Concentration-dependent inhibitory effects of apolipoprotein E on Alzheimer's beta-amyloid fibril formation in vitro.
Concentration-dependent inhibitory effects of apolipoprotein E on Alzheimer's beta-amyloid fibril formation in vitro.
复制标题
载脂蛋白 E 对阿尔茨海默病 β-淀粉样原纤维形成的体外浓度依赖性抑制作用。
作者：
H. Naiki;Fumitake Gejyo;Kazuya Nakakuki
最近，许多研究小组在体外研究了载脂蛋白 E (apoE) 对 β-淀粉样原纤维 (betaAf) 形成的影响。然而，他们的数据有些争议，迄今为止还没有对 apoE 作用的精确动力学评估。我们在体外检查了人 apoE 对 betaAf 形成的影响，从不同浓度的新鲜制备的 β-淀粉样蛋白 (1-40) (beta1-40) 开始，并使用硫黄素 T 荧光光谱法。当 50 µM 的 beta1-40 与摩尔比为 1:1000 至 1:100 的 apoE 一起孵育时，观察到 apoE 的剂量依赖性抑制作用。体外 betaAf 形成的成核和延伸阶段均受到 apoE 的抑制。另一方面，当300μM的β1-40与1:100摩尔比的apoE一起孵育时，apoE的抑制作用完全消失。然后，我们利用最近建立的体外 betaAf 延伸一级动力学模型，重点研究了 apoE 对体外 betaAf 形成延伸阶段的抑制作用的动力学 [Naiki, H., & Nakakuki, K. (1996) Lab.投资。 74、374-383]。数据的数学处理表明，apoE 通过与 beta1-40 形成复合物，从而消除反应混合物中游离的 beta1-40，从而在体外抑制 betaAf 的延伸。三种主要重组 apoE 异构体与 beta1-40 的平衡关联常数实际上是相同的。这些结果表明，apoE 对体外 βAf 形成的影响是不同的，并且可以解决有关体外 β-淀粉样蛋白 -apoE 相互作用的一些争议。
Recently, many research groups have examined the effect of apolipoprotein E (apoE) on beta-amyloid fibril (betaAf) formation in vitro. However, their data were somewhat controversial and no exact kinetic assessment of the role of apoE has thus far been available. We examined the effect of human apoE on betaAf formation in vitro, starting with various concentrations of freshly prepared beta-amyloid(1-40) (beta1-40) and using fluorescence spectroscopy with thioflavine T. When 50 microM of beta1-40 was incubated with a 1:1000 to 1:100 molar ratio of apoE, a dose-dependent inhibitory effect of apoE was observed. Both the nucleation and extension phases of betaAf formation in vitro were inhibited by apoE. On the other hand, when 300 microM of beta1-40 was incubated with a 1:100 molar ratio of apoE, the inhibitory effect of apoE was completely abolished. We then focused our study on the kinetics of the inhibitory effect of apoE on the extension phase of betaAf formation in vitro, utilizing the recently established first-order kinetic model of betaAf extension in vitro [Naiki, H., & Nakakuki, K. (1996) Lab. Invest. 74, 374-383]. The mathematical treatment of the data suggests that apoE inhibits the extension of betaAf in vitro, by making a complex with beta1-40, thus eliminating free beta1-40 from the reaction mixture. The equilibrium association constant with beta1-40 was practically the same among the three major recombinant apoE isoforms. These results indicate that the effects of apoE on betaAf formation in vitro is differential and could settle some of the controversy about beta-amyloid-apoE interaction in vitro.