REGULATION AND PROCESSING OF AMYLOID PRECURSOR PROTEIN GENES AND GENE PRODUCTS

工作概述：本项目的一个主要重点是发现淀粉样蛋白前体蛋白（APP）在阿尔茨海默病（AD）的病因和病理中的作用。这种蛋白质的正常生理作用也在研究中。APP的加工和早老素（PS）突变对APP加工的影响直接关系到AD老年斑中a - β肽的细胞内生成和细胞外沉积的增加，因此APP的研究很重要。APP的加工也产生分泌形式的蛋白质，可能具有神经营养特性。阿尔茨海默病患者的大脑表现出选择性和大量的神经元丢失。鉴于我们实验室最近的证据表明，在细胞培养中，家族性AD突变形式的APP和PS的过度表达导致凋亡细胞死亡，我们对研究这种细胞死亡的机制感兴趣。我们实验室的目标之一是发现APP或PS突变如何导致AD中特定的神经元细胞损失。先前我们发现，在稳定转染的PC12细胞中，突变形式的APP过表达导致细胞内淀粉样蛋白c端APP片段的产生增加。这伴随着数天内凋亡细胞死亡的增加。我们最近发现，通过腺病毒介导的基因转移过度表达FAD突变体APP可导致初代皮质神经元细胞死亡。细胞内羧基端APP片段和a -肽增多。通过检测与寡核小体结合的DNA片段的ELISA方法测量，过表达导致72小时内凋亡细胞死亡增加约4倍。此外，野生型APP过表达导致细胞死亡增加，表明正常APP水平升高可能具有神经毒性特征。原代皮质神经元的细胞死亡，乃至阿尔茨海默病的细胞死亡，可能是由于APP的神经毒性和神经营养加工产物的生成不平衡。这些产物可能在细胞内起作用，也可能具有旁分泌作用，被释放到介质中，影响邻近细胞。这些结果为针对凋亡通路和APP加工的特定元件进行AD治疗干预提供了理论依据。-淀粉样蛋白，a -肽，衰老，神经变性，细胞凋亡

Summary of Work:A major focus of this project is to discover the role of the Amyloid Precursor Protein (APP) in the etiology and pathology of Alzheimers Disease (AD). The normal physiological role of this protein is also under investigation. APP is important to study since the processing of APP and the effect of Presenilin (PS) mutations on APP processing bear directly on the increased intracellular production and extracellular deposition of A-beta peptides in senile plaques of AD. The processing of APP also generates secreted forms of the protein which may have neurotrophic properties. Brains of AD patients exhibit selective and massive neuronal loss. In light of recent evidence from our laboratory showing that, in cell culture, over-expression of familial AD mutated forms of APP and PS causes apoptotic cell death, we are interested in examining the mechanisms involved in this cell death. One of the aims of our laboratory is to discover how APP or PS mutations lead to specific neuronal cell loss in AD.Previously we showed that over-expression of mutated forms of APP in stably transfected PC12 cells led to an increased production of intracellular, amyloidogenic C-terminal fragments of APP. This was accompanied by increased apoptotic cell death over several days. We recently showed that over-expression of FAD mutant APP by adenovirus-mediated gene transfer leads to cell death of primary cortical neurons. Increased amounts of carboxyl terminal APP fragments and A-beta peptide were detected intracellularly. The overexpression led to about a four-fold increase in apoptotic cell death over 72 hours as measured by an ELISA method detecting fragmented DNA bound to oligonucleosomes. In addition wild-type APP over-expression caused an increase in cell death suggesting that elevated levels of normal APP may have neurotoxic characteristics. This cell death in primary cortical neurons and by extension, in AD, may result from an imbalance in the generation of neurotoxic and neurotrophic processing products of APP. These products may act within the cell or may have a paracrine effect, being released into the media to affect neighboring cells. These results provide a rationale for targeting particular elements of apoptotic pathways and APP processing for therapeutic intervention in AD. - amyloid, A-beta peptide, aging, neurodegeneration, apoptosis