Alcohol Effects on the Proteome and Transcriptome of Fetal Neural Stem Cell-Derived Extracellular Vesicles: Mechanism for Alcohol Teratogenesis

酒精对胎儿神经干细胞来源的细胞外囊泡蛋白质组和转录组的影响：酒精致畸机制

• 批准号: 10292946
• 负责人: Dae Chung
• 金额: $ 3.45万
• 依托单位: TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10292946
• 关键词: Address; Adult; Affect; Alcohol abuse; Alcoholism; Alcohols; Biological; Biological Assay; Brain; Cells; Cessation of life; Competence; Data; Data Analytics; Developmental Disabilities; Differentiation and Growth; Dose; Endocrine; Ethanol; Eukaryotic Cell; Exhibits; Exposure to; Family; Fetal Alcohol Exposure; Fetal Alcohol Spectrum Disorder; Fetal Development; Fetal alcohol effects; Gene Activation; Gene Expression; Goals; Growth; Human; Immunoprecipitation; Impairment; Inflammation; Inflammatory; Inflammatory Response; Inflammatory Response Pathway; Institutes; Intervention; Liquid Chromatography; Mediating; Mediator of activation protein; Messenger RNA; MicroRNAs; Mission; Mitochondrial Proteins; Molecular; Molecular Chaperones; Multivariate Analysis; Mus; NFIA gene; Neurons; Nonsense Codon; Nonsense-Mediated Decay; Nucleic Acids; Organelles; Parents; Pathway interactions; Pattern; Pattern recognition receptor; Pregnancy; Prevention; Proteins; Proteome; Publishing; RNA; RNA-Binding Proteins; Research; Research Methodology; Role; Scientist; Second Pregnancy Trimester; Sex Differences; Signal Transduction; Stress; Time; Training; Transcript; Translating; Translation Initiation; Translations; alcohol effect; alcohol exposure; alcohol related problem; alcohol response; career; cytokine; data analysis pipeline; disability; extracellular vesicles; fetal; inhibitor/antagonist; intercellular communication; interest; mouse model; nanoscale; nerve stem cell; neurobehavioral; neurodevelopment; neurogenesis; neuromechanism; neuroprotection; non-genetic; novel; overexpression; protein complex; protein expression; receptor; relating to nervous system; response; sex; skills; stem cell division; stem cell growth; stem cell proliferation; stem cell survival; stem cells; tandem mass spectrometry; teratogenesis; tool; transcription factor; transcriptome; transcriptome sequencing; transcriptomics; vesicular release

Project Summary Prenatal alcohol exposure (PAE) can result in decreased neurogenic capacity of neural stem cells (NSCs), which, during the early-to-mid gestation period, give rise to most neurons of the adult brain. This loss of neurogenic capacity explains, in part, neurobehavioral and brain growth deficits associated with Fetal Alcohol Spectrum Disorders (FASD). However, the mechanisms that mediate the persistent effects of episodic PAE on fetal NSCs are unknown. We and others have recently identified a novel class of sub-200 nanometer-sized extracellular vesicles (EVs) secreted by NSCs, as a means for intercellular transfer of biomolecules including proteins and nucleic acids. I hypothesize that EVs constitute endocrine-like organelles that propagate the effects of ethanol throughout the stem cell microenvironment in time and space. My preliminary and published data support the hypotheses that ethanol (i) alters the sequestration of proteins into EVs released by fetal NSCs, in a dose-dependent manner, and, by interfering with protein expression in EVs, (ii) results in loss of stem cell renewal and diminished neurogenesis. Using mass spectrometric and transcriptomic tools and cell and molecular biological analyses of EV function in a murine model, planned and completed studies address the following specific aims: Aim 1 seeks to identify and validate the effects of alcohol exposure on proteins and RNAs in EVs and associated NSCs. My studies show that EVs from parent NSCs exposed to moderate doses of ethanol (Mod-EtoHEVs) exhibit a significant increase in proteins of the Nonsense- Mediated Decay (NMD), whereas EVs obtained following NSC exposure to higher levels of ethanol (Hi-EtoHEVs) overexpress mitochondrial proteins that constitute a Danger-Associated Molecular Pattern (mito-DAMP) signal. These data will be validated and compared to the ethanol-affected intracellular proteome of parent NSCs. Aim 2 will evaluate the mediating role of Mod-EtoHEV-derived NMD on NSC proliferation, differentiation, and death, under basal and ethanol-exposure conditions. Our working hypothesis is that Mod-EtoHEVs transfer NMD components to recipient cells to confer compensatory neuroprotection. Aim 3 will evaluate the mediating role of Hi-EtoHEV-derived mito-DAMP on NSC proliferation, differentiation, and death. Our working hypothesis is that the secretion of a ‘danger’ signal in response to high levels of ethanol exposure triggers a pro-inflammatory cytokine response in recipient NSCs, resulting in impaired neurogenesis. My studies are expected to uncover novel intercellular communication pathways that mediate the effects of PAE on early neural maturation. This training plan will equip me with technical and research methodology skills and develop specific competency with data analytic and statistical approaches for proteome and transcriptome assessment. The plan’s focus on dissecting and disrupting the connection between PAE and FASD integrates well with my career goal to become a scientist with a focus on the role of early environmental mediators of developmental disability.

