含CpG基序寡核苷酸对慢性乙型肝炎患者外周血树突状细胞的免疫激活研究

Immunostimulatory role of CpG-containing oligodeoxynucleotide on the monocyte-derived dendritic cell in patients with chronic hepatitis B

目的研究含非甲基化 CpG 基序的寡核苷酸(CpG-ODN)对慢性乙型肝炎患者(CHB)外周血树突状细胞(DC)表型和功能的影响。方法以 CD14磁性分选微珠分离 CHB 患者外周血高纯度单核细胞;以重组人粒细胞巨噬细胞集落刺激因子(hGM-CSF)、重组人白细胞介素-4(hIL-4)诱导扩增 DC;以 CpG-ODN 刺激 DC 成熟,并与肿瘤坏死因子(TNF)-α比较,评价其对 DC 表达表面分子人类白细胞组织相容性抗原(HLA)-DR、CD86、CD1a,分泌 IL-12p70以及刺激同种 T 细胞增殖能力的影响。结果与 non-ODN 和磷酸盐缓冲液(PBS)组比较,CpG-ODN 能明显提高 CHB 患者外周血 DC 表面分子 HLA-DR、CD86的表达,使 IL-12分泌增加,刺激同种异体 T 细胞增殖的能力亦显著增强(P=0.017和0.023),但不能明显提高 CD1a 的表达;CpG-ODN 的上述刺激作用接近或略逊于hTNF-α,但差异无统计学意义(P>0.05)。结论 CpG-ODN 与 hTNF-α一样能够促进 CHB 患者外周血 DC 成熟进而增强其抗原提呈功能。

Objective To investigate the immunostimulatory role of CpG-containing oligodeoxynucleotide（CpG-ODN） on dendritic cell（DC） from patients with chronic hepatitis B. Methods Purified monocytes were isolated from the peripheral blood of CHB and healthy volunteers by combination of lymphocyte separating medium and CD14 magnetic cell sorting microbeads. Monocyte-derived DC were generated from monocyte culture in terms of addition of recombined human granulocyte-macrophage colony stimulating factor（hGM-CSF） and interleukin-4 （hIL-4）. At the fifth day of culture, CpG-ODN or other control reagents such as hTNF-α and non-ODN were added respectively to the medium,18 hours later, cells were collected for the detection of surface molecules （HLA-DR/CD86/ CDla）, IL-12p70 levels in the supernatant and stimulating capacity to allogeneic T lymphocytes. Resalts Compared with control groups of non-ODN and PBS,both the expression rates of H LA-DR and CD86, and IL-12pT0 level in culture supernatant of DC from CHB treated with CpG-ODN increased obviously, and the stimulating capacity of DC to allogeneic T lymphocytes in mixed lymphocyte reaction improved significantly（P=0. 017 and 0. 023）. Above effects of CpG-ODN were similar to that of hTNF-α with no significant differences（P〉0.05）. However, neither CpG-ODN nor hTNF-α failed in improving the expression rates of CDla. Conclusions CpG-ODN, like hTNF-α, has remarkable immunostimulatory effect on the differentiation and maturation of monocyte-derived DC from patients with chronic hepatitis B.