e抗原阳性慢性乙型肝炎患者外周血树突状细胞干扰素β的表达

Expression of IFN-beta on peripheral blood dendritic cells in HBeAg positive chronic hepatitis B

目的探讨慢性乙型肝炎患者树突状细胞干扰素β(IFN-β)表达与乙型肝炎病毒(HBV)感染后宿主免疫清除障碍的关系。方法选择e抗原阳性慢性乙型肝炎患者52例,健康对照48例,用免疫磁珠细胞分选乙型肝炎e抗原(HBeAg)法从外周血获得纯化的CD14+单核细胞,并用hGM-CSF、hIL-4诱导单核细胞成为未成熟的髓样树突状细胞(mDC),加入polyI:C(25μg/ml)刺激后获得成熟的mDC,在刺激后0h、12h、24h、48h用流式细胞仪检测细胞表面分化抗原CD86、HLA-DR、CD1a,real-time PCR检测IFN-β的表达变化。用ELISA方法检测mDC细胞上清液中IFN-β、IL-12、IL-10的浓度变化。结果两组mDC上0hIFN-β mRNA表达水平及细胞上清液中IFN-β浓度差异无统计学意义(P>0.05)。健康对照组mDC在12hIFN-β mRNA表达水平及IFN-β浓度与0h相比显著升高(P<0.05),较患者组0h、12h、24h、48hIFN-β表达显著升高(P<0.05)。而患者组mDC刺激后12h、24h和48hIFN-β mRNA及IFN-β表达水平与0h相比无显著变化。与0h比较,健康对照组12h、24h、48hCD86表达水平较患者组显著上升(P<0.05)。两组之间CD1a、HLA-DR表达差异无统计学意义。二组IL-10及IL-120h表达水平差异无统计学意义(P>0.05),患者组IL-10表达24h及48h与0h相比明显上升(P<0.05),而对照组12h、24h、48hIL-10表达水平没有明显上升。与之相反,患者组12h、24h、48hIL-12表达与0h相比表达水平没有明显上升,对照组在12hIL-12表达水平与0h相比差异有统计学意义(P<0.05),24h、48hIL-12表达水平继续升高(P<0.05)。结论慢性HBV感染者mDC受polyI:C刺激后IFN-β表达异常,协同刺激因子CD86表达低下,可能造成宿主对HBV感染的免疫清除障碍,导致疾病慢性化。

Objective To explore the roles of IFN-β expression on dendritic cells （DCs） in chronic hepatits B. Methods Monocytes were isolated from fresh peripheral blood of 22 healthy volunteers （HV） and 28 chronically HBV infected patients （CH）. DCs were induced and proliferated in the culture medium with rhGM-CSF and rhIL-4. We stimulated DCs with polyI： C and examined HLA-DR, CD86, and CDla expression at 0 h, 24 h and 48 h by Flow Cytometry. The mRNA expression of IFN β were quantified by real-time PCR. The concentrations of IFN-β, IL-12 and IL-10 in supernatant were determined by ELISA. Results The concentration of IFN-β had no significant changes at 0 h, 12 h, 24 h and 48 h in CH groups, whereas, there was a significant increase at 12 h in HV groups. The concentration of IL-10 displayed a significant increase at 12 h in CH groups, but there was no change in HV groups. In contrast, the concentration of IL-12 had a significant increase at 12 h in HV groups, hut there was no change in CH groups. The rate of CD86 expressions were increased after PolyI： C stimulation, the increased rate are 12. 6%, 23.8%, 20.7% in CH groups, and 31. 0%, 43.4%, 44.6% in HV groups at 12 h, 24 h and 48 h after PolyI：C stimulation, respectively. There was a significant increase of the expression level of CD86 in HV groups （P〈0.01）. In contrast, only slight increase was found in CH groups （31.0% vs 12.6%）. The differences （P〈0. 01） between the groups were maintained at 24 h and 48 h. No significant differences were detected in HLA-DR and CDI a between two groups. Conclusion The expression level of IFN-β.IL- 10,IL-12 has not increased in CH groups. A marked increase of the expression level of CD86 is observed in HV groups. Our results suggest that abnormal expression of IFN-β,IL-10,IL-12 and CD86 may lead to persistent infection of HBV.