关键词: C57BL/6小鼠, 骨髓源CD103+DC, 分离培养, LPS, 功能特征

Abstract: In order to establish a method of inducing and culturing CD103⁺ dendritic cells (CD103⁺ DCs) from C57BL/6 mice bone marrow in vitro,and investigate the effects of LPS on their morphology and functional feature. The bone marrow cells (BMCs) were isolated and cultured in the presence of recombinant granulocyte-macrophage colony-stimulating factor (GM-CSF) and fms-like tyrosine kinase 3 ligand (FLT3L), and then observed the effect of LPS on their morphology, phenotype and function. The results showed that the modality of BMCs diversified after being cultured in GM-CSF and FLT3L for 3 d, and grew clustered-liked.As time went on, the cell colonies became larger and larger. When culturing for 13 d, the BMC colonies started separating, typical morphology with dendritic processes could be observed. A number of morphologically typical dendritic cells were observed after culturing for 15 d. The dendritic structure became more obvious after BMCs were treated with LPS for 24 h under scan electron microscope. The induced BMCs were able to express CD103 on the cell surface, and the positive rate of CD103⁺DC was over 90%. The percentage of phagocytosis of CD103⁺ DCs was 25.70% in the group of RPMI-1640, but the percentage of phagocytosis in the group of LPS dropped to 10.33%. In the group of RPMI-1640, 41.31% CD103⁺DCs expressed MHC-II, 13.79% expressed CD83. The CD103⁺DCs treated by LPS showed a higher MHC-II expression rate of 68.10%, and increase expression rate in CD83 (24.71%). MTT assay showed that the ability of CD103⁺DC treated with LPS to stimulate the proliferation of the naive allergenic T cells was stronger than those CD103⁺DC untreated with LPS. In conclusion, a large number of dendritic cells can be generated by culturing BMCs from C57BL/6 mice in vitro,and LPS shows positive effect on CD103⁺ DC mature.

Key words: C57BL/6 mice, bone marrow derived CD103+DC, isolation and culture, LPS, functional characteristics