Cytokine productions from murine and human mast cells

小鼠和人类肥大细胞产生的细胞因子

基本信息

批准号：
05454285
负责人：
NAKAHATA Tatsutoshi
金额：
$ 3.33万
依托单位：
The University of Tokyo (1994)Shinshu University (1993)
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1994
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05454285/
关键词：
murine mast cells human cultured mast cell production of cytokines mRNA CD34+cells stem cell factor cross-linking of FcepsilonRI 大量培養 CD34^+細胞 BMMC 肥満細胞の大量培養インターロイキン-3(IL-3)stem cell factor(SCF)

项目摘要

To establish a large scale culture method for murine and human mast cells, we have examined the effects of various cytokines on the development of mast cells from murine bone marrow cells and human cord blood CD34^+ cells. In murine, the addition of interleukin-3 (IL-3) to cultures of bone marrow cells gives rise to pure cultures of mast cells (bone marrow-derived maust cells : BMMC). Addition of stem cell factor (SCF), IL-4 and IL-10 to culture with IL-3 stimulated the development of BMMC.Although stem cell factor (SCF), IL-4 and IL-10 alone failed to support BMMC development, SCF in combinations with IL-4 or IL-10 significantly stimulated the proliferation of BMMC.In contrast with BMMC,IL-3 alone could not support the proliferation of CTMC,which were purified from mouse peritoneal cells, as well as IL-4 or IL-10. SCF alone supported a small number of colonies from CTMC.IL-3 and IL-4 but not IL-10 enhanced SCF-dependent CTMC proliferation IL-3 stimulated CTMC proliferation in the pres … More ence of IL-4 or IL-10.In human, we obtained both tryptase-positive and chymase-positive mast cells by culturing CD34^+ cells, which were isolated from cord blood mononuclear cells using a cell sortor, in the presense of SCF and IL-6 or IL-11. When CD34^+ cells were cultured with SCF and IL-6 or IL-11, mast cells with tryptase-positive granules were first detected after 2 weeks of culture, and reached to almost 100% purity after 100 days of culture when 20-30% of the cells were immunohistologically stained for chymase. Our cultured human mast cells contained a large amount of histamine and tryptase, and expressed functional IgE receptors on their cell surface. Single cell culture of CD34^+ cells with a clone-sorting method indicated that both tryptase-positive and chymase-positive mast cells were originated from a common hemopoietic progenitor. Phenotipical analyzes of human mast cell progenitors using a clone-sorting system indicated that committed ones were CD34^+CD38^+ and uncommitted ones were CD34^+CD38^-.Using reverse transcriptase (RT)-PCR,we demonstrated that murine BMMC and cultured CTMC,and cultured human mast cells, which were pretreated with IgE and anti-IgE monoclonal antibodies, express mRNAs of TNF,GM-CSF,IL-3, IL-4, IL-6 and IL-13. These results suggest that both murine and human mast cells can release not only a variety of chemical mediators such as histamine and leukotorienes but also various cytokines. Less

为了建立小鼠和人类肥大细胞的大规模培养方法，我们研究了各种细胞因子对小鼠骨髓细胞和人类脐带血CD34^+细胞发育肥大细胞的影响。将干细胞因子 (SCF)、IL-4 和 IL-10 添加到骨髓细胞培养物中，产生肥大细胞的纯培养物（骨髓来源的 maust 细胞：BMMC）。 IL-3刺激BMMC的发育。虽然单独的干细胞因子(SCF)、IL-4和IL-10不能支持BMMC的发育，但SCF与IL-4或IL-10联合显着刺激BMMC的增殖。与BMMC相比，单独的IL-3不能支持从小鼠腹膜细胞中纯化的CTMC的增殖，而单独的IL-4或IL-10则支持少量的集落。 CTMC.IL-3 和 IL-4 但不包括 IL-10 增强 SCF 依赖性 CTMC 增殖 IL-3 在 IL-4 或 IL-10 存在的情况下刺激 CTMC 增殖。在人类中，我们获得了类胰蛋白酶阳性在存在 SCF 和 IL-6 或 IL-11 的情况下，通过培养 CD34^+ 细胞（使用细胞分选仪从脐带血单核细胞分离）来分离食糜酶阳性肥大细胞。 CD34^+细胞与SCF和IL-6或IL-11一起培养，培养2周后首次检测到带有类胰蛋白酶阳性颗粒的肥大细胞，培养100天后纯度接近100%（当20-30%时）我们培养的人类肥大细胞含有大量的组胺和类胰蛋白酶，并在其细胞表面表达功能性 IgE 受体。使用克隆分选方法进行的 CD34^+ 细胞的单细胞培养表明，类胰蛋白酶阳性和糜酶阳性肥大细胞均源自人类肥大细胞祖细胞的共同造血祖细胞，使用克隆分选系统表明定型。那些是CD34^+CD38^+，未承诺的是CD34^+CD38^-。使用逆转录酶（RT）-PCR，我们证明了小鼠BMMC和培养的CTMC以及经IgE和抗IgE单克隆抗体预处理的培养的人肥大细胞均表达TNF、GM-CSF、IL-3、IL-4、IL-6和IL-13的mRNA。小鼠和人类的肥大细胞不仅可以释放多种化学介质，例如组胺和白三烯，还可以释放多种细胞因子。