GENERATION OF AN INDUCIBLE SYSTEM IN THE UTERINE STROMA FOR IMPLANTATION STUDIES

用于植入研究的子宫间质中诱导系统的生成

• 批准号: 8358586
• 负责人: Liang Ma
• 金额: $ 23.64万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-03 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8358586
• 关键词: Accounting; Alleles; Binding; Chimera organism; Complex; Decidual Cell Reactions; Development; Doxycycline; Embryo; Embryo Loss; Embryonic Development; Event; Failure; Fertilization in Vitro; Gene Deletion; Gene Expression; Gene Targeting; Generations; Genes; Genetic; Genetic Recombination; Growth Factor; Hormonal; Human; Implant; Infertility; Knock-in Mouse; Knock-out; Knowledge; Lead; Mediating; Molecular; Mouse Strains; Mus; Nature; Organogenesis; Partner in relationship; Pathway interactions; Pharmaceutical Preparations; Phase; Placentation; Pregnancy; Procedures; Process; Production; Reporter; Research; Research Personnel; Southern Blotting; System; Technology; Tetracyclines; Tissues; Uterus; Validation; blastocyst; carcinogenesis; design; embryonic stem cell; failure Implantation; field study; homologous recombination; implantation; improved; intercellular communication; interest; natural Blastocyst Implantation; overexpression; postnatal; reproductive; tool; uterine receptivity

DESCRIPTION (provided by applicant): Human infertility is a global problem and failure of embryo implantation accounts for a significant percentage of pregnancy failure during both natural pregnancy and in vitro fertilization procedures. Implantation is an extremely complicated process requiring precisely controlled hormonal, growth factor signaling and cell-cell contacts which coordinate interactions between the competent blastocysts and the receptive uterus. In the past two decades we have greatly improved our knowledge on this subject by using sophisticated mouse genetics. However, our understanding of implantation is still rudimentary. Implantation research including the study of uterine receptivity and decidualization has benefited greatly from the Cre/LoxP technology which allows functional study of many genes in a tissue specific manner. Although inducible Cre/LoxP systems have been widely used in other fields of studies, their use in implantation studies has been limited by the steroidal nature of most of the inducers which interferes with the implantation process. The currently used Pgr-Cre and Amhr2-Cre lines are not inducible and each has their own limitations for implantation studies. Thus there is urgent need to develop an inducible tissue-specific Cre system for conditional deletion of genes in the uterus during implantation. In this proposal, we propose to knock rtTA into the endogenous Gli2 locus to generate a tetracycline-inducible line which in corporation with tetO-Cre can drive inducible Cre expression in the uterine stroma during implantation. In Aim I we will use BAC recombineering to generate a knock-in construct which will be used for gene targeting in ES cells and eventually germline chimera production. In Aim II, we will assess the usefulness of this knock-in allele in implantation studies. This mouse strain should be a valuable tool for researchers studying implantation as well as embryonic or postnatal organogenesis and carcinogenesis. PUBLIC HEALTH RELEVANCE: This project proposes to generate a tissue-specific inducible Cre system in the uterine stroma which can be used to knockout or overexpress gene of interest in the peri-implantation uterine stroma. This strain of mice will be an invaluable tool for reproductive biologists studying uterine receptivity, decidualization and placentation.

描述（由申请人提供）：人类不育是一个全球性问题，胚胎植入失败占自然妊娠和体外受精过程中妊娠失败的很大比例。着床是一个极其复杂的过程，需要精确控制激素、生长因子信号和细胞-细胞接触，以协调有能力的囊胚和接受子宫之间的相互作用。在过去的二十年里，我们通过使用复杂的小鼠遗传学极大地提高了我们在这个问题上的知识。然而，我们对植入的理解仍然是初步的。包括子宫容受性和蜕膜化研究在内的着床研究极大地受益于Cre/LoxP技术，该技术允许以组织特异性方式对许多基因进行功能研究。虽然诱导型Cre/LoxP系统已广泛用于其他研究领域，但它们在植入研究中的使用受到大多数诱导剂的甾体性质的限制，这会干扰植入过程。目前使用的Pgr-Cre和Amhr 2-Cre系是不可诱导的，并且对于植入研究各自具有其自身的局限性。因此，迫切需要开发一种可诱导的组织特异性Cre系统，用于在着床期间有条件地删除子宫中的基因。在这个提议中，我们建议敲入内源性Gli 2基因座的rtTA产生四环素诱导系，其与tetO-Cre的公司可以驱动诱导型Cre在植入过程中在子宫基质中的表达。在目标I中，我们将使用BAC重组工程产生敲入构建体，其将用于ES细胞中的基因靶向并最终用于生殖系嵌合体的产生。在目的II中，我们将评估这种敲入等位基因在植入研究中的有用性。这种小鼠品系应该是研究植入以及胚胎或出生后器官发生和致癌作用的研究人员的宝贵工具。 公共卫生相关性：本项目拟在子宫间质中构建组织特异性的可诱导Cre系统，用于在着床期子宫间质中敲除或过表达目的基因。这种小鼠品系将成为生殖生物学家研究子宫容受性、蜕膜化和胎盘形成的宝贵工具。