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(f) DCs isolated from unexpanded C57BL/6 mice were used to activate Thy1.1+ OTII cells in vitro, which were then adoptively transferred into separate Thy1.2+ C57BL/6 recipient mice, followed by three inhaled OVA challenges. Thy1.1+ OTII cells in the BAL and lung from recipients of DC activated OTII cells were enumerated. n = 9–23 mice per group total from three to six independent experiments. To ensure that Flt3L did not cause the lung-homing advantage of lung DC-activated T cells, we repeated these experiments using DCs isolated from mice that were not treated with Flt3L. Because these untreated mice did not have an expanded population of DCs, tissues were pooled from up to 20 mice per experiment to obtain adequate numbers of DCs. Single cell preparations of unexpanded tissues were rested overnight. The next day, nonadherent cells were collected, and CD11c+ cells were isolated with purity >97% (not depicted). DCs were used to activate CD4+ T cells in vitro, yielding similar levels of T cell differentiation and activation (not depicted), followed by adoptive transfer experiments as described above. Despite the presence of some alveolar macrophages in the lung DC preparations from unexpanded mice, lung DC-activated T cells trafficked significantly more efficiently into the BAL and lung compared with spleen and SLN DC-activated T cells (Fig.1f, left, BAL: 9- and 10-fold, respectively; right, lung: 2.5- and 6.6-fold, respectively). 如前所述，作者为了排除 Flt3L对于T细胞迁移特性的影响，只能牺牲了一大批的小鼠获取足够多的各组织的DC，实验结果与使用Flt3L类似。 教育部发话了，大批量克隆班主任可不行，校领导只能把各个城市叫Lung DC、叫spleen DC 和 SLN DC的人都召唤过来了。虽然最终教出来的效果跟克隆是一样的。 |
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