Antifungal activity of amyloid beta as a driver of dementia and AD pathogenesis.

β 淀粉样蛋白的抗真菌活性是痴呆和 AD 发病机制的驱动因素。

• 批准号: 10711875
• 负责人: ANGIE GELLI
• 金额: $ 37.42万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-15 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10711875
• 关键词: Abeta clearance; Adult; Aged, 80 and over; Alzheimer' s Disease; Alzheimer' s disease patient; Amyloid beta-Protein; Amyloid beta-Protein Precursor; Animal Model; Animals; Antifungal Agents; Area; Autopsy; Binding Proteins; Blood - brain barrier anatomy; Blood brain barrier dysfunction; Brain; Brain region; Candida; Cell Wall; Cells; Central Nervous System; Cerebrospinal Fluid; Chitin; Cryptococcus; Cryptococcus neoformans; Cryptococcus neoformans infection; Data; Decision Making; Dementia; Diagnosis; Endothelium; Environment; EphA2 Receptor; Exhibits; Freezing; Functional disorder; Fungal Components; Fungal DNA; Fungal Spores; Gene Expression Profile; Genes; Health; Human; Immunity; Impaired cognition; Impairment; Infection; Inflammation; Inflammatory; Inhalation; Innate Immune Response; Invaded; Mediating; Mediator; Memory Loss; Meningoencephalitis; Mission; Moods; Mus; Mycoses; Nature; Neurodegenerative Disorders; Neurologic Symptoms; Neuronal Injury; Neurons; Oxidative Stress; Pathogenesis; Patients; Polysaccharides; Process; Proliferating; Property; Proteins; Regulation; Reporting; Reproduction spores; Research; Research Project Grants; Role; Sampling; Senile Plaques; Signal Transduction; Structure; Structure of parenchyma of lung; Temperament; Testing; Tissues; United States National Institutes of Health; abeta deposition; aerosolized; amyloid precursor protein processing; antimicrobial; blood damage; brain endothelial cell; brain tissue; experience; extracellular; fungus; improved; in vitro Model; in vivo; long term memory; mouse model; neuroinflammation; neurotropic; parent grant; parent project; pathogenic fungus; therapeutic development

PROJECT ABSTRACT The blood-brain barrier (BBB) is a highly-restrictive structure that protects the central nervous system (CNS). Few fungal pathogens, such as Cryptococcus neoformans (Cn), can breach the BBB and invade the CNS. Fungal spores are prevalent in our environment and we become infected following inhalation of aerosolized spores. Once inhaled, spores of Cn proliferate in lung tissue and disseminate to the CNS due to their neurotropic nature. Cn is the leading cause of fungal meningoencephalitis in adults and it is often misdiagnosed as Alzheimer’s Disease (AD) due to similar neurological symptoms. Indeed, cognitive impairment was reported in 61% of subjects at least one year following Cn infection diagnosis. In AD, inflammatory and infectious processes promote BBB dysfunction underscoring the potentially critical role of the amyloid precursor protein (APP) in endothelial inflammation of the BBB and its subsequent dysfunction. We resolved the transcriptional signature of human brain microvascular endothelial cells (BMECs) infected with Cn. BMECs challenged with Cn showed significant dysregulation of several genes essential to the proper function of the brain endothelium (i.e. BBB), including EphA2-receptor tyrosine kinase, a key mediator of BBB dysfunction during Cn challenge (based on studies from our parent grant). We also identified APBB3 (amyloid beta precursor binding protein 3) as a potential mediator of BBB dysfunction. APBB3 is an uncharacterized protein whose role in APP processing/regulation has never been examined. Our results are consistent with similar changes reported in the abnormal BBB in AD pathogenesis. Reduced BBB integrity is associated with neuroinflammation, neuronal injury, oxidative stress and faulty clearance of amyloid beta (A). Based on our data we propose that fungal brain infection causes BBB dysfunction in part by dysregulating APP and reducing A clearance which would further damage the BBB. Our data are consistent with recent reports that found various brain regions from AD patients at autopsy infected with different species of fungi. Fungal DNA, proteins and structures were identified in frozen brain tissue from AD patients, but not from control patient tissue. Moreover fungal material was detected intra- and extracellularly in neurons from AD patients. Chitin polysaccharides, a key component of fungal cell walls, were also identified and human AD cerebral spinal fluid samples further confirmed the presence of fungi, including Cryptococcus and Candida. Indeed, following these recent discoveries, and the proposed antimicrobial properties of A a new AD hypothesis referred to as “antimicrobial protection hypothesis” has emerged. This hypothesis suggests that deposition of A plaque in brain can initiate early innate immune responses, where A entraps and neutralizes the invading fungal pathogen under normal conditions. The proposed supplement research project will address 2 specific aims that are an extension of the parent project and are fundamentally relevant to the pathogenesis of AD. The following specific aims will test the hypothesis that brain fungal infection dysregulates APP via APBB3 and the resulting antifungal immunity activity of amyloid beta (A) is mediated by EphA2- signaling. Specific aim 1 will examine the role of APP in BBB dysfunction in a mouse model of fungal brain infection. APP and APBB3 activity and their effects on BBB integrity will be examined in wild type and ephA2-/- mice infected with Cn. The physical interaction of A peptides with fungal cells will be examined in vivo and direct effects of A peptides on BBB integrity will be assessed in BBB spheroids, a highly-relevant in vitro model of the BBB. Specific aim 2 will determine whether A exhibits antifungal activity or induces antifungal immunity activity via EphA2-mediated signaling. We have performed several animal studies to resolve mechanisms of fungal brain pathogenesis by examining EphA2 activity in BBB dysfunction, however we have not examined neurological markers of AD in the context of cryptococcal brain infection. Given our expertise in animal models of fungal infection and experience working with in vitro models of the BBB, the studies proposed below are a natural segue to exploring an exciting and potentially ground breaking area of AD pathogenesis and therapeutic development.

