Role of apoE-mediated meningeal lymphatic remodeling in the pathophysiology of Alzheimer’s disease

摘要 年龄和载脂蛋白Eε4(ApoE 4)基因等位基因的表达是两个主要的危险因素 阿尔茨海默氏症。最近的研究表明，衰老会导致骨质疏松症患者脑液排出障碍。 发现于哺乳动物脑膜硬脑膜的真实淋巴管网络。有趣的是，早期的损失 脑膜淋巴引流与更严重的淀粉样蛋白β(Aβ)病理、血管和小胶质细胞有关 阿尔茨海默病小鼠脑淀粉样变性模型的激活和空间记忆缺陷。然而， APOE4的表达对脑膜淋巴功能的影响知之甚少。我们的初步数据 显示APOE4在人源化敲入小鼠中的表达导致有害的脑膜重塑 淋巴管和减少脑脊液(CSF)进入颈淋巴结内的引流。有趣的是， 我们还表明，脑膜周围淋巴管髓系细胞是apoE蛋白的主要局部来源，并成为 在中年APOE4小鼠中被激活。这些观察结果使我们提出了压倒一切的假设 APOE4的表达干扰脑膜固有髓系和淋巴管内皮细胞之间的串扰， 这将导致脑膜淋巴管的脑脊液引流减少，并加剧 阿尔茨海默病患者的神经炎症和脑淀粉样变性。在目标1中，我们计划使用构成或 研究APOE4对细胞自主和非自主影响的条件性基因敲除小鼠模型 不同年龄脑膜淋巴功能障碍的研究，确定潜在的先天免疫转录 特征，并确定淋巴生长因子和/或细胞因子的变化。在目标2中，我们将测试 脑膜淋巴引流早期损害促进认知功能衰退的假说 ApoE4小鼠，微调老年APOE4小鼠脑膜淋巴管可改善神经元功能和 以性别依赖的方式认知。为了评估这一点，将调节脑膜淋巴引流，使用腺- 促进淋巴管消融或扩张并评估其下游影响的相关病毒 老年男女APOE的神经转录体、突触可塑性和行为表现 人性化的老鼠。在目标3中，将测试操纵脑膜淋巴功能的治疗结果。 在敲击式阿尔茨海默病小鼠脑淀粉样变性模型中，APP-SAA表达APOE3或 载脂蛋白E4.在导致2个月和10个月大的小鼠脑膜淋巴管丢失或扩张后，我们将 评估不同的实验结果，包括行为表现、神经元活动、脑间质 流体Aβ(体内微透析法)、可溶性和不溶性Aβ聚集体以及先天和适应性的分布 大脑及其边缘组织中的免疫细胞激活。总之，这项提议将使我们能够发现小说 脑膜淋巴减少的机制，将成为未来项目的基石，并可能导致 治疗APOE4等位基因携带者的新疗法，这些携带者患阿尔茨海默病的风险更大。

ABSTRACT Aging and expression of apolipoprotein E ε4 (APOE4) gene alleles represent two major risk factors for Alzheimer’s disease. Recent studies have shown that aging leads to impaired brain fluid drainage by the bona fide lymphatic vascular network that is found in the mammalian meningeal dura. Interestingly, early loss of meningeal lymphatic drainage has been linked to worse amyloid-β (Aβ) pathology, vascular and microglial activation, and spatial memory deficits in Alzheimer’s disease mouse models of brain amyloidosis. However, little is known about the effects of APOE4 expression on meningeal lymphatic function. Our preliminary data shows that expression of APOE4 in humanized knock-in mice leads to a deleterious remodeling of meningeal lymphatic vessels and reduced drainage of cerebrospinal fluid (CSF) into the cervical lymph nodes. Interestingly, we also show that meningeal peri-lymphatic myeloid cells are a main local source of apoE protein and become activated in middle-aged APOE4 mice. These observations led us to raise the overarching hypothesis that expression of APOE4 disturbs the crosstalk between meningeal innate myeloid and lymphatic endothelial cells, which will subsequently lead to reduced CSF drainage by meningeal lymphatics, and exacerbated neuroinflammation and brain amyloidosis in Alzheimer’s disease. In Aim 1, we plan to use constitutive or conditional knockout mouse models to investigate the cell autonomous and non-autonomous effects of APOE4 on meningeal lymphatic dysfunction at different ages, determine the underlying innate immune transcriptional signatures, and identify alterations in lymphatic growth factors and/or cytokines. In Aim 2, we will test the hypothesis that an early impairment in meningeal lymphatic drainage promotes a faster cognitive decay in APOE4 mice, and that fine-tuning meningeal lymphatics in aged APOE4 mice ameliorates neuronal function and cognition in sex-dependent manner. To assess this, will modulate meningeal lymphatic drainage, using adeno- associated viruses that promote lymphatic vessel ablation or expansion, and evaluate the downstream effects on the neuronal transcriptomes, synaptic plasticity, and behavioral performances of aged female and male APOE humanized mice. In Aim 3, the therapeutic outcomes of manipulating meningeal lymphatic function will be tested in a knock-in Alzheimer’s disease mouse model of brain amyloidosis, the APP-SAA, expressing APOE3 or APOE4. After inducing meningeal lymphatic vessel loss or expansion in 2- and 10-month-old mice, we will evaluate different experimental outcomes including behavioral performance, neuronal activity, brain interstitial fluid Aβ (by in vivo microdialysis), soluble and insoluble Aβ aggregates, and the profiles of innate and adaptive immune cell activation in the brain and its border tissues. Altogether, this proposal will allow us to uncover novel mechanisms of meningeal lymphatic decline that will serve as cornerstones for future projects and might lead to novel therapies to treat APOE4 allele carriers that are at a greater risk of developing Alzheimer’s disease.