Doi H, Nakagawa S, Nagata K, Hata M, Kasahara S, Sakata T, Suzuki R, Nanno M, Sugamura K, Itoh T
Department of Anatomy, Tohoku University School of Medicine, Sendai, Japan.
Eur J Immunol. 1991 Mar;21(3):783-92. doi: 10.1002/eji.1830210335.
To investigate the role of thymic nurse cells (TNC) in activation and differentiation of fetal CD4-CD8- (double-negative) thymocytes, we have co-cultured murine fetal thymocytes (14-15 days of gestation) with an established murine TNC clone. We show here that TNC induced the growth of the fetal double-negative thymocytes in the presence of recombinant interleukin 2 (rIL2). Activated fetal thymocytes markedly formed lymphocyte-TNC complexes and proliferated extensively after 5 days in the co-culture. The activated fetal thymocytes in this co-culture condition remained double negative after 10 days in culture. None of them gave rise to phenotypically and functionally competent lymphocytes during this period. TNC alone and the supernatant of TNC had no effect on activation. The presence of both TNC and rIL2 was necessary for the growth of fetal thymocytes in our system. The proliferation of fetal thymocytes was inhibited by a monoclonal antibody against mouse IL2 receptors (IL2R). The fetal thymocytes could be maintained further in this co-culture condition. The prolonged cultivation of fetal thymocytes resulted in the establishment of the fetal thymocyte line and its several clones. CD4 single-positive cells of activated fetal thymocytes first appeared 14 days after the onset of culture and their number increased, whereas CD8+ cells or CD4CD8 double-positive cells were not observed. These results indicate that fetal CD4-CD8- thymocytes underwent phenotypic change after long periods of culture. All established clones of fetal thymocytes are CD4 single positive showing lymphocyte-TNC interactions but do not express CD3 complex. Northern blot analysis detected mRNA for the gamma T cell receptor, but no messages for the delta, alpha or beta T cell receptor. Chemical cross-linking of 125I-labelled IL2 revealed that the 90-kDa band (presumably considered to be the IL2R beta chain) was clearly present in IL2-responsive fetal clones, whereas freshly isolated day 14-15 fetal thymocytes lacked the band. Taken together, TNC might be involved in the differentiation and/or expansion of murine fetal thymocytes by inducing IL2R beta chain, which forms the functional IL2R together with IL2R alpha chain and CD4, one of the T cell accessory molecules, on the cell surface through direct cell-cell interaction.
为了研究胸腺哺育细胞（TNC）在胎儿CD4-CD8-（双阴性）胸腺细胞激活和分化中的作用，我们将小鼠胎儿胸腺细胞（妊娠14-15天）与已建立的小鼠TNC克隆进行了共培养。我们在此表明，在重组白细胞介素2（rIL2）存在的情况下，TNC诱导了胎儿双阴性胸腺细胞的生长。激活的胎儿胸腺细胞在共培养5天后明显形成淋巴细胞-TNC复合物并大量增殖。在这种共培养条件下激活的胎儿胸腺细胞在培养10天后仍保持双阴性。在此期间，它们均未产生表型和功能上合格的淋巴细胞。单独的TNC和TNC的上清液对激活没有影响。在我们的系统中，TNC和rIL2的存在对于胎儿胸腺细胞的生长是必需的。抗小鼠IL2受体（IL2R）的单克隆抗体抑制了胎儿胸腺细胞的增殖。胎儿胸腺细胞可以在这种共培养条件下进一步维持。胎儿胸腺细胞的长期培养导致建立了胎儿胸腺细胞系及其多个克隆。激活的胎儿胸腺细胞的CD4单阳性细胞在培养开始后14天首次出现，其数量增加，而未观察到CD8+细胞或CD4CD8双阳性细胞。这些结果表明，胎儿CD4-CD8-胸腺细胞在长期培养后发生了表型变化。所有已建立的胎儿胸腺细胞克隆均为CD4单阳性，显示淋巴细胞-TNC相互作用，但不表达CD3复合物。Northern印迹分析检测到γT细胞受体的mRNA，但未检测到δ、α或βT细胞受体的信息。125I标记的IL2的化学交联显示，90-kDa条带（可能被认为是IL2Rβ链）在对IL2有反应的胎儿克隆中明显存在，而新鲜分离的第14-15天胎儿胸腺细胞缺乏该条带。综上所述，TNC可能通过诱导IL2Rβ链参与小鼠胎儿胸腺细胞的分化和/或扩增，IL2Rβ链与IL2Rα链以及T细胞辅助分子之一的CD4在细胞表面形成功能性IL2R，通过直接的细胞-细胞相互作用实现。
