Department of Biochemistry and Molecular Biology and Molecular, Cellular and Integrative Neuroscience Program, Colorado State University, Fort Collins, CO, USA.
J Alzheimers Dis. 2011;24(4):681-91. doi: 10.3233/JAD-2011-101899.
Amyloid-β (Aβ) peptides, 36-43 amino acids in length, are produced from β- and γ-secretase cleavage of the amyloid-β protein precursor (AβPP), and are one of the causative agents of Alzheimer's disease (AD). Here we show that an ELISA can detect total rodent Aβ without interference from physiological concentrations of human Aβ. In cultured dissociated rat cortical neurons and rat and mouse hippocampal organotypic slices, we apply the assay to measure the production of Aβ in response to treatment with hydrogen peroxide, a known stimulator of Aβ secretion, or human Aβ dimer/trimer (Aβd/t), fractionated from the culture medium of 7PA2 cells. Peroxide increases Aβ secretion by about 2 fold, similar to results from previous reports that used a different assay. Of greater significance is that physiologically relevant concentrations (~250 pM) of human Aβd/t increase rodent Aβ secretion from cultured rat cortical neurons by >3 fold over 4 days. Surprisingly, neither treatment with peroxide nor human Aβd/t leads to accumulation of intracellular Aβ. Human Aβd/t increased >2 fold the Aβ secreted by organotypic hippocampal slices from tau knock-out mice whether or not they expressed a human tau transgene, suggesting tau plays no role in enhanced Aβ secretion. Together, these results support an Aβ-mediated feed-forward mechanism in AD progression.
淀粉样β（Aβ）肽长 36-43 个氨基酸，由β-和γ-分泌酶切割淀粉样β蛋白前体（AβPP）产生，是阿尔茨海默病（AD）的致病因素之一。在这里，我们展示了一种 ELISA 可以检测总啮齿动物 Aβ，而不会受到生理浓度人 Aβ的干扰。在培养的分离大鼠皮质神经元和大鼠及小鼠海马器官切片中，我们应用该测定法来测量 Aβ 的产生，以响应过氧化氢（一种已知的 Aβ 分泌刺激剂）或人 Aβ 二聚体/三聚体（Aβd/t）的处理，Aβd/t 从 7PA2 细胞的培养基中分离。过氧化氢使 Aβ 分泌增加约 2 倍，与先前使用不同测定法的结果相似。更重要的是，生理相关浓度（约 250 pM）的人 Aβd/t 在 4 天内使培养的大鼠皮质神经元中的啮齿动物 Aβ 分泌增加了 >3 倍。令人惊讶的是，无论是用过氧化氢处理还是用人 Aβd/t 处理，都不会导致细胞内 Aβ 的积累。人 Aβd/t 增加了来自 tau 敲除小鼠的器官型海马切片中 Aβ 的分泌，无论它们是否表达人 tau 转基因，这表明 tau 在增强 Aβ 分泌中不起作用。总之，这些结果支持 AD 进展中的 Aβ 介导的正反馈机制。
