1 Laboratory of Neuropathology, Institute of Pathology, Centre for Clinical Research at the University of Ulm, Ulm, Germany.
Brain. 2014 Mar;137(Pt 3):887-903. doi: 10.1093/brain/awt362. Epub 2014 Feb 10.
Alzheimer's disease is characterized by the deposition of amyloid-β peptide in the brain. N-terminal truncation resulting in the formation of AβN3pE and phosphorylation at serine 8 have been reported to modify aggregation properties of amyloid-β. Biochemically, soluble, dispersible, membrane-associated, and insoluble, plaque-associated amyloid-β aggregates have been distinguished. Soluble and dispersible amyloid-β aggregates are both in mixture with the extracellular or intracellular fluid but dispersible aggregates can be cleared from proteins in solution by ultracentrifugation. To clarify the role of phosphorylated amyloid-β and AβN3pE in soluble, dispersible, membrane-associated, and plaque-associated amyloid-β aggregates in the pathogenesis of Alzheimer's disease we studied brains from 21 cases with symptomatic Alzheimer's disease, 33 pathologically preclinical Alzheimer's disease cases, and 20 control cases. Western blot analysis showed that soluble, dispersible, membrane-associated and plaque-associated amyloid-β aggregates in the earliest preclinical stage of Alzheimer's disease did not exhibit detectable amounts of AβN3pE and phosphorylated amyloid-β. This stage was referred to as biochemical stage 1 of amyloid-β aggregation and accumulation. In biochemical amyloid-β stage 2, AβN3pE was additionally found whereas phosphorylated amyloid-β was restricted to biochemical amyloid-β stage 3, the last stage of amyloid-β aggregation. Phosphorylated amyloid-β was seen in the dispersible, membrane-associated, and plaque-associated fraction. All cases with symptomatic Alzheimer's disease in our sample fulfilled biochemical amyloid-β stage 3 criteria, i.e. detection of phosphorylated amyloid-β. Most, but not all, cases with pathologically preclinical Alzheimer's disease had biochemical amyloid-β stages 1 or 2. Immunohistochemistry confirmed the hierarchical occurrence of amyloid-β, AβN3pE, and phosphorylated amyloid-β in amyloid plaques. Phosphorylated amyloid-β containing plaques were, thereby, seen in all symptomatic cases with Alzheimer's disease but only in a few non-demented control subjects. The biochemical amyloid-β stages correlated with the expansion of amyloid-β plaque deposition and with that of neurofibrillary tangle pathology. Taken together, we demonstrate that AβN3pE and phosphorylated amyloid-β are not only detectable in plaques, but also in soluble and dispersible amyloid-β aggregates outside of plaques. They occur in a hierarchical sequence that allows the distinction of three stages. In light of our findings, it is tempting to speculate that this hierarchical, biochemical sequence of amyloid-β aggregation and accumulation is related to disease progression and may be relevant for an increasing toxicity of amyloid-β aggregates.
阿尔茨海默病的特征是淀粉样β肽在大脑中的沉积。据报道，N 端截断导致 AβN3pE 的形成和丝氨酸 8 的磷酸化，可改变淀粉样β的聚集特性。从生化角度来看，已经区分了可溶性、分散性、膜相关和不溶性、斑块相关的淀粉样β聚集物。可溶性和分散性的淀粉样β聚集物都与细胞外或细胞内液混合，但分散性的聚集物可以通过超速离心从溶液中的蛋白质中清除。为了阐明磷酸化淀粉样β和 AβN3pE 在阿尔茨海默病发病机制中可溶性、分散性、膜相关和斑块相关淀粉样β聚集物中的作用，我们研究了 21 例有症状的阿尔茨海默病患者、33 例病理性临床前阿尔茨海默病患者和 20 例对照患者的大脑。Western blot 分析显示，在阿尔茨海默病最早的临床前阶段，可溶性、分散性、膜相关和斑块相关的淀粉样β聚集物未检测到 AβN3pE 和磷酸化淀粉样β。这一阶段被称为淀粉样β聚集和积累的生化阶段 1。在生化淀粉样β阶段 2 中，另外发现了 AβN3pE，而磷酸化淀粉样β仅限于生化淀粉样β阶段 3，即淀粉样β聚集的最后阶段。磷酸化的淀粉样β出现在分散性、膜相关和斑块相关的部分。我们样本中所有有症状的阿尔茨海默病病例均符合生化淀粉样β阶段 3 的标准，即检测到磷酸化淀粉样β。大多数（但不是全部）有病理学临床前阿尔茨海默病的病例具有生化淀粉样β 1 或 2 期。免疫组织化学证实了淀粉样β、AβN3pE 和磷酸化淀粉样β在淀粉样斑块中的分层发生。因此，所有有症状的阿尔茨海默病病例都可见到含有磷酸化淀粉样β的斑块，但只有少数非痴呆对照受试者可见。生化淀粉样β阶段与淀粉样β斑块沉积的扩张以及神经原纤维缠结病理的扩张相关。综上所述，我们证明 AβN3pE 和磷酸化淀粉样β不仅可在斑块中检测到，也可在斑块外的可溶性和分散性淀粉样β聚集物中检测到。它们按照一个层次顺序出现，允许区分三个阶段。根据我们的发现，人们不禁推测，这种淀粉样β聚集和积累的分层生化序列与疾病进展有关，可能与淀粉样β聚集物的毒性增加有关。
