INHERITED HYPOTHALAMIC NEURODEGENERATION

遗传性下丘脑神经变性

基本信息

批准号：
2734236
负责人：
DAVID R REPASKE
金额：
$ 12.77万
依托单位：
CINCINNATI CHILDRENS HOSP MED CTR
依托单位国家：
美国
项目类别：
财政年份：
1997
资助国家：
美国
起止时间：
1997-09-30 至 2002-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2734236
关键词：
PC12 cells arginine vasopressin autosomal dominant trait diabetes insipidus gene expression gene mutation gene targeting genetic models genetically modified animals histology hypothalamus immunocytochemistry immunoelectron microscopy laboratory mouse neural degeneration neurogenetics neurophysins peptide hormone biosynthesis

项目摘要

Autosomal dominant neurohypophyseal diabetes insipidus (ADNDI) is an inherited endocrine disorder caused by a progressive and specific degeneration of the magnocellular neurons that produce the hormone, vasopressin. The genetic locus of ADNDI is the gene that encodes arginine vasopressin (AVP) and the AVP binding protein, neurophysin II (NPII), and three distinct classes of mutations that cause the disease have been defined. The overall goals of this grant are to investigate the molecular basis for this disease, specifically addressing the basis of dominant inheritance and the molecular and cellular mechanisms by which the mutations cause neurodegeneration. The hypothesis is that dominant negative mutations in the AVP-NPII gene cause ADNDI, and that the mutant alleles encode an AVP-NPII precursor that is misprocessed, accumulates, and causes degeneration of the magnocellular neurons. Specific aim 1 of the proposal is to test the hypothesis that mutant AVP-NPII alleles exert dominant negative effects. This will be tested by creation of transgenic mouse models via introduction of mutant human AVP-NPII genes. If features of ADNDI do not occur in these transgenic animals, the alternative hypothesis, that ADNDI mutations cause the disease by eliminating one of the two functional AVP-NPII alleles, will be tested in another mouse model by deleting one normal AVP-NPII allele using homologous recombination. Specific aim 2 is to test the hypothesis that mutations that cause ADNDI alter the preproAVP-NPII precursor to disrupt proper processing and allow a toxic accumulation of misprocessed precursor. The hypothalami of the mouse models for ADNDI will be evaluated by histology, immunohistochemistry and electron microscopy to investigate for magnocellular neuron degeneration. Expression of cloned mutant and normal human AVP-NPII cDNAs in PC12 cells and in vitro transcription, translation, and processing by microsomal membranes will be employed to allow detailed study of altered cell biology and biochemistry in the presence of ADNDI mutations. These experiments will define the genetic, molecular, and cellular pathophysiology of ADNDI and contribute to knowledge of other degenerative neurological diseases.

常染色体显性神经垂体尿崩症 (ADNDI) 是一种由进行性和特异性的遗传性内分泌紊乱引起的产生激素的大细胞神经元退化，加压素。 ADNDI 的遗传位点是编码精氨酸加压素 (AVP) 和 AVP 结合蛋白、神经素 II (NPII)，以及导致该疾病的三类不同类型的突变已被定义。这笔赠款的总体目标是调查这种疾病的分子基础，特别是解决显性遗传以及分子和细胞机制这些突变会导致神经退行性变。假设是 AVP-NPII 基因的显性失活突变会导致 ADNDI，并且突变等位基因编码一个被错误处理的 AVP-NPII 前体，积累并导致大细胞神经元变性。该提案的具体目标 1 是检验突变体的假设 AVP-NPII 等位基因发挥显着的负面影响。这将通过以下方式进行测试通过引入突变人类创建转基因小鼠模型 AVP-NPII 基因。如果这些转基因中没有出现 ADNDI 的特征动物，另类假设，ADNDI 突变导致通过消除两个功能性 AVP-NPII 等位基因之一来治疗疾病，将通过删除一个正常的 AVP-NPII 等位基因在另一种小鼠模型中进行测试使用同源重组。具体目标2是检验假设导致 ADNDI 的突变改变了 preproAVP-NPII 前体破坏正常的加工过程并导致有毒物质的积累处理不当的前体。 ADNDI 小鼠下丘脑模型将通过组织学、免疫组织化学和电子学进行评估显微镜检查大细胞神经元变性。克隆突变体和正常人 AVP-NPII cDNA 在 PC12 中的表达细胞和体外转录、翻译和加工将采用微粒体膜来详细研究改变的 ADNDI 突变存在时的细胞生物学和生物化学。这些实验将定义遗传、分子和细胞 ADNDI 的病理生理学并有助于其他知识退行性神经系统疾病。