妊娠不同时期胎盘单核-巨噬细胞来源树突样细胞刺激脐血T淋巴细胞的特性比较

Comparisons of immunological characteristics of placental monocyte-macrophage-derived dendritic-like cells in stimulating cord blood T lymphocytes in different periods of pregnancy

目的:观察妊娠不同阶段胎盘来源的树突样细胞(DC样细胞)刺激脐血淋巴细胞增殖活化特点的差异,进一步研究胎盘免疫细胞在妊娠耐受及分娩发动中的作用。方法:机械分离妊娠中、晚期胎盘中的单个核细胞,用磁珠分离法获取CD14+细胞,以内皮(逆)穿越系统诱导其分化为DC样细胞,并用流式细胞仪分析细胞表型,ELISA法检测细胞培养上清中白细胞介素(IL)-12、IL-10水平,CCK-8法检测其激发脐血淋巴细胞增殖能力,流式细胞术检测其所刺激的脐血T淋巴细胞内细胞因子。结果:足月妊娠胎盘来源的经诱导的细胞高表达与免疫激活有关的细胞表面标志物CD80、CD86;培养上清中检出较高水平的IL-12,极低水平的IL-10,具有较强的刺激脐血淋巴细胞增殖的能力;能刺激脐血T淋巴细胞分化形成较多的IFN-γ产生细胞,而分化形成的IL-10产生细胞则较少。但相比之下,中期妊娠胎盘来源的经诱导的细胞则低表达CD80、CD86(P<0.05),培养上清中检出较高水平的IL-10(P<0.05),但检测出的IL-12浓度极低(P<0.05),刺激脐血淋巴细胞的能力较弱(P<0.05);刺激脐血T淋巴细胞分化形成较多的IL-10产生细胞(P<0.05),而分化形成的IFN-γ产生细胞较少(P<0.05)。结论:妊娠不同时期胎盘来源的单核-巨噬细胞分化形成的DC样细胞免疫学特性不同,高表达CD80、CD86、IL-12的DC样细胞细胞在妊娠免疫耐受中止及分娩发动中发挥着一定的作用,而低表达CD80、CD86,高表达IL-10的细胞可能参与了妊娠过程中的免疫耐受。

Objective：To observe the differences of characteristics in activation and proliferation of cord blood lymphocytes stimulated by placenta-derived dendritic-like cells in different stages and further study the role of placental immune cells in pregnancy tolerance and labor onset.Methods：Mononuclear cells were separated mechanically from the mid-term placenta and late placenta.CD14＋ cells were obtained by MACS and induced to differentiate into dendritic-like cells through the transebdothelial trafficking system.Flow cytometry was performed to analyze dendritic-like cell phenotype and ELISA assay was performed to detect the levels of IL-12,IL-10 in culture supernatant.CCK-8 assay was used to detect the ability of stimulating the proliferation of cord blood lymphocytes and flow cytometry was used to detect intracellular cytokine produced by stimulated cord blood T lymphocytes.Results：CD14＋ monocyte-macrophage obtained from decidual tissue were inoculated into the endothelial monolayer,after two-way-induced culture,cells derived from full-term placenta had an appearance of dendritic morphology changes and highly expressed cell surface markers related to immune activation of dendritic cells,such as CD80,CD86;simultaneously,higher level of IL-12 and very low level of IL-10 were detected in the culture supernatant,and cells had stronge ability of stimulating the cord blood lymphocytes proliferation.They could stimulate cord blood lymphocytes to differentiate into the cells that most of them producing IFN-γ and less of them producing IL-10.However,the induced cells derived from mid-term placenta,expressed low-level CD80 and CD86（P〈0.05）,in the culture supernatant,higher level of IL-10（P〈0.05） and very low level of IL-12（P〈0.05） were detected.Cells had weaker ability in stimulating cord blood lymphocytes（P〈0.05）,and stimulated cord blood lymphocytes to differentiate to form more cells producing IL-10（P〈0.05） and less cells producing IFN-γ（P〈0.05）.Conclusion：Dendritic-like cells differentiated from placenta-derived monocyte-macrophages in different periods of pregnancy have different immunological characteristics,dendritic-like cells highly expressed CD80、CD86 and high-level IL-12 play a certain role in the termination of immune tolerance in pregnancy and labor onset,dendritic-like cells expressed low-level CD80、CD86,and high-level IL-10 may be involved in immune tolerance during pregnancy.