Department of Laboratory Medicine, Shimane University School of Medicine, Izumo, Japan.
FEBS J. 2015 Feb;282(3):463-76. doi: 10.1111/febs.13151. Epub 2014 Dec 15.
Amyloid β (Aβ), a 39-42 amino acid peptide derived from amyloid precursor protein, is deposited as fibrils in Alzheimer's disease brains, and is considered to play a major role in the pathogenesis of the disease. We have investigated the effects of a water-soluble Zn-phthalocyanine, ZnPc(COONa)₈, a macrocyclic compound with near-infrared optical properties, on Aβ fibril formation in vitro. A thioflavin T fluorescence assay showed that ZnPc(COONa)₈ significantly inhibited Aβ fibril formation, increasing the lag time and dose-dependently decreasing the plateau level of fibril formation. Moreover, it destabilized pre-formed Aβ fibrils, resulting in an increase in low-molecular-weight species. After fibril formation in the presence of ZnPc(COONa)₈, immunoprecipitation of Aβ₁₋₄₂ using Aβ-specific antibody followed by near-infrared scanning demonstrated binding of ZnPc(COONa)₈ to Aβ₁₋₄₂. A study using the hydrophobic fluorescent probe 8-anilino-1-naphthalenesulfonic acid showed that ZnPc(COONa)8 decreased the hydrophobicity during Aβ₁₋₄₂ fibril formation. CD spectroscopy showed an increase in the α helix structure and a decrease in the β sheet structure of Aβ₁₋₄₀ in fibril-forming buffer containing ZnPc(COONa)₈. SDS/PAGE and a dot-blot immunoassay showed that ZnPc(COONa)₈ delayed the disappearance of low-molecular-weight species and the appearance of higher-molecular-weight oligomeric species of Aβ₁₋₄₂. A cell viability assay showed that ZnPc(COONa)₈ was not toxic to a neuronal cell line (A1), but instead protected A1 cells against Aβ₁₋₄₂-induced toxicity. Overall, our results indicate that ZnPc(COONa)₈ binds to Aβ and decreases the hydrophobicity, and this change is unfavorable for Aβ oligomerization and fibril formation.
淀粉样蛋白β（Aβ）是一种 39-42 个氨基酸的肽，源自淀粉样前体蛋白，在阿尔茨海默病大脑中沉积为纤维，被认为在疾病的发病机制中起主要作用。我们研究了一种水溶性锌酞菁，ZnPc（COONa）₈，一种具有近红外光学性质的大环化合物，对体外 Aβ 纤维形成的影响。噻唑蓝 T 荧光测定法显示，ZnPc（COONa）₈ 显著抑制 Aβ 纤维形成，增加了潜伏期并剂量依赖性地降低了纤维形成的平台水平。此外，它使预先形成的 Aβ 纤维不稳定，导致低分子量物质增加。在 ZnPc（COONa）₈ 存在下形成纤维后，用 Aβ 特异性抗体进行 Aβ₁₋₄₂ 免疫沉淀，然后进行近红外扫描，证明 ZnPc（COONa）₈ 与 Aβ₁₋₄₂ 结合。使用疏水性荧光探针 8-苯胺-1-萘磺酸进行的研究表明，ZnPc（COONa）₈ 降低了 Aβ₁₋₄₂ 纤维形成过程中的疏水性。CD 光谱显示，在含有 ZnPc（COONa）₈ 的纤维形成缓冲液中，Aβ₁₋₄₀ 的α 螺旋结构增加，β 片层结构减少。SDS/PAGE 和斑点印迹免疫测定显示，ZnPc（COONa）₈ 延迟了低分子量物质的消失和 Aβ₁₋₄₂ 更高分子量寡聚物的出现。细胞活力测定显示，ZnPc（COONa）₈ 对神经元细胞系（A1）没有毒性，反而可以保护 A1 细胞免受 Aβ₁₋₄₂ 诱导的毒性。总的来说，我们的结果表明，ZnPc（COONa）₈ 与 Aβ 结合并降低疏水性，这种变化不利于 Aβ 寡聚化和纤维形成。
