Saito H, Ebisawa M, Tachimoto H, Shichijo M, Fukagawa K, Matsumoto K, Iikura Y, Awaji T, Tsujimoto G, Yanagida M, Uzumaki H, Takahashi G, Tsuji K, Nakahata T
Division of Allergy, National Children's Medical Research Center, Tokyo, Japan.
J Immunol. 1996 Jul 1;157(1):343-50.
To establish the method for generating a large number of mature human mast cells, we cultured cord blood mononuclear cells (CBMC) in several conditions in the presence of Steel factor (SF). Among several cytokines tested, IL-6 enhanced SF-dependent mast cell growth from purified CD34+ cells for more than 8 wk in culture. When CBMC were cultured instead of CD34+ cells, IL-6 enhanced the mast cell development in the presence but not in the absence of PGE2. PGE2 enhanced the SF- and IL-6-dependent development of mast cells from CBMC probably by blocking granulocyte-macrophage CSF (GM-CSF) secretion from accessory cells, because 1) PGE2, or anti-GM-CSF enhanced the mast cell development induced by SF and IL-6 from CBMC, but not from CD34+ cells; 2) GM-CSF inhibited the enhancing effect of IL-6 on the mast cell development from CD34+ cells; and 3) PGE2 inhibited GM-CSF secretion from CBMC. The mast cells cultured in the presence of SF, IL-6, and PGE2 for >10 wk were 99% pure, and seemed to be functionally mature, because 1) they contained 5.62 micrograms of histamine and 3.46 micrograms of tryptase per 10(6) cells; and 2) when sensitized with human IgE and then challenged with anti-human IgE, the cells released a variety of mediators such as histamine, and an increase in intracellular Ca2+ was found in advance of the activation of membrane movement by using a confocal laser-scanning microscope. Electron-microscopic analysis revealed that some of the cultured mast cells are morphologically mature since they filled with scroll granules and contained crystal granules.
为建立生成大量成熟人肥大细胞的方法，我们在多种条件下于存在Steel因子（SF）的情况下培养脐血单个核细胞（CBMC）。在测试的几种细胞因子中，白细胞介素-6（IL-6）增强了纯化的CD34⁺细胞在培养中超过8周的SF依赖性肥大细胞生长。当培养CBMC而非CD34⁺细胞时，IL-6在存在前列腺素E2（PGE2）的情况下而非在其不存在时增强肥大细胞发育。PGE2可能通过阻断辅助细胞分泌粒细胞-巨噬细胞集落刺激因子（GM-CSF）来增强CBMC来源的肥大细胞的SF和IL-6依赖性发育，因为：1）PGE2或抗GM-CSF增强了CBMC而非CD34⁺细胞由SF和IL-6诱导的肥大细胞发育；2）GM-CSF抑制了IL-6对CD34⁺细胞来源的肥大细胞发育的增强作用；3）PGE2抑制了CBMC分泌GM-CSF。在存在SF、IL-6和PGE2的情况下培养超过10周的肥大细胞纯度为99%，且似乎在功能上成熟，因为：1）每10⁶个细胞含有5.62微克组胺和3.46微克类胰蛋白酶；2）用人IgE致敏然后用抗人IgE攻击时，这些细胞释放多种介质如组胺，并且使用共聚焦激光扫描显微镜发现在膜运动激活之前细胞内Ca²⁺增加。电子显微镜分析显示，一些培养的肥大细胞在形态上成熟，因为它们充满了卷轴状颗粒并含有晶体颗粒。
