腺病毒载体转染人未成熟树突状细胞对其成熟特性的影响

Influence of adenovirus transfection on the maturation characteristics of human immature dendritic cells

目的观察重组腺病毒载体AdEASY-增强型绿色荧光蛋白(EGFP)转染人未成熟树突状细胞(imDC)后,其表型特征及免疫学功能的变化,并探讨白细胞介素(IL)10对腺病毒转染诱导imDC成熟的抑制作用。方法贴壁法分离人脐带血来源的单核细胞,利用重组粒细胞巨噬细胞集落刺激因子(GM-CSF)和IL-4诱导分化imDC。对照组为常规培养的imDC,转染组用AdEASY-EGFP转染imDC,IL-10组用IL-10处理转染后细胞。流式细胞仪检测细胞表面成熟标志[CD86、CD83和人类白细胞DR抗原(HLA-DR)],混合淋巴细胞反应(MLR)检测其刺激同种异体未致敏T淋巴细胞的增殖能力。结果腺病毒转染imDC后,转染组细胞成熟表型表达率分别为CD86:46±10、CD83: 38±7、HLA-DR:82±10,均较对照组(10±7、8±3、68±8)显著上调,且刺激T淋巴细胞增殖的能力显著加强(SI>2．0)。IL-10组处理的imDC上述表型表达率分别为CD86:8±5、CD83:9±3、HLA-DR: 63±12,与对照组比较差异无统计学意义(P>0．05),其刺激T淋巴细胞增殖的能力也明显下降。结论腺病毒能有效转染imDC,但在转染后有促进其成熟的趋势;用IL-10能有效抑制该成熟状态。

Objective To observe the changes in the phenotype characteristics and immune function after transfection of cord blood derived immature dedritics cells（imDC） with Adeasy-EGFP adenovirus vector. and to explore the function of IL-10 in inhibition of imDC maturation, Methods Immature dendritic cells were generated from human cord blood （CB） monocyte cultured with rhGM-CSF and rhIL-4. The recombinant adenovirus vector AdEASY-EGFP was transduced into immature dendritic cells on the third day with or without adding IL-10. The expression of cell maturation marker CD83, CD86 and HLA-DR were determined with flow cytometry. Allogenetic mixed leukocyte reaction（MLR） was used to examine the imDC＇s ability to promote T cell proliferation. Results The expression of surface maturation markers of imDC after transfection with adenovirus were significantly up-regulated （ CD86 ： 46 ± 10 ; CD83 ： 38 ± 7 ; HLA-DR ： 82 ± 12） , and its ability to promote T cell proliferation was also obviously increased（ SI 〉 2.0）. However, the expression of surface maturation markers of imDC after IL-10 treatment had lower mature penotypes expression after transduction （ CD86 -8 ± 5 ; CD83 ： 9 ± 3 ; HLA-DR ： 63 ±12 ） , and T cell stimulating ability was decreased comparing with adenovirus transfection groups. Conclusion Adenovirus can be transduced into imDC with high efficiency, but transfection itself can promote imDC＇s maturation. IL-10 treatment can inhibit the tendency to maturation stimulated by adnovirus transduction efficiently.