MANIPULATING MHC GENE EXPRESSION IN ES CELL CHIMERAS

这项研究的主要目标是从发展的角度分析 主要组织相容性I类产物的调节表达 复杂的老鼠。特别是，控制MHG的可能性 基因表达可能在母体对胎儿的耐受性中起关键作用 将检查同种异体移植物。我们设计了一个策略 研究H-2D δ异位表达的后果， 移植抗原在发育中的胚胎。一个承载着 与H-2D编码区融合的人β-肌动蛋白基因启动子 δ基因已被引入多能胚胎干（ES）细胞中。 虽然H-2D δ蛋白在细胞表面没有表达，但在细胞内没有表达。 未分化的ES细胞，大量的H-2D δ膜 在体外诱导分化的ES细胞上检测到糖蛋白。一 拟议调查的主要目的是确定这些 转染的ES细胞系可以贡献于所有的体细胞组织， 植入正常胚胎。我们是否应该努力获得活产 如果嵌合体成功，下一个目标将是确定 引入的H-2D Delta基因可以在生殖系中传播。应该 转基因嵌合体未能发展到术语，我们将描述 发育失败，并确定异常表型是否 可归因于特定细胞谱系的缺陷。的空间和时间 将检测I类抗原在发育胚胎中的表达 使用免疫组织化学和原位杂交技术。我们将 研究异位MHC基因表达的可能性， 胚胎谱系可能会引发针对 胚胎移植一个特别重要的问题是， 母体淋巴细胞浸润并破坏异常胚胎。这些 实验将有希望导致更清楚地了解调节 小鼠早期发育过程中MHC基因的表达及相关过程 母亲和胎儿的互动。

The broad objective of this research is to analyse developmentally regulated expression of Class I products of the Major Histocompatability Complex in the mouse. In particular, the possibility that control of MHG gene expression may play a key role in maternal tolerance of the fetal allograft will be examined. We have designed a strategy that will allow us to study the consequences of ectopic expression of H-2D delta transplantation antigens in the developing embryo. A construct carrying the human Beta-actin gene promoter fused to the coding regions of the H-2D delta gene has been introduced into pluripotent embryonic stem (ES) cells. Although H-2D delta protein was not expressed at the cell surface in undifferentiated ES cells, significant amounts of H-2D delta membrane glycoproteins are detected on ES cells induced to differentiate in vitro. A major aim of the proposed investigation is to determine whether these transfected ES cell lines can contribute to all the somatic tissues when incorporated into normal embryos. Should our efforts to obtain live born chimeras be successful, the next goal will be to determine whether the introduced H-2D Delta gene can be transmitted in the germ line. Should transgenic chimeras fail to develop to term, we will describe the developmental failure and determine whether the abnormal phenotype (s) is attributable to defects in specific cell lineages. The spatial and temporal expression of Class I antigens in the developing embryos will be examined using immunohistochemical and in situ hybridization techniques. We will investigate the possibility that ectopic MHC gene expression in the extra- embryonic lineages may trigger a maternal immune response directed against the fetal allograft. A particularly important point will be to test whether maternal lymphocytes infiltrate and disrupt the abnormal embryos. These experiments will hopefully lead to a clearer understanding regulation of MHC gene expression during early mouse development and processes involved in maternal-fetal interactions.