CpG-ODN联合HBsAg体外对慢性乙肝患者外周血树突状细胞STAT、SOCS通路的影响

Effect of CpG-ODN combined with HBsAg on the intracellular signaling pathway of monocyte-derived dendritic cell from chronic hepatitis B patients

目的研究含非甲基化CpG基序的免疫刺激寡核苷酸(CpG-ODN)联合重组HBsAg对慢性乙肝(CHB)患者外周血树突状细胞(dendriticcell,DC)表面标志、功能及胞浆信号传导与转录激活因子(STAT)、细胞信号转导抑制因子(SOCS)表达的影响。方法由CHB患者和健康者外周血单核细胞诱导扩增DC,以CpG2ODN或联合HBsAg刺激DC,并与TNF2α比较,评价其对DC表面标志、分泌IL212p70及刺激同种T细胞增殖能力的影响;应用Westernblot法检测DC胞浆STAT1、3、4、5、6以及SOCS1、3蛋白的表达。结果与PBS组比,TNF2α、CpG-ODN或联合HBsAg能明显提高CHB患者DC表面分子HLA-DR表达,IL-12p70分泌增加,刺激同种T细胞增殖能力增强,CpG-ODN联合HBsAg还能明显提高CD1a的表达;CpG-ODN或联合HBsAg、TNF2α对DC胞浆STAT1、4、6和SOCS1、3的表达呈不同程度增强作用,对STAT3、5的表达无明显影响。结论CpG-ODN与TNF-α一样能促进CHB外周血DC分化、成熟;CpG2ODN或联合HBsAg能增强DC的特异性抗原提呈作用;其作用机制可能是通过调节DC细胞内STAT1、4、6以及SOCS1、3等信号蛋白的表达。

Objective To investigate the effect of synthetic nonmethylated CpG containing oligodeoxynu-eleotide（CpG-ODN） combined with recombined HBsAg polypepfide on the intracellular signaling pathways of monocyte-derived dendritic eells（DC） in patients with chronic hepatitis B（CHB）. Methods Purified monocytes were isolated from the peripheral blood of CHB and healthy volunteers by combination of HISTOPAQUE-1.077 and CD14 Mien）Beads. Cells were collected for detection of surface molecules（HLA-DR/CD86/CD1a）, IL-12 p70 levels in the supernatant and of stimulating capacity to allogenie T lymphocytes. The expression of signaling molecules, activators of transcription（STAT1, 3, 4, 5, 6） and suppressors of cell signaling（SOCS1, 3） in DC were detected by Western blot assay. Results As compared to control group, the expression rates of HLA-DR of DC in CHB with the group of CpG-ODN or group combined with HBsAg were obviously increased; both IL-12 p70 level and the stimulating capacity of DC to allogenic T lymphocytes were significantly improved in CpG-ODN group and CpG-ODN/HBsAg combination group（ P〈0.01 and 0.05, respectively） ; in particular, the expression of CDla increased only in the CpG-ODN/HBsAg combination group. As compared to increased control group, both CpG-ODN （or combined with HBsAg） and TNF-α enhanced the expression of cytoplasmic phosphorylated STAT1, 4, 6 and SOCS1, 3 in DC of CHB, but have no effect on expression of STAT3, 5. Conclusion CpG-ODN（or combined with HBsAg）, like TNF-α, has remarkable stimulatory effect to the impaired phenotype and function of monocyte-derived DC in patients with chronic hepatitis B possibly through regulating the expression of signal molecules of STAT1, 4, 6 and SOCS1, 3 in DC.