ALZHEIMERS ABETA, APOLIPOPROTEINS & BLOOD-BRAIN BARRIER

阿尔茨海默病 ABETA，载脂蛋白

• 批准号: 2735675
• 负责人: Berislav V Zlokovic
• 金额: $ 27.53万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-01 至 2000-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2735675
• 关键词: Alzheimer's disease; Baculoviridae; aging; amyloid proteins; apolipoprotein E; blood brain barrier; guinea pigs; high density lipoproteins; high performance liquid chromatography; immunocytochemistry; intermolecular interaction; membrane transport proteins; protein isoforms; protein sequence; recombinant proteins; synthetic peptide; very low density lipoprotein; western blottings

Amyloid beta (Abeta) is a fibrillar component of amyloid in senile plaques and cerebral blood vessels in Alzheimer's Disease (AD), and a soluble peptide (sAbeta) normally present in body fluids. The origin of Abeta deposited in brain and blood vessels in AD is uncertain, and whether or not sAbeta is the direct precursor of amyloid it is not known. The current concept states that a key question in amyloidogenesis is to determine what factors are responsible for the conformational alteration of sAbeta into its fibrillar form. The unifying hypothesis of this proposal is that the blood-brain barrier (BBB) may play an important role in cerebrovascular and brain amyloidogenesis by regulating transport of sAbeta and two amyloid-associated proteins shown to bind sAbeta, apolipoproteins E (apo E) and J (apo J). In support of this hypothesis we have recently provided the evidence that a synthetic peptide homologous to major form of circulating sAbeta, sAbeta(1-40), is taken up into the brain via specific mechanism, and possibly as an sAbeta-apo J complex. In contrast, blood- brain transport of sAbeta-apo E was negligible. Cerebrovascular permeability to apo J was significant, while apo E had a limited access across the BBB, indicating that the apo E found within the brain is produced locally. Our pilot in vitro experiments demonstrated that sAbeta(1-40) binds to apo J with the affinity that is about 10 times higher than for apo E. Studies conducted in vivo indicated that sAbeta(1- 40) binding to apo J and apo E results in remarkable 114-fold difference in barrier permeability to their respective complexes. In the present proposal, we plan to characterize the mechanisms of sAbeta transport at the BBB in greater detail. Specific hypotheses to be tested are: 1. Binding of sAbeta to apo J and apo E isoforms 2, 3 and 4 modulates cerebrovascular permeability to sAbeta; 2. sAbeta peptides are transported across the BBB by a specific transporter and/or receptor; 3. Transport of sAbeta complexed to apo J and apo E isoforms incorporated into high and very low density lipoprotein particles, i.e., HDL and VLDL, may be mediated by HDL and/or low density lipoprotein (LDL) BBB receptors; 4. Aging alters transport mechanisms at the BBB predisposing to accumulation of sAbeta and amyloid-associated proteins in cerebral microvessels and brain parenchyma. The guinea-pig with the Abeta sequence identical to humans will be used. The affinity of sAbeta binding to apolipoproteins will be determined in vitro. Transport studies will be performed in the in situ vascular perfused brain, in conjunction with the capillary depletion technique, radio-HPLC end immunocytochemical analysis. The putative sAbeta transporter and/or receptor will be isolated from BBB plasma membranes. Proposed studies should help us understand transport mechanisms of sAbeta at the BBB, with an ultimate goal to develop strategies that may reduce accumulation of sAbeta and amyloid-associated proteins in brain and cerebral microvessels, thereby decelerating and/or preventing conformational transformation of sAbeta and its potential cytotoxic effects.

β淀粉样蛋白（Abeta）是老年斑中淀粉样蛋白的纤维成分 和阿尔茨海默病 (AD) 中的脑血管，以及可溶性 肽（sAbeta）通常存在于体液中。阿贝塔的由来 AD 中大脑和血管中的沉积尚不确定，是否或 sAbeta 不是淀粉样蛋白的直接前体，目前尚不清楚。目前的 概念指出淀粉样蛋白生成的一个关键问题是确定什么 因素导致 sAbeta 构象改变为 它的纤维形式。该提案的统一假设是 血脑屏障（BBB）可能在脑血管疾病中发挥重要作用 通过调节 sAbeta 和两种物质的运输来调节脑淀粉样蛋白生成 淀粉样蛋白相关蛋白可结合 sAbeta、载脂蛋白 E (apo E) 和 J (apo J)。为了支持这一假设，我们最近提供了 有证据表明合成肽与主要形式同源 循环的 sAbeta，sAbeta(1-40)，通过特定的 机制，并且可能作为 sAbeta-apo J 复合物。相比之下，血液—— sAbeta-apo E 的脑转运可以忽略不计。脑血管 apo J 的通透性显着，而 apo E 的通透性有限 穿过 BBB，表明在大脑中发现的 apo E 是 本地生产。我们的体外试验表明 sAbeta(1-40) 与 apo J 结合的亲和力约为 10 倍 高于 apo E。体内进行的研究表明 sAbeta(1- 40) 与 apo J 和 apo E 的结合导致显着的 114 倍差异 其各自复合物的屏障渗透性。在现在 根据提案，我们计划描述 sAbeta 运输机制 BBB 更详细。需要检验的具体假设是： 1. sAbeta 与 apo J 和 apo E 同工型 2、3 和 4 的结合进行调节 脑血管对 sAbeta 的通透性； 2. sAbeta肽的转运 通过特定的转运蛋白和/或受体穿过血脑屏障； 3. 运输 sAbeta 与 apo J 和 apo E 亚型复合，并纳入高和 极低密度脂蛋白颗粒，即 HDL 和 VLDL，可能是 由 HDL 和/或低密度脂蛋白 (LDL) BBB 受体介导； 4. 衰老改变了 BBB 的转运机制，导致积累 脑微血管中 sAbeta 和淀粉样蛋白相关蛋白的含量 脑实质。与 Abeta 序列相同的豚鼠 人类将会被利用。 sAbeta 与载脂蛋白结合的亲和力 将在体外测定。运输研究将在 原位血管灌注脑，结合毛细血管耗竭 技术，放射性高效液相色谱结束免疫细胞化学分析。假定的 sAbeta 转运蛋白和/或受体将从 BBB 质膜中分离出来。 拟议的研究应有助于我们了解 sAbeta 的转运机制 在 BBB，最终目标是制定可能减少 sAbeta 和淀粉样蛋白相关蛋白在大脑中的积累 脑微血管，从而减缓和/或预防 sAbeta的构象转变及其潜在的细胞毒性 影响。