EXPRESSION OF MAC-2 IN IMPLANTATION SITES OF MICE

MAC-2 在小鼠植入部位的表达

• 批准号: 2202182
• 负责人: HARRY M WEITLAUF
• 金额: $ 13.96万
• 依托单位: TEXAS TECH UNIVERSITY HEALTH SCIS CENTER
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-12-01 至 1997-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2202182
• 关键词: antisense nucleic acid; cell differentiation; decidua; early embryonic stage; embryo /fetus culture; embryo implantation; galactosides; gene expression; immunocytochemistry; immunoprecipitation; in situ hybridization; laboratory mouse; laboratory rabbit; lectin; ligands; messenger RNA; monoclonal antibody; neutralizing antibody; passive immunization; placenta; protein biosynthesis; protein purification; protein structure function; trophoblast; uterus

It has recently been shown that a soluble beta-galactoside specific lectin (Mac-2) is expressed on mouse blastocysts just prior to the time of embryo attachment to the uterine epithelium and that it accumulates in large amounts at the feto-maternal interface within developing implantation sites. Because Mac-2 has been associated with a variety of important functions including cell-recognition/adhesion and local immunomodulation in other cellular environments, its localization to the region of contact between the implanting conceptus and its mother led us to undertake experiments to establish the ontogeny of this lectin in the implantation site as well as to determine what role it plays in pregnancy. The spatio-temporal pattern of expression of Mac-2 in early implantation sites and deciduomata will be established by immunohistochemistry using the anti-Mac-2 monoclonal antibody (M3/38) on frozen sections of the mouse uterus at various stages of implantation and placental development. Because Mac-2 is a soluble extracellular protein that binds to the cell surface by means of its carbohydrate binding site, it will be important to compare the spatio-temporal pattern of expression of the protein with that for its mRNA. That will be done by means of in situ hybridization histochemistry using a labeled antisense mRNA probe on frozen sections of the uterus adjacent to those used to examine distribution of the protein. Additional studies will be undertaken to resolve the functional role of this soluble lectin in the processes of implantation and placentation. First, a monospecific function perturbing polyclonal antiserum against Mac-2 will be used to determine whether immunoneutralization of the lectin interferes with attachment and trophoblastic outgrowth in vitro or perturbs the processes of implantation or placentation in vivo. And second, the endogenous uterine ligands for Mac-2 will be isolated by means of immunoprecipitation (again with M3/38) and a monospecific function perturbing polyclonal antiserum will be prepared in a rabbit against the purified ligands(s). If that effort proves to be successful, the resulting antiserum will be used both for immunocytochemistry to establish the spatio-temporal pattern of expression of the Mac-2 ligand in the implantation site for comparison with that for the lectin and its mRNA, and for passive immunization of pregnant females in hopes of perturbing ligand function in situ during implantation and placentation.

最近已经显示出可溶性β-半乳糖苷特异性凝集素 （MAC-2）在胚胎之前在小鼠胚泡上表达 附着子宫上皮，并积聚 发育植入的feto母亲界面的量 站点。因为Mac-2与各种重要 包括细胞识别/粘附和局部免疫调节的功能 在其他细胞环境中，其定位到接触区域 在植入的概念和母亲之间，使我们承担了 实验以在植入中建立该凝集素的个体发育 站点以及确定其在怀孕中的作用。 MAC-2在早期植入中表达的时空表达模式 将使用免疫组织化学建立位点和deciduomata 小鼠冷冻切片上的抗MAC-2单克隆抗体（M3/38） 子宫处于植入和胎盘发育的各个阶段。 因为MAC-2是一种结合细胞的可溶细胞外蛋白 通过其碳水化合物结合位点表面，对 比较蛋白质的表达时空模式 因为它的mRNA。这将通过原位杂交来完成 在冷冻切片上使用标记的反义mRNA探针的组织​​化学 子宫与用于检查蛋白质分布的子宫相邻。 将进行其他研究以解决 该可溶性凝集素在植入和胎盘的过程中。 首先，单型功能会扰动多克隆抗血清反对 MAC-2将用于确定凝集素的免疫中性化是否 干扰体外的附着和滋养细胞的生长或 在体内植入或胎盘的过程。和 其次，将通过均值隔离用于MAC-2的内源性子宫配体 免疫沉淀（再次带有M3/38）和一个单特异性功能 扰动多克隆抗血清将在兔子中准备 纯化的配体。如果这项努力被证明是成功的，那么 抗血清将用于免疫细胞化学来确定 MAC-2配体表达的时空模式 与凝集素及其mRNA相比的植入部位， 并为怀孕的女性进行被动免疫，希望扰动 配体功能在植入和胎盘期间原位。