APO D CHOLESTEROL STORAGE AND NEURODEGENERATION

描述(调查人员摘要)：本提案旨在划定 人类神经退行性变的分子和细胞神经生物学 精神障碍，Niemann-Pick病C型(NPC)。鼻咽癌及其动物模型， 胆固醇储存障碍(CSD)小鼠已被映射到 18号染色体的昆虫体周围区域，尽管缺陷基因有 还没有被确认。另外两个神经学小鼠突变体， 共济失调(Ax)和扭转(Tw)与CSD基因座紧密连锁。一个 鼻咽癌和慢性阻塞性肺疾病的显著特征是溶酶体的异常聚集 胆固醇和其他与衰减有关的脂类 哺乳动物细胞诱导的正常内环境平衡反应 脂蛋白摄取(刺激胆固醇酯合成， 抑制从头合成胆固醇和下调低密度脂蛋白 受体活性)。调查人员已经证实， 在30kD的胆固醇结合蛋白中，载脂蛋白D是 培养的CSD星形胶质细胞缺乏，并有细胞内 一种新的46kD载脂蛋白免疫反应蛋白的保留 突变细胞的生化标记物。调查人员提议 确定这个新的46kD载脂蛋白D免疫反应物的身份和作用 CSD中的蛋白质。为了进一步研究载脂蛋白D作为一种 神经性和非神经性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 研究无细胞体系中载脂蛋白D-配体相互作用的技术 在单个活细胞中。由于载脂蛋白D是Lipocalin的成员 小分子疏水配体载体蛋白家族(例如 包括视网膜结合蛋白、β-乳球蛋白和气味结合 蛋白质)，这些研究将提供关于结构的新信息- 载脂蛋白D与胆固醇及相关配体结合的功能相关性 以及对配体运输机制的新见解 一般是通过脂钙蛋白。鼻咽癌和慢性阻塞性肺病的主要表型特征 小鼠反映神经退行性变伴髓鞘过少和选择性 小脑浦肯野细胞丢失。调查人员已经确定 分泌出强有力的有丝分裂原和生长因子，碱性成纤维细胞 培养的星形胶质细胞的碱性成纤维细胞生长因子(BFGF)缺乏。这个不足之处是 与CSD脑内少突胶质细胞成熟缺陷有关。 此外，蛋白激酶C(PKC)的表达已被证明 对髓鞘蛋白基因的表达起关键作用 少突胶质细胞的发育，在CSD脑中受到限制。这个 因此，研究人员将调查生长因子的作用。 尤其是碱性成纤维细胞生长因子和载脂蛋白D在少突胶质细胞成熟和 PKC在CSD脑髓鞘减退中的作用。最后，在确认后 关于人类鼻咽癌基因，研究人员提议克隆小鼠 人类基因的同源性，以建立其与 人类突变并确定tw和ax是否是由 类似的遗传缺陷(S)。这些研究将提供新的见解 人鼻咽癌及其小鼠模型CSD的分子缺陷 有助于更好地了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.