Jarnjak-Jankovic Silvija, Hammerstad Hege, Saebøe-Larssen Stein, Kvalheim Gunnar, Gaudernack Gustav
Department of Pediatric Research, The National Hospital, Oslo, Norway.
BMC Cancer. 2007 Jul 3;7:119. doi: 10.1186/1471-2407-7-119.
Dendritic cells (DCs) are professional antigen-presenting cells with the ability to induce primary T-cell responses and are commonly produced by culturing monocytes in the presence of IL-4 and GM-CSF for 5-7 days (Standard DC). Recently, Dauer and co-workers presented a modified protocol for differentiation of human monocytes into mature DCs within 48 hours (Fast DC). Here we report a functional comparison of the two strategies for generation of DCs from human monocytes with adaptions for large-scale clinical use.
The Elutra Cell Selection System was used to isolate monocytes after collection of leukapheresis product. The enriched monocytes were cultured in gas permeable Teflon bags with IL-4 and GM-CSF for 24 hours (Fast DC) or 5 days (Standard DC) to obtain immature DCs. The cells were then transfected with mRNA from the leukemia cell line Jurkat E6 by electroporation and incubated for additional 24 h or 2 days in the presence of pro-inflammatory cytokines (TNFalpha, IL-1beta, IL-6 and PGE2) to obtain mature DCs.
Mature Fast DC and Standard DC displayed comparable levels of many markers expressed on DC, including HLA-DR, CD83, CD86, CD208 and CCR7. However, compared to Standard DC, mature Fast DC was CD14high CD209low. Fast DC and Standard DC transfected with Jurkat E6-cell mRNA were equally able to elicit T cell specifically recognizing transfected DCs in vitro. IFNgamma-secreting T cells were observed in both the CD4+ and CD8+ subsets.
Our results indicate that mature Fast DC are functional antigen presenting cells (APCs) capable of inducing primary T-cell responses, and suggest that these cells may be valuable for generation of anti-tumor vaccines.
树突状细胞（DCs）是专业的抗原呈递细胞，具有诱导初始T细胞反应的能力，通常通过在IL-4和GM-CSF存在的情况下培养单核细胞5至7天来产生（标准DC）。最近，道尔及其同事提出了一种改良方案，可在48小时内将人单核细胞分化为成熟DC（快速DC）。在此，我们报告了两种从人单核细胞生成DC的策略的功能比较，并对其进行了大规模临床应用的适应性调整。
使用Elutra细胞分选系统在采集白细胞分离产品后分离单核细胞。将富集的单核细胞在透气的聚四氟乙烯袋中用IL-4和GM-CSF培养24小时（快速DC）或5天（标准DC）以获得未成熟DC。然后通过电穿孔用白血病细胞系Jurkat E6的mRNA转染细胞，并在促炎细胞因子（TNFα、IL-1β、IL-6和PGE2）存在的情况下再孵育24小时或2天以获得成熟DC。
成熟的快速DC和标准DC在DC上表达的许多标志物水平相当，包括HLA-DR、CD83、CD86、CD208和CCR7。然而，与标准DC相比，成熟的快速DC CD14高CD209低。用Jurkat E6细胞mRNA转染的快速DC和标准DC在体外同样能够引发特异性识别转染DC的T细胞。在CD4+和CD8+亚群中均观察到分泌IFNγ的T细胞。
我们的结果表明，成熟的快速DC是能够诱导初始T细胞反应的功能性抗原呈递细胞（APC），并表明这些细胞可能对生成抗肿瘤疫苗有价值。
