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Development of high-level expression vectors in embryonic and hematopoietic stem cells and generati of GM-CSF and IL-3 of receptor transgenic mice

胚胎干细胞和造血干细胞高水平表达载体的研制及受体转基因小鼠GM-CSF和IL-3的产生

基本信息

批准号：
04559003
负责人：
ARAI Ken-ichi
金额：
$ 11.65万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Developmental Scientific Research (B)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1994
项目状态：
已结题

项目摘要

Transgenic mice expressing the human GM-CSF (hGM-CSF) high affinity receptor were generated. Expression of the receptor is under control of the H-2L^d class I promoter, resulting in its expression on numerous hematopoietic cells, including splenocytes, thymocytes, and bone marrow cells. Murine GM-CSF (mGM-CSF) does not bind to the human receptor, therefore hGM-CSF receptor(hGMR)transgenic mice display normal phenotypes in the absence of exogenous hGM-CSF.In methylcellulose colony assays which examine bone marrow hematopoietic progenitors from hGMR transgenic mice, addition of hGM-CSF lead to the formation of multiple lineage colonies, including erythrocyte colonies whose genration normally depends on erythropoietin. In the presence of mGM-CSF,only granulocyte and macrophage colonies were generated. These data suggest that cytokine responsiveness of various myeloid progenitors is regulated at the level of receptor expression rather than through downstream signal transduction machinery. We also examined effects of hGM-CSF signalling on the differentiation of thymocytes from hGMR transgenic mice. hGM-CSF can transduce proliferation signals to thymic subsets when they express hGMR,while thymocytes does not respond to mGM-CSF.Addition of hGM-CSF to fetal thymic organ cultures inhibited thymocyte differentiation in stage-specific manner. In vivo administration of hGM-CSF into hGMR mice also showed that hGM-CSF supports myeloid lineage cell proliferation and differentiation but inhibits thymocyte differentiation.

获得了表达人GM-CSF高亲和力受体的转基因小鼠。该受体的表达受H-2L^d I类启动子的控制，导致其在包括脾细胞、胸腺细胞和骨髓细胞在内的大量造血细胞上表达。小鼠粒-巨噬细胞集落刺激因子(mGM-CSF)不与人受体结合，因此hGM-CSF受体(HGMR)转基因小鼠在没有外源hGM-CSF的情况下表现出正常的表型。在检测hGMR转基因小鼠骨髓造血祖细胞的甲基纤维素集落试验中，hGM-CSF的加入会导致多个谱系集落的形成，包括通常依赖促红细胞生成素的红细胞集落。在mGM-CSF存在的情况下，只产生粒细胞和巨噬细胞集落。这些数据表明，不同髓系祖细胞的细胞因子反应是在受体表达水平上调节的，而不是通过下游的信号转导机制。我们还检测了hGM-CSF信号对hGMR转基因小鼠胸腺细胞分化的影响。HGM-CSF在表达hGMR时可将增殖信号传导至胸腺细胞亚群，而胸腺细胞对mGM-CSF无反应。HGM-CSF体内给药也表明，hGM-CSF促进髓系细胞的增殖和分化，但抑制胸腺细胞的分化。