Ni K, O'Neill H
Division of Biochemistry and Molecular Biology, School of Life Sciences, Australian National University, Canberra ACT, Australia.
Br J Haematol. 1999 Apr;105(1):58-67.
Long-term stroma-dependent cultures from murine spleen have been previously shown to support dendritic cell (DC) development in vitro. Secondary cultures have now been established using a splenic stromal cell layer overlaid with cells from different lymphoid sites. Cells resembling DCs can be generated in vitro from unfractionated murine lymphoid cells in the absence of added growth factors. Bone marrow (BM) cultures are the most successful but some cultures have been derived from spleen and thymus. Large numbers of mononuclear cells with dendritic morphology can be generated from BM within 2 weeks and cell production in many cultures has been maintained for at least 6 months. A significant proportion of cells binds antibodies specific for DC markers. No lymphoid cells, mast cells or granulocytes are detectable in culture by antibody and histochemical staining and light and electron microscopy. As with cells generated in primary cultures, cells grown in secondary cultures are equally potent stimulators of both syngeneic and allogeneic mixed lymphocyte reactions, confirming their function as antigen-presenting cells. They are also capable of endocytosing and presenting protein antigen to the D10.G4.1 Th2 clone and to unprimed T cells. This study confirmed the presence of DC precursors in multiple lymphoid sites which can be induced to proliferate in the presence of a spleen stromal cell monolayer. The secondary culture system provides an ideal in vitro model for investigation of the development of DC from different tissue sites. It also provides a stable and continuous resource of cells for further studies on DC development.
先前已表明，来自小鼠脾脏的长期依赖基质的培养物能够在体外支持树突状细胞（DC）的发育。现在已经建立了二级培养体系，该体系使用覆盖有来自不同淋巴组织部位细胞的脾基质细胞层。在不添加生长因子的情况下，未分级的小鼠淋巴细胞能够在体外生成类似DC的细胞。骨髓（BM）培养最为成功，但也有一些培养物来源于脾脏和胸腺。两周内可从骨髓中产生大量具有树突状形态的单核细胞，许多培养物中的细胞生成已维持至少6个月。相当一部分细胞能结合针对DC标志物的特异性抗体。通过抗体、组织化学染色以及光镜和电镜检查，在培养物中未检测到淋巴细胞、肥大细胞或粒细胞。与原代培养中产生的细胞一样，二级培养中生长的细胞同样是同基因和异基因混合淋巴细胞反应的有效刺激物，这证实了它们作为抗原呈递细胞的功能。它们还能够内吞蛋白质抗原并将其呈递给D10.G4.1 Th2克隆和未致敏的T细胞。本研究证实了多个淋巴组织部位存在DC前体，这些前体在脾基质细胞单层存在的情况下可被诱导增殖。二级培养体系为研究不同组织部位DC的发育提供了理想的体外模型。它还为DC发育的进一步研究提供了稳定且持续的细胞来源。
