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PLACENTAL AMINO ACID TRANSPORT: MOLECULAR ASPECTS

胎盘氨基酸转运：分子方面

基本信息

批准号：
2025200
负责人：
VADIVEL GANAPATHY
金额：
$ 14.87万
依托单位：
AUGUSTA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1989
资助国家：
美国
起止时间：
1989-04-01 至 1998-11-30
项目状态：
已结题

项目摘要

The primary objective of the project is to obtain a clear understanding of the mechanisms involved in the transport of a unique amino acid, taurine, across the human placenta. Taurine, though a non-protein amino acid, is the most abundant free amino in several tissues (e.g. brain, heart, and retina) of the human fetus. In animals, induction of taurine deficiency in the mother during pregnancy produces deleterious effects on the developing fetus, causing functional impairment of those organs which accumulate high levels of taurine. The effects of taurine deficiency in utero include retinal degeneration, cardiac myopathy and impairment of cerebellar development. These studies indicate that taurine plays an important role in the growth and development of the fetus. Paradoxically however, neither the human fetus nor the human placenta can synthesize taurine endogenously. Maternal-to-fetal transfer across the placenta is the only mechanisms available for the fetus to obtain this important nutrient. A model is proposed for the transport of taurine across the placenta according to which the syncytiotrophoblast expresses two similar, yet distinct, taurine transporters, one in the maternal-facing brush border membrane and the other in the fetal-facing basal membrane and a functional coordination between these two transporters bring about the transfer of taurine from mother to fetus. The project seeks to obtain supporting evidence for this model, by establishing the molecular and biochemical identity of these two transporters. This goal will be achieved by isolating the cDNAs encoding these transporters from a human placental cDNA library and by functionally expressing the cloned transporters in mammalian cells and in Xenopus laevis oocytes. Polyclonal and monoclonal antibodies against the cloned transport proteins will be generated to be used in immunohistochemical studies to establish the polarized localization of the two taurine transporters in the syncytiotrophoblast. Characterization of taurine transport across the brush border and basal membranes will also be done with purified membrane vesicles from normal placentas. These studies will be complemented with an intact cell system in which placental choriocarcinoma cells will be grown on permeable filters allowing access to the brush border and basolateral membranes for differential assessment and characterization of taurine transport across these membranes. Another goal of the project is to investigate the hormonal regulation of the placental taurine transporter(s) and the cellular mechanisms involved therein. It is proposed that the transporter is regulated by posttranslational modification involving phosphorylation and that protein kinase C and calcineurin (a Ca2+- calmodulin-dependent phosphoprotein phosphatase) participate in this process. The taurine transporter cDNA(s) and the antitransporter antibodies will provide excellent experimental tools for these studies. This project will result in a thorough understanding of the molecular and cellular mechanisms responsible for the maternal-to-fetal transfer of taurine across the human placenta.

该项目的主要目标是获得一个清晰的认识，一种独特氨基酸的运输机制，牛磺酸穿过人类胎盘牛磺酸虽然是一种非蛋白质氨基酸是几种组织（例如脑，心脏和视网膜）。在动物中，牛磺酸的诱导母亲在怀孕期间的缺乏会产生有害的影响对发育中的胎儿造成功能性损伤它们积累了大量的牛磺酸牛磺酸的作用子宫内缺乏症包括视网膜变性、心肌病和小脑发育障碍。这些研究表明牛磺酸在生长发育中起着重要的作用，胎儿然而，奇怪的是，无论是人类胎儿还是人类胎盘可以内源性合成牛磺酸。母胎移植是胎儿唯一可以利用的机制获得这种重要的营养素。提出了一个模型的运输牛磺酸穿过胎盘的能力表达两种相似但不同的牛磺酸转运蛋白，其中一种在一个位于母侧刷状缘膜，另一个位于胎儿侧刷状缘膜基底膜和这两者之间的功能协调转运蛋白使牛磺酸从母体转移到胎儿。该项目旨在通过以下方式获得该模型的支持证据：确定这两种病毒的分子和生化特性运输机这一目标将通过分离编码这些转运蛋白来自人胎盘cDNA文库，在哺乳动物细胞中功能性表达克隆的转运蛋白，非洲爪蟾卵母细胞中。多克隆和单克隆抗体将产生克隆的转运蛋白，免疫组织化学研究，以建立极化定位合胞体滋养层中的两种牛磺酸转运蛋白。牛磺酸跨刷状缘和基底膜转运的特征膜也将用来自正常人的纯化的膜囊泡来完成。胎盘这些研究将补充一个完整的细胞系统其中胎盘绒毛膜癌细胞将生长在可渗透的过滤器允许进入刷状缘和基底外侧膜，牛磺酸跨膜转运的差异评估和表征这些膜。该项目的另一个目标是调查胎盘牛磺酸转运蛋白的激素调节和其中涉及的细胞机制。拟议将转运蛋白受翻译后修饰的调节，磷酸化和蛋白激酶C和钙调神经磷酸酶（Ca2 +- 钙调蛋白依赖性磷蛋白磷酸酶）参与了这一过程过程牛磺酸转运蛋白cDNA和反转运蛋白抗体将为这些研究提供极好的实验工具。该项目将导致对分子和负责母胎转移的细胞机制牛磺酸穿过人类胎盘