项目摘要 血脑屏障（BBB）是保护中枢神经系统的高度限制性结构 (CNS) 很少有真菌病原体，例如新型隐球菌 (Cn)，可以突破 BBB 并侵入 中枢神经系统 (CNS) 真菌孢子在我们的环境中普遍存在，我们在吸入真菌孢子后就会受到感染。 一旦吸入，Cn 孢子就会在肺组织中增殖并传播到中枢神经系统。 它们的神经营养性质是成人真菌性脑膜脑炎的主要原因。 由于类似的神经系统症状，被误诊为阿尔茨海默病（AD）。 在 AD 中，61% 的受试者在 Cn 感染诊断后至少一年内出现功能障碍。 炎症和感染过程促进 BBB 功能障碍，强调了潜在的关键作用 淀粉样前体蛋白 (APP) 在 BBB 内皮炎症及其随后的功能障碍中的作用。 我们解析了感染的人脑微血管内皮细胞（BMEC）的转录特征 受到 Cn 攻击的 BMEC 显示出对正常功能至关重要的几个基因的显着失调。 脑内皮（即 BBB）的功能，包括 EphA2 受体酪氨酸激酶（BBB 的关键介质） Cn 挑战期间的功能障碍（基于我们家长资助的研究）我们还确定了 APBB3（淀粉样蛋白）。 β 前体结合蛋白 3) 作为 BBB 功能障碍的潜在介质，是一种尚未表征的蛋白。 其在 APP 加工/调节中的作用从未被检查过，我们的结果与此一致。 AD 发病机制中血脑屏障异常的类似变化与血脑屏障完整性降低有关。 神经炎症、神经元损伤、氧化应激和β淀粉样蛋白 (A) 的错误清除。 根据数据，我们认为真菌性脑部感染导致 BBB 功能障碍的部分原因是 APP 失调和减少 A 清除会进一步损害 BBB，我们的数据与最近发现的报告一致。 尸检时 AD 患者的各个大脑区域感染了不同种类的真菌 DNA， 在 AD 患者的冷冻脑组织中鉴定出蛋白质和结构，但在对照患者中未鉴定出蛋白质和结构 此外，在 AD 患者的神经元细胞内和细胞外均检测到了真菌物质。 多糖是真菌细胞壁的关键成分，也被鉴定出，并且人类 AD 脑脊髓 事实上，液体样本进一步证实了真菌的存在，包括隐球菌和念珠菌。 这些最近的发现以及 A 提出的抗菌特性被称为新的 AD 假说 随着“抗菌保护假说”的出现，该假说表明 A 斑块的沉积。 大脑中的 A 可以启动早期先天免疫反应，其中 A 捕获并中和入侵的真菌 正常条件下的病原体。拟议的补充研究项目将解决两个具体目标。 它们是父项目的延伸，与 AD 的发病机制根本相关。 以下具体目标将检验脑真菌感染通过 APBB3 调节 APP 的假设，以及 β 淀粉样蛋白 (A) 的抗真菌免疫活性由 EphA2- 信号传导介导。 图 1 将检查 APP 在真菌脑感染小鼠模型中 BBB 功能障碍中的作用。 将在感染 Cn 的野生型和 ephA2-/- 小鼠中检查其活性及其对 BBB 完整性的影响。 将在体内检查 A 肽与真菌细胞的物理相互作用以及 A 的直接影响 肽对 BBB 完整性的影响将在 BBB 球体（一种高度相关的 BBB 体外模型）中进行评估。 具体目标2将确定A是否表现出抗真菌活性或诱导抗真菌免疫活性 通过 EphA2 介导的信号传导，我们进行了多项动物研究来解决真菌的机制。 通过检查 BBB 功能障碍中的 EphA2 活性来研究脑发病机制，但我们尚未检查 鉴于我们在动物模型方面的专业知识，我们在隐球菌脑部感染的情况下发现了 AD 的神经学标志物。 根据真菌感染的经验和 BBB 体外模型的工作经验，下面提出的研究是 自然地继续探索 AD 发病机制中令人兴奋且具有潜在突破性的领域， 治疗的发展。