Generation of Inducible Immortalized Dendritic Cells with Proper Immune Function In Vitro and In Vivo
Generation of Inducible Immortalized Dendritic Cells with Proper Immune Function In Vitro and In Vivo
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DOI：
10.1371/journal.pone.0062621
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发表时间：
2013-04-23
期刊：
影响因子：
3.7
通讯作者：
Brenner, Sebastian
中科院分区：
文献类型：
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作者：
Richter, Cornelia;Thieme, Sebastian;Brenner, Sebastian
树突状细胞是天然免疫的专职抗原提呈细胞，在维持免疫应答平衡中起着关键作用。树突状细胞的研究主要受到体内数量少和体外维护困难的限制。我们将表达可诱导的SV40大T抗原的转基因小鼠的骨髓细胞分化为树突状细胞。经地塞米松和强力霉素永生化后，这些细胞在长期培养中是稳定的。在没有地塞米松和强力霉素(去诱导)的情况下，树突状细胞表现出原代细胞的特性，其特征是表达经典的树突状细胞表面标志CD11c、CD11b、MHCII、CD40和CD86。此外，去诱导的脂多糖激活的树突状细胞分泌IL-1β、IL-6、TNFα和IL-12。去诱导的、负载卵清蛋白的树突状细胞将CD4(+)T细胞极化为Th1、Th17和Th2细胞，表明它们具有正确的抗原提呈特性。与小鼠气管内应用携带卵清蛋白的原始树突状细胞一致，去诱导树突状细胞的应用导致淋巴细胞向肺内募集。综上所述，我们成功地使用条件永生化技术扩增了树突状细胞。所产生的树突状细胞具有原代树突状细胞特有的免疫表型，将有助于进一步研究树突状细胞的免疫调节特性。
Dendritic cells are the professional antigen presenting cells of innate immunity and key players in maintaining the balance of immune responses. Studies with dendritic cells are mainly limited by their low numbers in vivo and their difficult maintenance in vitro. We differentiated bone marrow cells from transgenic mice expressing an inducible SV40 large T-antigen into dendritic cells. When immortalized by dexamethasone and doxycycline, these cells were stable in long-term culture. In the absence of dexamethasone and doxycycline (de-induction), dendritic cells displayed properties of primary cells, characterized by expression of classical dendritic cell surface markers CD11c, CD11b, MHCII, CD40 and CD86. Furthermore, de-induced lipopolysaccharide activated dendritic cells secreted IL-1 beta, IL-6, TNF alpha and IL-12. De-induced, Ovalbumin-loaded dendritic cells polarize CD4(+) T cells into Th1, Th17 and Th2 cells, indicating their correct antigen presenting property. Consistent with intratracheal application of Ovalbumin-loaded primary dendritic cells into mice, the application of de-induced dendritic cells resulted in recruitment of lymphocytes to the lungs. In summary, we successfully expanded dendritic cells using conditional immortalization. The generated dendritic cells demonstrate the characteristic immunophenotype of primary dendritic cells and will facilitate further studies on immunomodulatory properties of dendritic cells.