Objective: To explore the effect of nerve growth factor (NGF) on the differentiation of murine bone marrow-derived dendritic cells (DCs) in vitro.
Methods: The bone marrow cells of femur and tibia from healthy C57B -L/6 mice were isolated and divided into 4 groups: a phosphate buffered saline (PBS) group (PBS group), a NGF group, a granulocyte monocyte colony stimulating factor (GM-CSF) plus interleukin 4 (IL-4) group (GM-CSF+IL-4 group), and a GM-CSF plus IL-4 and NGF group (n=6 in each group). The positive rate of CD11c+ and the proportion of CD8a- were compared at the 7th day among the different groups by flow cytometry. The immature DCs were acquired by classic methods with GM-CSF and IL-4. The purified DCs were obtained by magnetic bead positive selection for CD11c+ cells. The immature DCs were divided into 4 groups: a PBS group, a NGF group, a LPS group, and a NGF+LPS group (n=6 in each group), which were incubated with PBS, NGF, LPS and NGF+LPS, respectively. Cytokine levels of IL-6, IL-10 and IL-12 were detected by ELISA after 24 hours..
Results: 1) the percentage of CD11c+ DCs in the NGF group were more than that in the PBS group, and lower than that in the the GM- CSF+IL-4 group (both P<0.05). There was no difference between the GM-CSF + IL-4 group and the NGF+GM-CSF+IL-4 group (P>0.05). CD8a- DCs were dominant in these four groups; 2) NGF could further up-regulate the LPS-induced cytokine secretion from DCs, such as IL-6, IL-10, and IL-12 (all P<0.05), but NGF alone had no such effect (all P<0.05).
Conclusion: NGF can promote the murine bone-marrow cells differentiation into CD11c+ DCs, with CD8a-subset; NGF could enhance LPS-induced cytokine secretion from DCs (IL-6, IL-10 and IL-12).
目的：在体外探讨神经生长因子(nerve growth factor，NGF)在刺激小鼠骨髓来源的树突状细胞(dendritic cells，DCs)分化过程中的作用。方法：首先取C57BL/6小鼠股骨、胫骨骨髓细胞，按培养过程中干预因素不同分为磷酸盐缓冲液(phosphate buffered saline，PBS)刺激组(PBS组，n=6)、NGF刺激组(NGF组，n=6)、粒巨噬细胞集落刺激因子(granulocyte monocyte colony stimulating factor，GM-CSF)和IL-4共刺激组(GM-CSF+IL-4组，n=6)、NGF加GM-CSF和IL-4共刺激组(NGF+GM-CSF+IL-4组，n=6)，培养第7天采用流式细胞仪比较各组间CD11c+细胞阳性率和CD8a−亚型比例；然后用GM-CSF+IL-4刺激培养未成熟的DCs，经CD11c免疫磁珠阳性分选得到纯化的未成熟DCs，将其分为PBS组(n=6)、NGF组(n=6)、脂多糖(lipopolysaccharide，LPS)组(LPS组，n=6)、NGF+LPS组(n=6)，分别用PBS，NGF，LPS，NGF+LPS诱导细胞成熟，24 h后酶联免疫吸附分析法(enzyme linked immunosorbent assay，ELISA)检测上清液中IL-6，IL-10和IL-12的水平。结果：1) NGF组CD11c+DCs百分比高于PBS组，低于GM-CSF+IL-4组(均P<0.05)；GM-CSF+IL-4组与NGF+GM-CS F+IL-4组间比较，差异无统计学意义(P>0.05)，且在4组中均以CD8a−DCs为主；2)NGF可以增强LPS诱导的DCs分泌IL-6，IL-10和IL-12(均P<0.05)，而单独NGF刺激对DCs分泌IL-6，IL-10和IL-12无影响(均P>0.05)。 结论：NGF可以诱导小鼠骨髓细胞向CD11c+DCs的定向分化，并且以CD8a−亚型为主；NGF能增强LPS诱导的DC分泌IL-6，IL-10和IL-12。.