MicroRNA Regulation of Phospholipid Homeostasis in Alzheimer's Disease Pathogenesis

MicroRNA 对阿尔茨海默病发病机制中磷脂稳态的调节

• 批准号: 10521283
• 负责人: Dongming Cai
• 金额: --
• 依托单位: JAMES J PETERS VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10521283
• 关键词: 3-Dimensional; Age Months; Alleles; Alzheimer' s Disease; Alzheimer' s disease pathology; Animals; Anti-Inflammatory Agents; Antiinflammatory Effect; Astrocytes; Attenuated; Autopsy; Axon; Behavioral; Brain; Brain region; Cells; Chelating Agents; Chronic; Coculture Techniques; Cognitive; Cognitive deficits; Complex; Copper; Cuprizone; Data; Data Set; Defect; Demyelinations; Development; Disease; Disease Progression; Down-Regulation; Enzymes; Etiology; Exhibits; Experimental Models; Female; Functional disorder; Funding; Gene Expression; Genes; Goals; Hippocampus; Homeostasis; Human; Impairment; Inflammation; Inflammatory; Inflammatory Response; Injections; Knock-in; Knock-in Mouse; Ligands; Lipopolysaccharides; Micro Array Data; MicroRNAs; Microglia; Modeling; Molecular; Molecular Profiling; Morphology; Mus; Myelin; Nature; Neurons; Pathogenesis; Pathogenicity; Pathology; Pathway interactions; Phagocytes; Phenotype; Phospholipids; Pilot Projects; Regulation; Resolution; Role; SYNJ1 gene; Sampling; Senile Plaques; System; TREM2 gene; Testing; Time; Viral; apolipoprotein E-3; apolipoprotein E-4; brain cell; cognitive performance; cohort; cytokine; dietary control; exosome; follow-up; genetic risk factor; glial activation; in vivo; induced pluripotent stem cell; inorganic phosphate; knock-down; male; motor deficit; mouse model; network models; neuroinflammation; novel; overexpression; prevent; protein expression; recruit; response; single-cell RNA sequencing; targeted treatment; tau Proteins; transcriptome sequencing; uptake; validation studies; vector

PROJECT SUMMARY APOE4 is the strongest genetic risk factor for sporadic AD with Ab-dependent and Ab-independent effects on disease pathogenesis. However, the molecular mechanisms underlying the pathogenic nature of APOE4 in AD are not fully elucidated. In previous funding period (07/01/2017-present), we have made significant progress toward understanding micro-RNA (miRNA) regulation of APOE4-induced brain phospholipid dysregulation in AD. We have uncovered a novel regulatory mechanism of miR-195 targeted at APOE4-associated cognitive deficits and lysosomal defects in AD. Notably, we identified miR-195 as a top miRNA candidate involved in the APOE- regulated brain phosphoinositol biphosphate (PIP2) pathway using human ROSMAP and mouse microarray data. Levels of miR-195 are significantly lower in APOE4+ human and mouse brains, and in human inducible pluripotent stem cells (iPSC)-derived neurons and astrocytes when compared to APOE4- counterparts. Over- expressing miR-195 reduces expression levels of its top target synaptojanin 1 (synj1), the brain PIP2 degrading enzyme. Elevating miR-195 ameliorates cognitive deficits and AD pathology in APOE4+ mice and rescues lysosomal defects in APOE4+ iPSC brain cells. Furthermore, our preliminary results support the role of miR-195 as an anti-inflammatory miRNA in regulating microglial function. Our single cell (sc)-RNA seq. analysis of E4FAD mouse brains with miR-195 over-expression suggests that miR-195 alters molecular signatures of microglia sub- clusters. APOE4+ microglia with lower miR-195 levels and higher synj1 expression at baseline, manifests with impaired phagocytic activities and lysosomal defects when compared to APOE3+ microglia. Down-regulation of synj1 or over-expression of miR-195 can rescue these phenotypes. Beside synj1, inflammatory genes pdcd4 and smad7 are predicted targets of miR-195 as well. Over-expression of miR-195 in microglia inhibits lipopolysaccharide (LPS)-induced increases in smad7 and pdcd4 expression, attenuates LPS-induced proinflammatory cytokine release and augments anti-inflammatory responses. In addition, exosomes derived from APOE4/4 astrocytes (ADEs) contain less miR-195 than those in APOE3/3 ADEs, and over-expression of miR-195 in APOE4/4 astrocytes increases miR-195 levels in ADEs which can attenuate LPS-induced pro- inflammatory cytokine release. Therefore, we hypothesize that miR-195 may exhibit anti-inflammatory effects through down-regulation of microglial synj1 to regulate lysosomal function, direct target at microglial inflammatory gene expression and responses, and modulation of neuro-inflammation and tau spread by exosomal miR-195. We propose to characterize the regulation of microglial function by miR-195 during AD pathogenesis in this renewal application. We will: 1) determine the impact of miR-195 on microglia function and APOE-regulated neuro-inflammation in AD in vivo (Aim 1) using cuprizone (CPZ)-induced inflammation in male and female EFAD mouse models (human ApoE4 knock-in at 5xFAD background); 2) to characterize the molecular mechanisms by which miR-195 regulates AD-associated neuro-inflammation (Aim 2) using microglial culture and 3-D co-culture system of mouse brain cells from EFAD and APOE KI mice, as well as from synj1-/-, APOE-/- and TREM2-/- mice (with manipulations of miR-195 levels); 3) to perform high resolution multiscale network modeling using scRNA- seq dataset from mouse brains (Aim 1) and RNA-seq and miR-seq datasets from microglia and 3-D co-culture system (Aim 2) to identify microglia-specific molecular signatures driven by miR-195; and 4) to validate identified microglial signature driven by miR-195 in postmortem human brain samples and investigate their correlation with the development of AD-associated neuro-inflammation during disease progression (Aim 2). The goals of this application aim to elucidate novel pathways and molecular signatures driven by miR-195 protective against APOE4-induced microglial dysfunction in AD pathogenesis, which will facilitate identification and development of a more personalized targeted therapeutic approach to AD-associated neuro-inflammation.

