Mouse Model of Myelin Basic Protein-Amyloid Beta Interactions in Brain

淀粉样蛋白(Ass)在大脑中的积聚，无论是作为实质斑块还是 脑血管沉积是阿尔茨海默病(AD)患者的一个重要病理特征 以及几种相关的疾病。Ass多肽来源于淀粉样蛋白ss-蛋白。 前体(Asspp)被ss-分泌酶和？-分泌酶连续的蛋白水解酶切割。这个 促进或阻碍ASS组装成沉积在大脑中的纤维结构的因素 在很大程度上仍然是不确定的。我们实验室最近的体外研究表明，髓鞘碱性 蛋白质(MBP)是大脑中髓鞘的重要组成部分，是ASS纤维的有效抑制物 并可保护培养的原代神经元免受天冬氨酸的毒性作用。尽管 Ass在大脑中的空间沉积与这一发现一致(即大脑白质富含MBP 在很大程度上没有纤维状ASS沉积)，MBP下降与 水平和Ass水平升高目前尚不清楚MBP是否确实影响Ass组装 和体内蓄积。 几个特征良好的人类asspp转基因小鼠模型已经被 产生形成AD样纤维淀粉样沉积的物质。来研究这场战争的后果 在这些已建立的ASPP中，缺乏MBP对纤维淀粉样蛋白的组装和沉积 转基因模型可以将它们培育到MBP基因敲除背景上。这样的一个 被称为颤抖鼠标的模型是存在的，但也有明显的缺点 它们不会形成髓鞘，在出生后几个月内死亡。不幸的是，人类的ASPP 转基因小鼠模型需要远远超过几个月的衰老才能发展出显著的 病理性淀粉样蛋白形成。而不是敲除整个MBP蛋白a的表达 我们计划采用的更复杂的方法将是突变一个高度特定的领域 在MBP蛋白上禁用特定功能。为此，在此的R21阶段 应用程序我们建议生成一种新的“敲入”鼠标模型，我们将在其中介绍 内源性小鼠Golli-MBP基因中特定KRG基序的丙氨酸突变。我们的 最近的研究发现，这个特定的KRG基序是与Ass结合的一个重要元件 多肽和抑制它们的纤维组装。由此产生的新的“敲入”模式将产生 缺乏结合天冬氨酸多肽并抑制其组装的能力的MBP。这些新颖的MBP- 将产生KRG/AAA敲击小鼠，并初步鉴定其存活、生长、 行为和髓鞘形成。 在成功完成此应用程序的R21阶段后，我们计划继续 在R33阶段，我们提出了跨越MBP-KRG/AAA敲击的两种不同的小鼠 出现纤维状淀粉样蛋白沉积的人asspp转基因小鼠模型。十字勋章 将对小鼠品系进行老化，并对其进行定量评估，包括 ASS多肽的沉积及其下游的病理和行为 后果。这些研究的完成将为潜在的生理学提供新的见解 控制致病淀粉样蛋白组装的机制，并可能导致新的途径 对这种病理过程的干预。

Accumulation of the amyloid ss-protein (Ass) in brain, either as parenchymal plaques or cerebrovascular deposits, is a key pathological feature of patients with Alzheimer's disease (AD) and several related disorders. The Ass peptides are derived from the amyloid ss-protein precursor (AssPP) by sequential proteolytic cleavages by ss- and ¿-secretase enzymes. The factors that either promote or impede Ass assembly into fibrillar structures that deposit in brain remain largely undefined. Recent in vitro work from our laboratory has shown that myelin basic protein (MBP), a prominent component of myelin in brain, is a potent inhibitor of Ass fibrillar assembly and can protect cultured primary neurons from the toxic effects of Ass. Although the spatial deposition of Ass in brain is consistent with this finding (i.e. brain white matter rich in MBP is largely devoid of fibrillar Ass deposits) and there is a relationship between decreased MBP levels and increased Ass levels it remains unknown if MBP does indeed influence Ass assembly and accumulation in vivo. Several well-characterized human AssPP transgenic mouse models have been generated that develop AD-like fibrillar amyloid deposits. To study the consequences of the absence of MBP on fibrillar amyloid assembly and deposition in these established AssPP transgenic models one could breed them onto an MBP gene knockout background. Such a model, known as the shiverer mouse, exists but comes with the significant shortcomings in that they do not form myelin and die within several months after birth. Unfortunately, human AssPP transgenic mouse models require aging well beyond several months to develop significant pathologic amyloid formation. Instead of knocking out expression of the entire MBP protein a more sophisticated approach that we plan to employ will be to mutate a highly specific domain on the MBP protein to disable a specific function. To this end, in the R21 Phase of this application we propose to generate a novel "knock in" mouse model where we will introduce alanine mutations into a specific KRG motif in the endogenous mouse Golli-MBP gene. Our recent studies have identified this specific KRG motif as an essential element for binding to Ass peptides and inhibiting their fibrillar assembly. The resulting new "knock in" model will produce MBP that lacks the ability to bind Ass peptides and inhibit their assembly. These novel MBP- KRG/AAA knock in mice will be generated and initially characterized for viability, growth, behavior, and myelination. After successful completion of the R21 Phase of this application we plan to proceed to the R33 Phase where we propose to cross the MBP-KRG/AAA knock in mice with two different human AssPP transgenic mouse models that develop fibrillar amyloid deposition. The crossed mouse lines will be aged and quantitatively evaluated for the acumulation, asembly, and deposition of Ass peptides and the resulting downstream pathological and behavioral consequences. Completion of these studies will provide new insight into potential physiological mechanisms that govern pathogenic amyloid assembly and may lead to new avenues for intervention into this pathologic process.