The role of the blood-brain barrier in Alzheimer s disease: Interaction of LRP1 and ABC transporter function

长期以来，人们已经知道神经毒性β-淀粉样肽（Aβ）的积累是阿尔茨海默病（AD）的特征性标志。Randy Bateman博士和其他人证明，Aβ肽的产生率在健康个体和AD患者之间没有差异。我们和其他人已经表明，健康和痴呆个体之间Aβ肽的清除功能发生了显著变化。因此，我们在本申请中提出进一步阐明脑中最大的Aβ处理结构-血脑屏障（BBB）的这种清除机制的细节。我们已经证明LRP 1、ABC转运蛋白B1和C1是血脑屏障的主要清除蛋白，并且这些清除机制很可能不是由这些转运蛋白中的一种介导的单一事件。我们的项目旨在破译Aβ清除的协同作用。我们将能够确定LRP 1单独、与特异性ABC转运蛋白组合或特异性ABC转运蛋白单独是否能够将大多数Aβ肽转运出大脑。因此，我们希望澄清这一过程背后的机制，并使用不同的独特的小鼠模型，我们将产生原代小鼠脑内皮细胞进行体外BBB转胞吞研究。为了验证我们的体外观察结果，我们将通过交叉相应的小鼠品系将这些实验设置转移到体内情况中，并分析BBB清除机制。此外，我们将通过使用仅感染BBB内皮的高度特异性AAV特异性地操纵活小鼠中的脑内皮细胞。使用这种方法，我们将尝试在大脑外执行增强清除方法。有趣的是，不仅有废物的清除机制描述了大脑。也有人提出，聚集形式的Aβ从外周组织进入大脑，引发淀粉样病变。在最近的一项手术连接AD模型小鼠与野生型小鼠的联体共生实验中，已显示人Aβ42在野生型动物的脑中蓄积。这可能意味着外周Aβ-肽有一条直接进入大脑的途径。为了分析这一点，我们将利用我们的体外和体内设置，并确定是否存在以及在何种程度上存在这样的转运途径。综上所述，我们将能够使用体外和体内应用的最新技术来破译BBB Aβ转运的分子机制。Pahnke和Pietrzik实验室的共同努力和专业知识提供了独特的模型系统，这些系统将被结合起来回答BBB如何以及在多大程度上影响大脑的清除以及Aβ肽可能流入大脑的重要问题。

It has been known for a long time that the accumulation of the neurotoxic beta-amyloid peptides (Aβ) is a characteristic hallmark of Alzheimer’s disease (AD). Dr. Randy Bateman and others demonstrated that the production rate of Aβ peptides does not differ between healthy individuals and patients with AD. We and others have shown the clearance function of Aβ peptides between healthy and demented individuals is dramatically altered. Therefore, we propose in this application to further clarify the details of this clearance mechanisms of the largest Aβ disposal structure of the brain, the blood brain barrier (BBB). We have already shown that LRP1, ABC transporters B1 and C1 are major clearance proteins at the BBB, and that these clearance mechanisms most likely are not singular event mediated by just one of these transporters. Our project aims to decipher the concerted action of the Aβ clearance. We will be able to determine whether LRP1 alone, in combination with specific ABC transporters or specific ABC transporters alone are capable to transport the majority of Aβ peptides out of the brain. Hence, we want to clarify the mechanisms behind this process and use different unique mouse models from which we will generate primary mouse brain endothelial cells to perform in vitro BBB transcytosis studies. To validate our in vitro observations, we will transfer these experimental setups into the in vivo situation by crossing the respective mouse strains and analyse BBB clearance mechanisms. Additionally, we will specifically manipulate the endothelial cells of the brain in a living mouse by using highly specific AAVs which only infect the BBB endothelium. Using this approach we will try to perform an enhanced clearance approach out of the brain. Interestingly, there are not only clearance mechanisms of waste products described out of the brain. It has also been proposed that aggregated forms of Aβ travel from peripheral tissues into the brain to seed amyloid pathology. In a recent parabiosis experiment of surgically joined AD model mice to wild-type mice it has been shown that human Aβ42 accumulates in the brains of wild-type animals. This may imply that there is a direct route of peripheral Aβ-peptides into the brain. To analyse this we will make use of our in vitro and in vivo settings and determine if and to which extend such transport routes exist.Taken together we will be able to decipher the molecular mechanism of BBB Aβ-transport using state of the art in vitro and in vivo applications. The combined efforts and expertise of the Pahnke and Pietrzik laboratories offer unique model systems which will be combined to answer the important question how and to which extent the BBB influences the clearance out of the brain and potentially the influx of Aβ-peptides into the brain.