项目摘要 产前酒精暴露（PAE）可以改善神经元干细胞（NSC）， 在早期到中间的妊娠期，这会引起成人大脑的大多数神经元。这种损失 神经源能力部分解释了与胎儿酒精相关的神经行为和脑生长定义 频谱障碍（FASD）。但是，介导情节PAE持续作用的机制 胎儿NSC是未知的。我们和其他人最近确定了一类新型纳米尺寸的新型 NSC分泌的细胞外蔬菜（EV），作为生物分子细胞间转移的一种手段 蛋白质和核酸。我假设电动汽车构成类似内分泌的细胞器，从而传播效果 整个干细胞微环境的乙醇在时间和空间中。 我的初步和发布的数据支持乙醇（i）改变蛋白质的假设 以剂量依赖性的方式释放出胎儿NSC释放的电动汽车，并通过干扰蛋白质表达 EV，（ii）导致干细胞更新和神经发生减少。使用质谱和 鼠模型中EV功能的转录组工具以及细胞和分子生物学分析，计划和 完成的研究解决了以下特定目的：目标1试图识别和验证酒精的影响 电动汽车和相关NSC中蛋白质和RNA的暴露。我的研究表明，父母NSC的电动汽车 暴露于中等剂量的乙醇（Mod-Etohevs）暴露于胡说八道的蛋白质显着增加 介导的衰减（NMD），而在NSC暴露于较高水平的乙醇（HI-Etohevs）之后获得的电动汽车获得 构成与危险相关的分子模式（MITO-DAMP）信号的过表达线粒体蛋白。 这些数据将经过验证并将其与受母体NSC的受乙醇影响的细胞内蛋白质组进行比较。目的 2将评估Mod-Etohev衍生的NMD在NSC增殖，分化和死亡中的中介作用， 在基本和乙醇暴露条件下。我们的工作假设是Mod-Et-Etohevs转移NMD 受体细胞的组件与会议补偿性神经保护作用。 AIM 3将评估中介作用 Hi-Etohev衍生的Mito-DAMP关于NSC增殖，分化和死亡。我们的工作假设是 对高水平乙醇暴露的响应的“危险”信号的分泌会触发促炎性细胞因子 受体NSC的反应，导致神经发生受损。 我的研究预计将发现新的细胞间通信途径，以介导PAE的影响 关于早期神经饱和。该培训计划将使我具备技术和研究方法论技巧和 使用蛋白质组和转录组的数据分析和统计方法开发特定的能力 评估。该计划的重点是解剖和破坏PAE和FASD集成之间的联系 我的职业目标是成为一名科学家，重点是 发育障碍。