APO D CHOLESTEROL STORAGE AND NEURODEGENERATION

描述（研究者摘要）：本提案旨在描述 人类神经退行性疾病的分子和细胞神经生物学 C型尼曼-匹克病（Niemann-Pick disease type C，NPC）鼻咽癌及其动物模型， 胆固醇储存障碍（CSD）小鼠已经被映射到 18号染色体的近着丝粒区，尽管缺陷基因具有 尚未被确认。另外两种神经学小鼠突变体， 共济失调（AX）和旋转（TW）与CSD基因座紧密连锁。一 NPC和csd的显著特征是溶酶体的异常积聚， 胆固醇和其他脂质与胆固醇和胆固醇的衰减有关 正常的稳态反应引起的哺乳动物细胞， 脂蛋白摄取（刺激胆固醇酯合成， 抑制胆固醇从头合成和下调LDL 受体活性）。调查人员已经确定， 在30 kD胆固醇结合蛋白中，载脂蛋白D是 缺乏培养的CSD星形胶质细胞， 保留一种新的46 kD载脂蛋白D免疫反应蛋白， 突变细胞的生化标记。研究人员建议 确定这种新的46 kD载脂蛋白D免疫反应性的身份和作用， CSD中的蛋白质为了进一步研究载脂蛋白D作为 神经和非神经细胞内胆固醇转运蛋白 细胞，研究人员将使用最先进的生物物理 研究无细胞系统中载脂蛋白D-配体相互作用的技术 和单个活细胞。由于载脂蛋白D是脂质运载蛋白的成员， 小疏水配体载体蛋白家族（其实例 包括视网膜结合蛋白、β-乳球蛋白和气味结合 这些研究将提供关于蛋白质结构的新信息- 载脂蛋白D与胆固醇和相关配体结合的功能相关性 并为配体转运机制提供了新的见解 由脂质运载蛋白控制NPC和CSD的主要表型特征 小鼠反映神经变性伴髓鞘形成不足， 小脑浦肯野细胞丧失。调查人员已经确定 分泌的强力丝裂原和生长因子，碱性成纤维细胞 从培养的星形胶质细胞的碱性成纤维细胞生长因子（bFGF）是缺乏的。这一缺陷是 与CSD脑中少突胶质细胞的成熟缺陷有关。 此外，蛋白激酶C（PKC）的表达已被证明是 对髓鞘蛋白基因的表达至关重要， 少突胶质细胞的发育在CSD脑中受到限制。的 因此，研究人员将研究生长因子的作用， 特别是bFGF，以及载脂蛋白D，在少突胶质细胞的成熟， PKC在CSD脑髓鞘形成不足中的作用。最后，在识别 研究人员建议克隆小鼠， 人类基因的同源物，以建立其与 人类基因突变，并确定tw和ax是否由 类似的遗传缺陷。这些研究将提供新的见解， 人NPC及其小鼠模型csd的分子缺陷，以及 从而更好地理解神经退行性变的机制 在这些遗传性疾病中。

DESCRIPTION (Investigator's Abstract): This proposal aims to delineate the molecular and cellular neurobiology of the human neurodegenerative disorder, Niemann-Pick disease type C (NPC). NPC and its animal model, the cholesterol storage disorder (csd) mouse have been mapped to the pericentomeric region of chromosome 18, although the defective gene has not yet been identified. Two additional neurological mouse mutants, the ataxic (ax) and twirler (tw) are closely linked to the csd locus. A salient feature of NPC and csd is the abnormal accumulation of lysosomal cholesterol and other lipids which is associated with an attenuation of the normal homeostatic responses elicited by mammalian cells with lipoprotein uptake (stimulation of cholesterol ester synthesis, suppression of de novo cholesterol synthesis and down-regulation of LDL receptor activity). The investigators have established that processing of the 30 kD cholesterol binding protein, apolipoprotein (apo) D is deficient in cultured csd astrocytes and there is intracellular retention of a novel 46 kD apo D-immunoreactive protein that serves as a biochemical marker of the mutant cells. The investigators propose to determine the identity and role of this novel 46 kD apo D-immunoreactive protein in csd. To further investigate the role of apo D as an intracellular cholesterol transport protein in neural and non-neural cells, the investigators will use state-of-the-art biophysical techniques to investigate apo D-ligand interactions in cell free systems and in single live cells. Since apo D is a member of the lipocalin family of small hydrophobic ligand carrier proteins (examples of which include retinal binding protein, beta-lactoglobulin and odorant binding protein), these studies will provide new information on the structure- function correlates of apo D binding to cholesterol and related ligands as well as provide new insights into the mechanisms of ligand transport by lipocalins in general. The major phenotypic features of NPC and csd mouse reflect neurodegeneration with hypomyelination and a selective loss of cerebellar Purkinje cells. The investigators have established that secretion of the potent mitogen and growth factor, basic fibroblast factor (bFGF) from cultured astrocytes is deficient. This deficiency is associated with a maturational defect of oligodendrocytes in csd brain. Furthermore, expression of protein kinase C (PKC) which has been shown to be crucial for the expression of myelin protein genes during oligodendrocyte development, is restricted in csd brain. The investigators will therefore investigate the role of growth factors especially bFGF, as well as apo D, in oligodendroglial maturation and of PKC in the hypomyelination of csd brain. Finally, upon identification of the human NPC gene, the investigators propose to clone the mouse homolog of human gene in order to establish its relationship to the human mutation and to determine whether tw and ax are caused by a similar genetic defect(s). These studies will provide new insights into the molecular defect of human NPC and its mouse model, csd, as well as lead to a better understanding of the mechanisms of neurodegeneration in these inherited disorders.