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目的探讨香加皮羽扇豆烷乙酸酯(CPLA)对人外周血单个核细胞(PBMC)来源的树突状细胞(DC)在体外分化成熟及免疫活性的影响。方法从人外周血分离单个核细胞,与细胞因子GM-CSF、IL-4共培养,于第5天加入DC的促成熟刺激剂TNF-α(阳性对照组)或CPLA。倒置显微镜和透射电镜下观察DC的形态;应用流式细胞术检测成熟DC的表面标志CD1a、CD83、CD80和CD86的表达情况;用ELISA检测DC培养上清中IL-12和IFN-γ的含量;用MTT法测定DC刺激T细胞增殖的能力。结果培养10d后,经CPLA刺激的PBMC呈现出典型DC的形态学特征;成熟DC的特征性表面分子CD1a、CD83、CD80和CD86表达水平均明显上调(P<0.05);细胞培养上清中IL-12和IFN-γ含量明显增高(P<0.05);刺激T细胞增殖的能力明显增强(P<0.05)。结论CPLA可诱导PBMC来源的DC分化成熟,并可促进其细胞因子的分泌,增强DC的免疫调节活性。
Objective To investigate the effects of lupinol acetate (CPLA) on the differentiation and maturation of dendritic cells (DCs) derived from human peripheral blood mononuclear cells (PBMCs) in vitro. Methods Mononuclear cells were isolated from human peripheral blood and co-cultured with the cytokines GM-CSF and IL-4. On the fifth day, the mature stimulant TNF-α (positive control group) or CPLA of DC was added. The morphology of DCs was observed under inverted microscope and transmission electron microscope; the expression of surface markers CD1a, CD83, CD80, and CD86 in mature DCs was detected by flow cytometry; the levels of IL-12 and IFN-γ in DC culture supernatants were measured by ELISA. The ability of DC to stimulate T cell proliferation was determined by MTT assay. Results After 10 days of culture, PBMCs stimulated by CPLA showed the morphological features of typical DCs; the expression levels of CD1a, CD83, CD80, and CD86 were significantly increased in mature DCs (P<0.05); IL in cell culture supernatants The levels of -12 and IFN-γ were significantly increased (P<0.05); the ability to stimulate T cell proliferation was significantly increased (P<0.05). Conclusion CPLA can induce the differentiation and maturation of PBMC-derived DCs, promote the secretion of cytokines, and enhance the immune regulatory activity of DCs.