阿尔茨海默病患者和非痴呆老人α2巨球蛋白抑制β淀粉样肽纤维形成能力的比较

Comparison of alpha2-macroglobulin to inhibite fibril formation of beta-amyloid peptide between patients with Alzheimer's disease and non-dementia control

目的 分析阿尔茨海默病患者和非痴呆老人及青年人的α2 巨球蛋白体外抑制β淀粉样肽纤维形成能力的差异,阐明α2巨球蛋白对β淀粉样肽降解代谢变化的影响。方法 使用浊度试验、硫磺素 T试验和电子显微镜负染方法,观察分析了β淀粉样肽的凝聚和纤维形成及26例阿尔茨海默病患者和 27 例非痴呆老人及 24 例青年人的α2 巨球蛋白对β淀粉样肽的凝聚和纤维形成的抑制作用。结果 12.5μmol/L浓度 Aβ1-42 肽在 pH7.4 的 PBS缓冲液中37℃恒温摇床温育72 h后电子显微镜负染可见大量的β淀粉样肽纤维形成。正常青年人的 50μmol/Lα2 巨球蛋白电镜下观察证实完全地抑制了β淀粉样肽纤维形成,浊度试验表明抑制率达(81.2±7.2)%,硫磺素 T试验抑制率达(78.7±7.7)%。非痴呆老年组等量α2巨球蛋白电镜下 2/27 例显示不全抑制,25/27 完全抑制,浊度和硫磺素 T试验抑制率分别为(73.0±8.7)%和(68.3±9.6)%,同青年组比较无显著性差异(P>0.05)。阿尔茨海默病患者等量α2巨球蛋白电镜下 15/26 例显示不全抑制,9/26 完全抑制,浊度和硫磺素 T试验抑制率分别为(50.0±6.87)%和(54.9±7.1)%,同青年组比较明显减低(P<0.01)。结论 阿尔茨海默病患者α2巨球蛋白体外抑制β淀粉样肽纤维形成能力明显减低。

Objective To explore the comparison of alpha2-macroglobulin to inhibit fibril formation of beta-amyloid peptide between patients with Alzheimer's disease and non-dementia control and to elucidate the effect of Alpha2-macroglobulin on the degradation metabolism of beta-amyloid peptide. Methods Both fibril formation of beta-amyloid peptide and fibril formation inhibition of alpha2-macroglobulin from 26 patients with Alzheimer's disease, 27 non-dementia control and 24 normal young people were determined by tests of turbidity and Thioflavine T fluorometric measurements and electronic microscopy negatively staining. Results The fibril formation of beta-amyloid peptide was observed by electron microscope when a freshly solubilized beta-amyloid (12.5μmol/L) was incubated in PBS, pH 7.4 with shaking at 37℃ for 3 days. Fibril formation of beta-amyloid peptide was completely prevented by using combined incubation of beta-amyloid peptide with alpha2-macroglobulin from 24/24 normal young group, 25/27 from non-dementia older people and 11/26 patients with Alzheimer's disease using electronic microsopic observation. Inhibation rate of beta-amyloid peptide aggregation for alpha2-macroglobulin was significantly lower in Alzheimer's disease group than in young people group \[(50.0±6.87)%vs. (81.2±7.2)% P<0.01 for turbidity test and (54.9±7.1)%vs. (78.7±7.7)% P>0.05 for thioflavine T assay\], but it was not significantly different in young peoples group and non dementia control \[(81.2±7.2)%vs. (73.0±8.7)% P> 0.05 for turbidity test and (78.7±7.7)%vs. (68.3±9.6)% P>0.05 for thioflavine T assay\]. Conclusion The ability of alpha2-macroglobulin from patients with Alzheimer's disease to prevent beta-amyloid peptide aggregation is reduced and it will help form the accelerate formation neurotoxic protofibrillar and fibril and impede clearance of beta-amyolid peptide from tissues such as brain.