论文部分内容阅读
为研究肝X受体(LXR)对小鼠骨髓来源耐受性树突状细胞的诱导作用,取C57BL/6纯系小鼠股骨骨髓,将骨髓细胞置于含rmGM-CSF(20ng/ml)、rmIL-4(20ng/ml)、10%FBS、含/或不含LXR激动剂TP0901317(5μmol/L)的RPMI1640培养基中生长。于第7天时检测细胞表型,指标包括:CD11c、CD80、CD86、MHCⅡ、CCR7、PD-L1等;同日加入LPS(100ng/ml)促成熟。LPS作用第3天时,检测细胞表型。以MACS分选未成熟和成熟CD11c+细胞,以丝裂霉素C处理细胞。另取BALB/c小鼠脾脏,MACS分选CD4+T淋巴细胞。将CD11c+细胞与CD4+T细胞混合培养,CCK-8法检测T细胞增殖情况。ELISA法检测细胞培养上清中IL-10水平。研究发现,TP0901317处理组:培养第7天时,CD11c+细胞约为70.2%;CD80、CD86和MHCⅡ在CD11c+细胞上为中低表达,表达水平在40.2%~50%;CD11c+CCR7+细胞约占78%;CD11c+PD-L1+细胞约占90%。以LPS处理促成熟第3天时,CD11c+细胞约为85%;CD80、CD86和MHCⅡ在CD11c+细胞上的表达水平没有明显差异;CD11c+CCR7+细胞约占86%;CD11c+PD-L1+细胞约占99%。混合淋巴细胞反应显示,TP0901317处理组:成熟CD11c+细胞与未成熟细胞一样,均没有刺激CD4+T细胞增殖的能力。而在dTP0901317未处理组:成熟CD11c+细胞具有很强的刺激CD4+T细胞增殖的能力。ELISA检测结果显示,与TP0901317未处理组比较,TP0901317处理组CD11c+细胞能够产生更高水平的IL-10,具有显著意义(P<0.05)。故而可以认为LXR活化能够诱导小鼠骨髓细胞向耐受性树突状细胞分化。
In order to study the induction of bone marrow-derived dendritic cells from liver X receptor (LXR) on murine bone marrow-derived cells, bone marrow cells of C57BL / 6 mice were treated with rmGM-CSF (20ng / ml) , RmIL-4 (20 ng / ml), 10% FBS with and without LXR agonist TP0901317 (5 μmol / L) The cell phenotypes were detected on the 7th day. The indicators included: CD11c, CD80, CD86, MHCⅡ, CCR7, PD-L1 and the like; LPS (100ng / ml) was added on the same day to promote maturation. On the 3rd day of LPS treatment, the cell phenotype was examined. Immature and mature CD11c + cells were sorted by MACS and cells treated with mitomycin C. Another BALB / c mouse spleen, MACS sorting CD4 + T lymphocytes. CD11c + cells and CD4 + T cells were mixed and cultured. The proliferation of T cells was detected by CCK-8 assay. The level of IL-10 in the cell culture supernatant was detected by ELISA. The results showed that CD11c + cells were about 70.2% on the 7th day after culture in TP0901317 treatment group. The expression levels of CD80, CD86 and MHCⅡ on CD11c + cells were low and 40.2% ~ 50%, while those on CD11c + CCR7 + cells were 78% ; CD11c + PD-L1 + cells account for about 90%. The percentage of CD11c + cells was about 85% on the third day after LPS treatment, and there was no significant difference in the expression of CD80, CD86 and MHCⅡ on CD11c + cells. CD11c + CCR7 + cells accounted for about 86% and CD11c + PD-L1 + cells accounted for about 99% %. Mixed lymphocyte reaction showed that, TP0901317 treatment group: mature CD11c + cells and immature cells, no stimulation of CD4 T cell proliferation. In the untreated group of dTP0901317: mature CD11c + cells have a strong ability to stimulate CD4 + T cell proliferation. The results of ELISA showed that compared with TP0901317 untreated group, CD11c + cells treated with TP0901317 could produce higher levels of IL-10, with significant significance (P <0.05). Therefore, LXR activation can be considered to induce the differentiation of mouse bone marrow cells into tolerogenic dendritic cells.