落新妇苷对小鼠骨髓来源树突状细胞成熟及免疫功能的抑制作用

Effects of astilbin on maturation and immunologic function of mouse bone marrow-derived dendritic cells

目的:探讨落新妇苷对小鼠骨髓来源树突状细胞(dendritic cell,DC)成熟和免疫功能的影响。方法:采用体外条件培养法得到小鼠骨髓来源未成熟树突状细胞,加入脂多糖刺激细胞成熟。落新妇苷(低浓度25μg/mL、中浓度50μg/mL、高浓度100μg/mL)处理细胞后,流式细胞仪分析各组细胞凋亡,细胞表面主要组织相容性抗原复合物分子Ⅰa和共刺激分子CD40、CD80和CD86表达;异硫氰酸荧光素标记葡聚糖内吞实验分析各组DC抗原吞噬功能;酶联免疫吸附测定法检测各组DC培养液上清白细胞介素12亚单位p40(p40 subunit of interleukin-12,IL-12p40)水平;氚-胸腺嘧啶核苷掺入法检测体外混合淋巴细胞反应(mixed lymphocyte reaction,MLR)中各组DC刺激同种反应性T细胞的增殖能力,酶联免疫吸附测定法检测MLR上清IL-2、IL-4、IL-10、γ干扰素水平。结果:各组DC凋亡率差异无统计学意义(P>0.05)。与脂多糖组相比,高、中、低浓度落新妇苷组细胞表面主要组织相容性抗原复合物分子Ⅰa和共刺激分子表达明显降低,抗原吞噬能力明显升高,分泌IL-12p40明显减少,刺激同种反应性T细胞增殖的能力显著下降(P<0.05)。与脂多糖组相比,高、中、低浓度落新妇苷组MLR上清中IL-2和γ干扰素水平明显降低(P<0.05),而IL-4、IL-10水平比较,差异无统计学意义(P>0.05)。结论:落新妇苷(25、50、100μg/mL)能够剂量依赖性地抑制体外培养小鼠骨髓来源DC的成熟,并对其免疫功能具有一定的负性调节作用。

Objective： To explore the effects of astilbin on the maturation and immunologic function of mouse bone marrow-derived dendritic cells （DCs）. Methods： Mouse bone marrow cells were cultured with recombinant mouse granulocyte-macrophage colony-stimulating factor and interleukin-4 （IL-4） for 5 days to get immature DCs （imDCs）, then the imDCs was cultured in the presence of 1 μg/mL lipopolysaccharide （LPS） or LPS （1 μg/mL） plus astilbin （25, 50, 100 μg/mL） for 48 h. Then, the cells were harvested, and the apoptosis, immunophenotypes and antigen phagocytosis capability of imDCs in LPS, and low-, medium- and high-dose astilbin groups were analyzed by flow cytometry. Contents of p40 subunit of interleukin-12 （IL-12p40） in the supernatants were detected with enzyme-linked immunosorbent assay （ELISA）. The stimulatory activity of the harvested cells on allogeneic T cells in mixed lymphocyte reactions （MLR） was tested by incorporation of 3H-thymidine, and the contents of IL-2, IL-4, IL-10 and interferon-γ （INF-γ） in the supernatants of MLR were examined by ELISA. Results： At the concentrations of 25 to 100 μg/mL, astilbin exhibited no toxicity on co-cultured DCs. Compared with the lipopolysaccharide, low-, medium- and high-dose astilbin could decrease the expression levels of major histocompatibility complex-Ⅰa （MHC-Ⅰa）, CD40, CD80 and CD86 molecules in DCs. DCs in the low-, medium- and high-dose astilbin groups exhibited weaker capabilities for antigen phagocytosis and less contents of IL-12p40 in the supernatants than in the LPS group. Furthermore, low-, medium- and high-dose astilbin showed weak activities in stimulating the proliferation of allogeneic T cells as compared with the LPS （P〈0.05）. Compared with the LPS, low-, medium- and high-dose astilbin could decrease IL-2 and INF-γ secretion from T cells in MLR but had no effect on IL-10 secretion. Conclusion： Astilbin can inhibit maturation of mouse bone marrow-derived DCs with dose-dependent effect and exert negative effects on immunologic function of the DCs.