项目概要 APOE4 是散发性 AD 最强的遗传危险因素，对 Ab 依赖性和非 Ab 依赖性的影响 疾病发病机制。然而，APOE4 在 AD 中致病的分子机制 尚未完全阐明。在上一个资助期间（2017年7月1日至今），我们取得了重大进展 旨在了解 AD 中 APOE4 诱导的脑磷脂失调的 micro-RNA (miRNA) 调节。 我们发现了一种针对 APOE4 相关认知缺陷的 miR-195 新型调控机制 AD 中的溶酶体缺陷。值得注意的是，我们确定 miR-195 是参与 APOE- 的顶级 miRNA 候选者。 使用人类 ROSMAP 和小鼠微阵列数据调节脑磷酸肌醇二磷酸 (PIP2) 通路。 APOE4+ 人类和小鼠大脑以及人类诱导型大脑中 miR-195 的水平显着降低 与 APOE4 对应物相比，多能干细胞 (iPSC) 衍生的神经元和星形胶质细胞。超过- 表达 miR-195 会降低其首要靶标 synaptojanin 1 (synj1) 的表达水平，大脑 PIP2 降解 酶。升高 miR-195 可改善 APOE4+ 小鼠的认知缺陷和 AD 病理并挽救 APOE4+ iPSC 脑细胞中的溶酶体缺陷。此外，我们的初步结果支持 miR-195 的作用 作为调节小胶质细胞功能的抗炎 miRNA。我们的单细胞 (sc)-RNA 测序。 E4FAD分析 miR-195 过度表达的小鼠大脑表明 miR-195 改变了小胶质细胞亚群的分子特征 集群。 APOE4+ 小胶质细胞在基线时具有较低的 miR-195 水平和较高的 synj1 表达，表现为 与 APOE3+ 小胶质细胞相比，吞噬细胞活性受损和溶酶体缺陷。下调 synj1 或 miR-195 的过度表达可以挽救这些表型。除了 synj1 之外，还有炎症基因 pdcd4 和 smad7 也是 miR-195 的预测靶标。小胶质细胞中 miR-195 的过度表达抑制 脂多糖 (LPS) 诱导的 smad7 和 pdcd4 表达增加，减弱 LPS 诱导的 促炎细胞因子释放并增强抗炎反应。此外，外泌体衍生 来自 APOE4/4 星形胶质细胞 (ADE) 的 miR-195 含量比 APOE3/3 ADE 中的要少，并且过表达 APOE4/4 星形胶质细胞中的 miR-195 会增加 ADE 中的 miR-195 水平，从而减弱 LPS 诱导的促 炎症细胞因子释放。因此，我们推测miR-195可能表现出抗炎作用 通过下调小胶质细胞synj1调节溶酶体功能，直接针对小胶质细胞炎症 基因表达和反应，以及外泌体 miR-195 对神经炎症和 tau 扩散的调节。 我们建议在 AD 发病过程中表征 miR-195 对小胶质细胞功能的调节 续订申请。我们将：1) 确定 miR-195 对小胶质细胞功能和 APOE 调节的影响 使用铜宗 (CPZ) 在男性和女性 EFAD 中诱导炎症来研究 AD 体内神经炎症（目标 1） 小鼠模型（5xFAD 背景下的人类 ApoE4 敲入）； 2) 表征分子机制 使用小胶质细胞培养和 3-D 共培养，miR-195 调节 AD 相关神经炎症（目标 2） 来自 EFAD 和 APOE KI 小鼠以及 synj1-/-、APOE-/- 和 TREM2-/- 小鼠的小鼠脑细胞系统 （通过操纵 miR-195 水平）； 3) 使用 scRNA 执行高分辨率多尺度网络建模 来自小鼠大脑的 seq 数据集（目标 1）以及来自小胶质细胞和 3-D 共培养的 RNA-seq 和 miR-seq 数据集 系统（目标 2）识别由 miR-195 驱动的小胶质细胞特异性分子特征； 4) 验证已识别的 死后人脑样本中由 miR-195 驱动的小胶质细胞特征，并研究它们与 疾病进展过程中 AD 相关神经炎症的发生（目标 2）。本次活动的目标 该申请旨在阐明由 miR-195 驱动的新途径和分子特征，以预防 APOE4 诱导的 AD 发病机制中的小胶质细胞功能障碍，这将有助于识别和发展 针对 AD 相关神经炎症的更加个性化的靶向治疗方法。