BIOCHEMISTRY OF LEWY BODIES AND GELSOLIN AMYLOID

路易体和凝溶胶蛋白淀粉样蛋白的生物化学

• 批准号: 3417343
• 负责人: BLAS FRANGIONE
• 金额: $ 23.81万
• 依托单位: NEW YORK UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-09-30 至 1995-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3417343
• 关键词: Alzheimer's disease; Parkinson's disease; amyloid proteins; antibody formation; brain disorder diagnosis; cerebrospinal fluid; cytoplasm; diagnosis design /evaluation; epitope mapping; gelsolin; histopathology; human genetic material tag; human tissue; immunocytochemistry; laboratory rabbit; molecular cloning; neural degeneration; neurochemistry; postmortem; protein sequence; western blottings

Lewy Bodies (LBs) are distinctive intraneural cytoplasmic inclusions of unknown pathogenesis. They are found in all cases of Parkinson's Disease (PD), and diffuse Lewy body disease (DLBD) (also known as the Lewy body variant of Alzheimer's disease). In addition, LBs are occasionally found in a few other pathological processes. The composition of LBs by direct biochemical methods and their role in these diseases processes remains unknown. Recently we have shown that antibodies raised to purified amyloid extracted from familial amyloidosis, Finnish type (FAF) immunoreact with LBs. This immunoreactivity is specific, as it was absorbed by the purified amyloid proteins and is unaffected by other antigens such as ubiquitin, neurofilament proteins and tubulin which have been shown by immunohistochemical methods to be present in LBs. FAF amyloid has been found to be an aberrant degradation product, starting at position 173, of gelsolin, an actin modulating protein. The amino acid sequence shows heterogeneity at the N-terminus and at position 15, where asparagine substitutes for aspartic acid. At the DNA level, a guanine to adenine transition corresponding to nucleotide 654 of the human gelsolin has been found in all FAF patients tested so far. Commercially available anti- gelsolin antibodies raised to the carboxy-terminus of the molecule (away from the amyloidogenic region) do not immunostain LBs. This raises the possibility that LBs also contain an abnormal fragment of gelsolin that is closer to the amino terminal of the molecule. We plan: (1) To perform an immunohistochemical survey of DLBD, Alzheimer's disease, and Parkinson's disease cases with the anti-FAF antibody, as well as with antibodies to intact gelsolin, fragments and synthetic peptides of different regions of the gelsolin molecule, to evaluate patterns of immunoreactivity and epitope mapping; (2) To characterize the composition of the purified LBs preparation by using Western blotting and direct amino acid sequencing to sequence gelsolin and/or its fragments; (3) To investigate the biological activity of gelsolin isolated from DLBD, PD, AD, and control CSF to see if there are any abnormalities; (4) To clone the gelsolin gene from familial PD patients and screen for any variations; (5) To investigate cerebral spinal fluid from patients with PD, DLBD, other neurodegenerative disorder, controls and AD for the presence of any abnormal gelsolin fragments that could be used as a diagnostic test. Together these studies will expand knowledge about LBs and the role gelsolin plays in their formation.

路易体（LB）是一种独特的神经内细胞质包涵体， 未知的发病机制。 它们存在于所有帕金森病病例中 (PD)和弥漫性路易体病（DLBD）（也称为路易体 阿尔茨海默病的变种）。 此外，偶尔会发现LB 在其他一些病理过程中。 LB的直接组成 生物化学方法及其在这些疾病过程中的作用仍然存在 未知 最近，我们已经证明，从纯化的淀粉样蛋白中提取的抗体 来自家族性淀粉样变性，芬兰型（FAF）与LB免疫反应。 这 免疫反应性是特异性的，因为它被纯化的淀粉样蛋白吸收， 蛋白质并且不受其它抗原如泛素的影响， 神经丝蛋白和微管蛋白， 免疫组化方法检测LB。 FAF淀粉样蛋白已经被 发现是从位置173开始的异常降解产物， 凝溶胶蛋白，一种肌动蛋白调节蛋白。 氨基酸序列显示 在N-末端和位置15处的异质性，其中天冬酰胺 天冬氨酸的替代品。 在DNA水平上，鸟嘌呤到腺嘌呤 对应于人凝溶胶蛋白的核苷酸654的转换已经被 在所有的FAF患者中发现的。 市售抗- 凝溶胶蛋白抗体上升到分子的羧基末端（远离 来自淀粉样蛋白生成区域）不免疫染色LB。 这就提出了一个 LB也可能含有凝溶胶蛋白的异常片段， 更靠近分子的氨基末端。 我们计划：（1）执行一项 DLBD、阿尔茨海默病和帕金森病免疫组化研究 具有抗FAF抗体的疾病病例，以及具有 完整的凝溶胶蛋白，不同区域的片段和合成肽 凝溶胶蛋白分子，以评估免疫反应性和表位的模式 （2）鉴定纯化的LB的组成 通过使用蛋白质印迹和直接氨基酸测序来制备， （3）研究凝溶胶蛋白及其片段的生物学特性， 从DLBD、PD、AD和对照CSF中分离的凝溶胶蛋白的活性，以观察 （4）克隆了日本血吸虫凝溶胶蛋白基因， （5）研究帕金森病患者的脑功能变化， 来自PD、DLBD、其他神经退行性疾病患者的脊髓液， 对照和AD中是否存在任何异常凝溶胶蛋白片段， 可以用作诊断测试。 这些研究将扩大 关于LB的知识和凝溶胶蛋白在其形成中的作用。