VITAMIN A AND REPRODUCTION

维生素 A 与生殖

• 批准号: 6520846
• 负责人: DAVID E ONG
• 金额: $ 23.8万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-09-30 至 2005-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6520846
• 关键词: chemical binding; chloramphenicol acetyltransferase; differential display technique; estrogens; estrus; female; gene expression; gene induction /repression; genetic promoter element; hormone regulation /control mechanism; in situ hybridization; laboratory rat; molecular cloning; reporter genes; retinoate; retinoid binding proteins; subtraction hybridization; tissue /cell culture; uterus; vitamin biosynthesis; vitamin metabolism

Vitamin A, retinol, is an essential nutrient that serves as precursor to the important hormone, retinoic acid (RA). Relatively little is known of the control of synthesis of RA from retinol in the normal, fully-developed animal and sites of action of RA are inferred, rather than demonstrated. Previous work has identified estrogen as a physiological signal which induces the synthesis of RA in the rat uterus and that coordinately directs cell-specific expression of the three cellular retinoid-binding proteins present in the uterus during the estrous cycle. Proposed studies will: l) Identify uterine genes that are under estrogen control indirectly, via RA stimulation. The techniques of differential display or subtractive hybridization followed by library screening or will be used to identify these genes. Candidate genes will be followed during the estrous cycle to confirm their physiological significance. 2) Demonstrate the site(s) of expression of the estrogen-stimulated RA responsive genes by in situ hybridization and immunolocalization during the estrous cycle. Demonstration of expression/non-expression of candidate genes in cells expressing cellular retinoic acid-binding protein will test the competing hypotheses that this protein either blocks or enhances the RA responsiveness of cells. 3) Establish the mechanism by which estrogen directly regulates cellular retinoic-acid binding protein (II) expression in the uterus. The promoter region of the rat gene will be cloned, dissected and tested using CAT reporter constructs in an estrogen responsive cell line. 4) Demonstrate the mechanism by which estrogen induces RA synthesis in the uterus. Specifically, is this induction a direct effect of estrogen on pre-existing enzymes, does it require transcription, or is it indirect? A novel radioreceptor assay capable of detecting small amounts of RA has been developed for this aim. In summary, the work to be accomplished here will allow dissection of the effects of the demonstrated estrogen-stimulated synthesis of RA signal that is part of a normal physiological process. This will provide important information on retinoic acid action in the unmanipulated, intact animal. Regulation of retinoic acid production by estrogen has direct importance for understanding/treating conditions such as endometriosis, breast cancer, and cancers of the female reproductive system.

维生素A，视黄醇，是一种必需的营养素，作为前体的重要激素，视黄酸（RA）。相对较少的是已知的控制合成的RA从视黄醇在正常的，完全发达的动物和网站的作用RA的推断，而不是证明。以前的工作已经确定雌激素作为一种生理信号，诱导合成RA在大鼠子宫和协调指导细胞特异性表达的三个细胞维甲酸结合蛋白存在于子宫在发情周期。提出的研究将：1）确定子宫基因，雌激素控制下间接，通过RA刺激。 差异显示技术或消减杂交技术以及文库筛选技术将被用来鉴定这些基因。候选基因将在发情周期中进行跟踪，以确认其生理意义。 2)通过原位杂交和免疫定位证明动情周期中雌激素刺激的RA应答基因的表达位点。在表达细胞视黄酸结合蛋白的细胞中候选基因的表达/不表达的证明将测试该蛋白阻断或增强细胞的RA反应性的竞争性假设。3)建立雌激素直接调节子宫中细胞视黄酸结合蛋白（II）表达的机制。大鼠基因的启动子区将被克隆、解剖并在雌激素应答细胞系中使用CAT报告基因构建体进行测试。4)证明雌激素诱导子宫内RA合成的机制。具体来说，这种诱导是雌激素对预先存在的酶的直接影响，它需要转录，还是间接的？一种新的放射受体测定能够检测少量的RA已开发用于此目的。总之，这里要完成的工作将允许解剖的影响，证明雌激素刺激合成的RA信号，这是一个正常的生理过程的一部分。这将为未操作的完整动物提供有关维甲酸作用的重要信息。雌激素调节视黄酸的产生对于理解/治疗子宫内膜异位症、乳腺癌和女性生殖系统癌症等疾病具有直接的重要性。