Role of inflammasomes in Alzheimer's Disease

项目概要/摘要 神经炎症是阿尔茨海默病（AD）的重要组成部分。然而，分子 炎症调节 AD 进展的机制尚未明确。我们发现β-淀粉样蛋白 1-42 激活 NLRP3 炎症小体，AD 患者一致有激活的证据 他们大脑中的炎症小体。为了测试 NLRP3 炎症小体在 AD 中的作用，我们培育了 APP/PS1 小鼠（ 基于淀粉样蛋白的 AD 小鼠模型）转化为 NLRP3、ASC 或 caspase-1 KO（这三种蛋白质组成 NLRP3 炎性体）并观察到这些小鼠完全免受许多 AD 特征的影响 包括学习/记忆缺陷和长时程增强异常。 NLRP3 炎症小体调节两种高度促炎细胞因子 IL-1β 和 IL-18 的表达 小胶质细胞中大量产生；此外，星形胶质细胞强烈表达pro-IL-18。 IL-1β 是 淀粉样斑块周围富含小胶质细胞，可引起发烧，这是一种强烈的急性期 反应、败血症综合征和焦亡细胞死亡。 IL-18 是 IL1 超家族的成员。与IL-不同 1beta、IL-18 不激活 NF-κB 或具有致热活性。尚不清楚 IL-1beta 或 IL-18 减少是造成炎性体缺陷 APP/PS1 小鼠的保护性表型的原因。我们 产生了 IL-18KO/APP/PS1 小鼠，令人惊讶的是，这些小鼠出现了致命的癫痫症， 左乙拉西坦治疗完全逆转。正如流行病学研究表明的那样，这是高度相关的 近三分之二的 AD 患者在病程中的某个时刻会出现癫痫发作。 在目标 1 中，我们将研究炎症小体依赖性细胞焦亡在小胶质细胞中的作用。 使用转基因小鼠研究 AD 的发病机制，其中炎症小体已被专门删除 APP/PS1 背景上的小胶质细胞。我们还将测试缺乏 Gasdermin D（a caspase-1 底物和细胞焦亡的最终效应分子）。在目标 2 中，我们将检查 IL-18 是否 抵消 IL-1β 在 AD 相关癫痫发作中的促癫痫作用。我们将使用遗传方法 （在其他 AD 小鼠模型中删除 IL-18）以及药理学方法（IL-1beta 功能丧失） 方法）在 IL-18KO/APP/PS1 小鼠中进行实验并评估动物的癫痫发作情况。在目标 3 中，我们将确定以下角色： IL-18 减少神经元网络活动并调节突触传递。我们将确定以下类型 IL-18KO/APP/PS1 小鼠中通过形态学和 电生理学研究以及使用免疫组织化学的生化分析。我们将专门 使用 floxed IL-18 转基因小鼠系检查小胶质细胞 IL-18 的作用。顺利完成这些 目标将阐明炎症体产生的细胞因子在 AD 中的作用，并可能产生新的转化 旨在专门阻止导致 AD 的炎症以及机械靶向的方法 癫痫发作会影响 AD 患者的生活质量。

PROJECT SUMMARY/ABSTRACT Neuroinflammation is an important component of Alzheimer's disease (AD). However, the molecular mechanism by which inflammation modulates AD progression is not defined. We discovered that beta-amyloid 1-42 activates NLRP3 inflammasomes and that AD patients uniformly have evidence of activated inflammasomes in their brains. To test the role of NLRP3 inflammasomes in AD, we bred APP/PS1 mice (an amyloid-based murine model of AD) into NLRP3, ASC, or caspase-1 KOs (the three proteins comprising the NLRP3 inflammasome) and observed that these mice were completely protected from numerous AD features including learning/memory deficits and abnormalities in long-term potentiation. NLRP3 inflammasomes regulate the expression of IL-1beta and IL-18, two highly proinflammatory cytokines abundantly produced in microglial cells; in addition, astrocytes are strong expresses of pro-IL-18. IL-1beta is abundant in microglial cells on the periphery of amyloid plaques and can cause fever, a strong acute phase response, sepsis syndrome, and pyroptotic cell death. IL-18 is a member of the IL1 superfamily. Unlike IL- 1beta, IL-18 does not activate NF-kappaB or have pyrogenic activity. It is unknown if IL-1beta or IL-18 reduction was responsible for the protective phenotype in inflammasome-deficient APP/PS1 mice. We generated IL-18KO/APP/PS1 mice and, surprisingly, these mice developed a lethal seizure disorder, which was completely reversed by levetiracetam therapy. This is highly relevant as epidemiologic studies suggest that almost two-thirds of AD patients have seizures at some point during the course of their disease. In Aim 1, we will examine the role of inflammasome-dependent pyroptosis in microglial cells in the pathogenesis of AD using transgenic mice in which the inflammasome has been specifically deleted from microglial cells on an APP/PS1 background. We will also test APP/PS1 mice deficient for Gasdermin D (a caspase-1 substrate and the final effector molecule of pyroptosis). In Aim 2, we will examine if IL-18 counterbalances the proepileptic effects of IL-1beta in AD-related seizures. We will use a genetic approach (deleting IL-18 in other AD mouse models) as well as a pharmacological approach (an IL-1beta loss-of-function approach) in IL-18KO/APP/PS1 mice and assess animals for seizures. In Aim 3, we will determine the role of IL-18 in reducing neuronal network activity and modulating synaptic transmission. We will identify the types of synapses that are dysregulated in IL-18KO/APP/PS1 mice by performing morphological and electrophysiological studies as well as biochemical analysis using immunohistochemistry. We will specifically examine the role of microglial IL-18 using a floxed IL-18 transgenic mouse line. Successful completion of these Aims will elucidate the role of inflammasome-generated cytokines in AD, and could result in novel translational approaches designed to specifically halt the inflammation that drives AD, as well as mechanistically target seizures that affect the quality of life of AD patients.