/beta heterodimer that serves as receptor for antigen on most human T lymphocytes. After mixed lymphocyte culture with irradiated allogeneic cells for 7 d and 3 additional days in rIL-2 (100 U/ml), cells underwent proliferation in three of five individuals tested. In addition, MLC-derived cells lysed 51Cr-labeled PHA-induced blasts derived from the allogeneic cells used as stimulator, but not allogeneic unrelated or autologous blast cells. No cytotoxicity against autologous or allogeneic target cells could be induced by culturing CD3+4-8-WT31- lymphocytes in MLC with irradiated autologous cells. Surface iodination of allogeneic MLC-activated CD3+4-8-WT31- cells followed by lysis in 1% digitonin and immunoprecipitation with anti-CD3 mAb indicated that the CD3-associated molecules consisted of a major 45-kD band and a minor band of 43 kD. Northern blot analysis showed that mRNA for the gamma chain was expressed at high levels, whereas mRNAs for alpha and beta chains were missing. These data support the notion that TCR-gamma rather than TCR-alpha/beta is expressed in allospecific CD3-4-8-WT31- cell populations. Clones were further derived from MLC-stimulated CD3+4-8-WT31- populations. All the seven clones studied in detail maintained the surface phenotype as well as the cytolytic pattern of the original MLC populations, thus only specific allogeneic PHA-induced blasts were lysed. NK-sensitive as well as NK-resistant tumor targets were variably susceptible to lysis; therefore, specific cytolytic activity against allogeneic cells was not necessarily linked to the expression of MHC-nonrestricted cytotoxicity against tumor cells.
这些实验旨在确定人类TCR-γ⁺细胞识别同种异体细胞的能力。通过用抗CD4和抗CD8单克隆抗体处理外周血E花环形成细胞来获得富含TCR-γ⁺的细胞群体。所得细胞群体为CD2⁺4⁻8⁻，表达不同比例的CD3⁺细胞（40% - 90%），且不与WT31单克隆抗体反应，WT31单克隆抗体对α/β异二聚体的框架决定簇具有特异性，该异二聚体是大多数人类T淋巴细胞上的抗原受体。在与经照射的同种异体细胞进行混合淋巴细胞培养7天，并在重组白细胞介素-2（100 U/ml）中再培养3天后，所检测的5名个体中有3名的细胞发生了增殖。此外，混合淋巴细胞培养衍生的细胞裂解了来自用作刺激细胞的同种异体细胞的51Cr标记的PHA诱导的母细胞，但不裂解同种异体无关或自体母细胞。通过在混合淋巴细胞培养中用经照射的自体细胞培养CD3⁺4⁻8⁻WT31⁻淋巴细胞，不能诱导对自体或同种异体靶细胞的细胞毒性。对同种异体混合淋巴细胞培养激活的CD3⁺4⁻8⁻WT31⁻细胞进行表面碘化，然后在1% digitonin中裂解并用抗CD3单克隆抗体进行免疫沉淀，结果表明与CD3相关的分子由一条主要的45-kD条带和一条次要的43 kD条带组成。Northern印迹分析显示γ链的mRNA高水平表达，而α链和β链的mRNA缺失。这些数据支持这样的观点，即在同种特异性CD3⁻4⁻8⁻WT31⁻细胞群体中表达的是TCR-γ而不是TCR-α/β。从混合淋巴细胞培养刺激的CD3⁺4⁻8⁻WT31⁻细胞群体中进一步获得克隆。详细研究的所有7个克隆都维持了原始混合淋巴细胞培养细胞群体的表面表型以及细胞溶解模式，因此仅裂解特定的同种异体PHA诱导的母细胞。NK敏感以及NK抗性的肿瘤靶细胞对裂解的敏感性各不相同；因此，针对同种异体细胞的特异性细胞溶解活性不一定与针对肿瘤细胞的MHC非限制性细胞毒性的表达相关。
