Ratta M, Rondelli D, Fortuna A, Curti A, Fogli M, Fagnoni F, Martinelli G, Terragna C, Tura S, Lemoli R M
Institute of Haematology and Medical Oncology Seràgnoli, University of Bologna, Italy.
Br J Haematol. 1998 Jun;101(4):756-65. doi: 10.1046/j.1365-2141.1998.00771.x.
Dendritic cells (DCs) are the most powerful professional antigen-presenting cells (APC), specializing in capturing antigens and stimulating T-cell-dependent immunity. In this study we report the generation and characterization of functional DCs derived from both steady-state bone marrow (BM) and circulating haemopoietic CD34+ cells from 14 individuals undergoing granulocyte colony-stimulating factor (G-CSF) treatment for peripheral blood stem cells (PBSC) mobilization and transplantation. Clonogenic assays in methylcellulose showed an increased frequency and proliferation of colony-forming unit-dendritic cells (CFU-DC) in circulating CD34+ cells, compared to that of BM CD34+ precursors in response to GM-CSF and TNF-alpha with or without SCF and FLT-3L. Moreover, peripheral blood (PB) CD34+ cells generated a significantly higher number of fully functional DCs, as determined by conventional mixed lymphocyte reactions (MLR), than their BM counterparts upon different culture conditions. DCs derived from mobilized stem cells were also capable of processing and presenting soluble antigens to autologous T cells for both primary and secondary immune response. Replacement of the early-acting growth factors SCF and FLT-3L with IL-4 at day 7 of culture of PB CD34+ cells enhanced both the percentage of total CD1a+ cells and CD1a+ CD14- cells and the yield of DCs after 14 d of incubation. In addition, the alloreactivity of IL-4-stimulated DCs was significantly higher than those generated in the absence of IL-4. Furthermore, autologous serum collected during G-CSF treatment was more efficient than fetal calf serum (FCS) or two different serum-free media for large-scale production of DCs. Thus, our comparative studies indicate that G-CSF mobilizes CD34+ DC precursors into PB and circulating CD34+ cells represent the optimal source for the massive generation of DCs. The sequential use of early-acting and intermediatelate-acting colony-stimulating factors (CSFs) as well as the use of autologous serum greatly enhanced the growth of DCs. These data may provide new insights for manipulating immunocompetent cells for cancer therapy.
树突状细胞（DCs）是最强大的专职抗原呈递细胞（APC），专门负责捕获抗原并刺激T细胞依赖性免疫。在本研究中，我们报告了来自稳态骨髓（BM）以及14名接受粒细胞集落刺激因子（G-CSF）治疗以进行外周血干细胞（PBSC）动员和移植的个体的循环造血CD34+细胞所衍生的功能性DCs的生成及特性。甲基纤维素中的克隆形成试验显示，与BM CD34+前体细胞相比，循环CD34+细胞中集落形成单位树突状细胞（CFU-DC）的频率和增殖在有或无干细胞因子（SCF）和FMS样酪氨酸激酶3配体（FLT-3L）的情况下，对粒细胞-巨噬细胞集落刺激因子（GM-CSF）和肿瘤坏死因子-α（TNF-α）的反应有所增加。此外，在外周血（PB）CD34+细胞在不同培养条件下产生的完全功能性DCs数量显著高于其BM对应细胞，这通过传统的混合淋巴细胞反应（MLR）得以确定。源自动员干细胞的DCs也能够处理可溶性抗原并将其呈递给自体T细胞以进行初次和二次免疫反应。在PB CD34+细胞培养第7天用白细胞介素-4（IL-4）替代早期作用生长因子SCF和FLT-3L，可提高培养14天后总CD1a+细胞和CD1a+ CD14-细胞的百分比以及DCs的产量。此外，IL-4刺激的DCs的同种异体反应性显著高于未使用IL-4产生的DCs。此外，在G-CSF治疗期间收集的自体血清在大规模生产DCs方面比胎牛血清（FCS）或两种不同的无血清培养基更有效。因此，我们的比较研究表明，G-CSF可将CD34+ DC前体细胞动员到PB中，循环CD34+细胞是大量生成DCs的最佳来源。早期作用和中晚期作用的集落刺激因子（CSF）的序贯使用以及自体血清的使用极大地促进了DCs的生长。这些数据可能为操纵免疫活性细胞用于癌症治疗提供新的见解。
