Preffer F I, Colvin R B, Leary C P, Boyle L A, Tuazon T V, Lazarovits A I, Cosimi A B, Kurnick J T
J Immunol. 1986 Nov 1;137(9):2823-30.
The phenotype of T lymphocyte subsets present in renal biopsies showing acute cellular allograft rejection in six patients on cyclosporine have been characterized in situ by immunoperoxidase staining, and after expansion in vitro in interleukin 2 (IL-2) by two-color flow cytometry, sorting, and functional analysis. After 8 to 42 days in organ culture, both Leu-3+ (CD4) and Leu-2+ (CD8) subsets were found in each culture, in a ratio that varied from 0.2 to 5.0, which was not significantly different than the results of in situ immunoperoxidase staining of the uncultured biopsy. The cultured cells were almost all Leu-4+ (CD3) T cells (89% +/- 4), which expressed the activation markers DR (82% +/- 6) and the IL 2 (CD25) receptor (15% +/- 4). The Leu-3+ cells were largely Leu-8- (90% +/- 6), whereas a minority of the Leu-2+ cells were Leu-15+ (CD11) (26% +/- 4). Only a small fraction of the Leu-2+ cells stained for Leu-7 (8% +/- 6). Functional analysis of FACS-purified Leu-2-3+ and Leu-2+3- populations indicated that both subsets proliferated in response to graft donor antigens in a mixed lymphocyte reaction (MLR) and produced IL 2. Only the Leu-2+3- population demonstrated donor-specific cytotoxic activity. A minor subpopulation in each culture were both Leu-3+ and Leu-2+ (2.0%). Leu-2+3+ cells from one biopsy were purified to homogeneity (99.8%), and were found to express the T cell antigen receptor complex Ti/CD3 (WT-31+, Leu-4+), but not the common thymocyte antigen CD1 (OKT6). The Leu-2+3+ cells neither responded in the MLR, nor showed any cytotoxic capacity. The Leu-2+3+ cells were capable of IL 2 but not interferon-gamma production. None of the purified cultures demonstrated NK activity. A subset of the purified Leu-2+3+ cells lost Leu-2+ during 1 to 3 wk in culture, and became Leu-2-3+. These studies provide evidence that the cells that infiltrate renal allografts during rejection include alloproliferative, lymphokine-producing cells of both Leu-2+ and Leu-3+ subsets. The Leu-2+3- cells are also highly cytotoxic against donor lymphocytes, indicating the presence of helper independent cytotoxic T cells. A minor population of Leu-2+3+ T cells that do not express donor specific function was also identified.
通过免疫过氧化物酶染色对6例接受环孢素治疗、肾活检显示急性细胞性同种异体移植排斥反应患者的肾活检组织中存在的T淋巴细胞亚群表型进行原位鉴定，并在体外经白细胞介素2（IL-2）扩增后，通过双色流式细胞术、分选和功能分析进行鉴定。在器官培养8至42天后，在每个培养物中均发现了Leu-3 +（CD4）和Leu-2 +（CD8）亚群，其比例在0.2至5.0之间变化，这与未培养活检组织的原位免疫过氧化物酶染色结果无显著差异。培养的细胞几乎全是Leu-4 +（CD3）T细胞（89%±4），其表达活化标志物DR（82%±6）和IL-2（CD25）受体（15%±4）。Leu-3 +细胞大多为Leu-8 -（90%±6），而少数Leu-2 +细胞为Leu-15 +（CD11）（26%±4）。仅一小部分Leu-2 +细胞对Leu-7呈阳性染色（8%±6）。对经荧光激活细胞分选术（FACS）纯化的Leu-2 - 3 +和Leu-2 + 3 -群体进行功能分析表明，两个亚群在混合淋巴细胞反应（MLR）中均对移植物供体抗原产生增殖反应并产生IL-2。只有Leu-2 + 3 -群体表现出供体特异性细胞毒性活性。每个培养物中的一小部分亚群同时为Leu-3 +和Leu-2 +（2.0%）。从一次活检中纯化得到均一性的Leu-2 + 3 +细胞（99.8%），发现其表达T细胞抗原受体复合物Ti/CD3（WT-31 +，Leu-4 +），但不表达普通胸腺细胞抗原CD1（OKT6）。Leu-2 + 3 +细胞在MLR中无反应，也未显示任何细胞毒性能力。Leu-2 + 3 +细胞能够产生IL-2，但不能产生干扰素-γ。纯化培养物均未显示自然杀伤（NK）活性。一部分纯化的Leu-2 + 3 +细胞在培养1至3周期间失去Leu-2 +，变为Leu-2 - 3 +。这些研究提供了证据，表明在排斥反应期间浸润肾同种异体移植物的细胞包括Leu-2 +和Leu-3 +亚群的同种异体增殖性、产生淋巴因子的细胞。Leu-2 + 3 -细胞对供体淋巴细胞也具有高度细胞毒性，表明存在辅助性独立细胞毒性T细胞。还鉴定出一小部分不表达供体特异性功能的Leu-2 + 3 + T细胞。
