Mayer T G, Fuller A A, Fuller T C, Lazarovits A I, Boyle L A, Kurnick J T
J Immunol. 1985 Jan;134(1):258-64.
To evaluate in situ lymphocyte responses in cell-mediated immune tissue injury, we have developed an approach for propagation of human allospecific T lymphocytes directly from tissue biopsies. We have utilized renal allograft tissue obtained from eight patients undergoing cellular rejection. Needle biopsy tissue was cultured in medium containing interleukin 2 (IL 2), including recombinant-DNA-produced IL 2. In each case, lymphoblasts migrated out of the tissue and increased in numbers, especially adjacent to the tissue. In two cases in which there was no cellular infiltrate present in the biopsy, no lymphocytes proliferated in vitro. Instead, fibroblasts eventually filled the wells from these allograft biopsies. The continued presence of the allograft tissue enhanced the viability and growth of the lymphoblasts in cultures from rejecting allografts. The isolated lymphoblasts had surface markers of mature OKT3+ lymphocytes of either OKT4+ or OKT8+ subsets. OKT8+ cells predominated. There was variability (41 to 97%) in the percentage of T lymphoblasts that bore surface HLA-DR antigens. In assays of lymphoblasts obtained from eight separate renal allografts, there was donor-specific cytotoxicity, and in all but two of the cases there was donor-induced proliferation. The specificity of the cytotoxic reaction was tested by using 51Cr-labeled, PHA-stimulated target cells prepared from a panel of HLA-typed donors. Proliferation was tested after 48 hr in the presence of mitomycin C-treated peripheral blood mononuclear cells as stimulator cells by using only 10(4) responder T lymphoblasts. Of particular note was that the cytotoxicity of the isolated lymphoblasts showed specificity against both "private" HLA class I alloantigens (of the allograft donor) as well as "public" cross-reacting epitopes. This method permits the propagation and functional characterization of in vivo-activated T lymphoblasts that are obtained from the actual sites of immune-mediated injury. Preliminary studies of other tissues with diverse inflammatory processes indicate the possible widespread applicability of obtaining in vivo-activated lymphocytes.
为了评估细胞介导的免疫组织损伤中的原位淋巴细胞反应，我们开发了一种直接从组织活检中扩增人同种异体特异性T淋巴细胞的方法。我们使用了从八名经历细胞排斥反应的患者身上获取的肾移植组织。针吸活检组织在含有白细胞介素2（IL-2）的培养基中培养，包括重组DNA产生的IL-2。在每种情况下，淋巴母细胞从组织中迁移出来并数量增加，尤其是在组织附近。在两例活检中没有细胞浸润的情况下，体外没有淋巴细胞增殖。相反，成纤维细胞最终填满了这些同种异体移植活检的孔。同种异体移植组织的持续存在增强了来自排斥同种异体移植的培养物中淋巴母细胞的活力和生长。分离出的淋巴母细胞具有OKT4 +或OKT8 +亚群的成熟OKT3 +淋巴细胞的表面标志物。OKT8 +细胞占主导。带有表面HLA-DR抗原的T淋巴母细胞百分比存在变异性（41%至97%）。在从八个单独的肾移植中获得的淋巴母细胞的检测中，存在供体特异性细胞毒性，除两例病例外，在所有病例中均有供体诱导的增殖。通过使用从一组HLA分型供体制备的51Cr标记、PHA刺激的靶细胞来测试细胞毒性反应的特异性。在存在丝裂霉素C处理的外周血单核细胞作为刺激细胞的情况下，仅使用10(4)个反应性T淋巴母细胞在48小时后测试增殖。特别值得注意的是，分离出的淋巴母细胞的细胞毒性对“私人”HLA I类同种异体抗原（同种异体移植供体的）以及“公共”交叉反应表位均表现出特异性。该方法允许对从免疫介导损伤的实际部位获得的体内活化T淋巴母细胞进行扩增和功能表征。对具有不同炎症过程的其他组织的初步研究表明，获取体内活化淋巴细胞可能具有广泛的适用性。
