Glia-neuron interaction in fetal alcohol spectrum disorders

胎儿酒精谱系障碍中神经胶质细胞与神经元的相互作用

• 批准号: 9114822
• 负责人: Marina Guizzetti
• 金额: --
• 依托单位: PORTLAND VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-10-01 至 2018-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9114822
• 关键词: Affect; Afghanistan; Age; Alcohol abuse; Alcohol consumption; Alcohols; Alteplase; Area; Astrocytes; Award; Biochemical Reaction; Blood; Brain; Choline; Communication; Confocal Microscopy; DNA; DNA Methylation; DNA Modification Methylases; Deoxycytidine; Development; Enzymes; Epigenetic Process; Ethanol; Event; Extracellular Matrix; Extracellular Matrix Degradation; Extracellular Matrix Proteins; Female of child bearing age; Fetal Alcohol Exposure; Fetal Alcohol Spectrum Disorder; Fibronectins; Gene Expression; Healthcare Systems; Hippocampus (Brain); Immunohistochemistry; In Vitro; Investigation; Kidney; Label; Laminin; Lead; Learning; Lecithin; Liver; Measures; Mediating; Membrane; Mental Retardation; Methionine; Military Personnel; Mission; Neuroglia; Neuronal Plasticity; Neurons; Nutrient; Phospholipids; Play; Population; Postnatal Care; Pregnant Women; Protein Isoforms; Proteins; Recommendation; Research; Risk Behaviors; Role; Services; Soldier; Supplementation; Testing; Urokinase; Vertebral column; Veterans; Woman; Women' s Health; alcohol effect; alcohol exposure; child bearing; cholinergic; density; dietary supplements; experience; in vitro testing; in vivo; inhibitor/antagonist; laminin-1; men; neurochemistry; neuron development; novel; postnatal; premature; prenatal intervention; prevent; promoter; protein expression; public health relevance

DESCRIPTION (provided by applicant): In utero alcohol exposure may lead to Fetal Alcohol Spectrum Disorders (FASD), which cause learning deficits. Alcohol abuse is a well-known problem in the veteran population, which includes a growing number of women of childbearing age. Some of the effects of ethanol in the developing brain have been attributed to a premature loss of neuronal plasticity. Astrocytes play a major role in mediating neuronal plasticity and in the formation of neuronal connectivity during development. We have recently shown that cholinergic stimulation of astrocytes in vitro increases the levels of extracellular matrix (ECM) proteins laminin and fibronectin, which trigger neuritogenesis in hippocampal neurons while ethanol decreases the levels of neuritogenic ECM in astrocytes and astrocyte-induced neuritogenesis. Neuritogenic ECM proteins can be modulated by changes in their rate of degradation. In preliminary studies we found that tissue plasminogen activator (tPA) and urokinase plasminogen activator (uPA), involved in the degradation of the ECM, are upregulated by ethanol. Our studies also suggest that the epigenetic mechanism of DNA methylation may be involved in this effect of ethanol, as ethanol inhibits DNA methyltrasferases (DNMTs) and tPA promoter DNA methylation in astrocytes, events associated with increased gene expression. We hypothesize that ethanol-induced inhibition of DNMT activity in astrocytes increases the expression of proteins involved in ECM degradation and inhibits hippocampal plasticity. Consistent with our findings, DNA methylation is delayed by ethanol during development. Ethanol- induced DNMT activity and DNA methylation changes in astrocytes and their consequences on neurons are unexplored. The most promising treatment for FASD is choline, which can be converted to phosphatidylcholine (PC), a major membrane phospholipid, and, in the liver and kidney only, to methionine and then to s- adenosylmethionine (SAM), the main methyl donor in the enzymatic reaction of DNA methylation catalyzed by DNMT enzymes. The neurochemical mechanisms by which choline supplementation ameliorates hippocampal functions affected by prenatal alcohol exposure remains elusive. Preliminary studies show that, in vitro, astrocyte treatment with both choline and SAM prevents ethanol-induced DNMT inhibition. The overarching hypothesis of this study is that ethanol inhibits DNMT activity, decreases tPA and uPA promoter DNA methylation, increases expression and release of uPA and tPA, decreases neuritogenic ECM, and inhibits neuritogenesis/dendritic arborization in the developing hippocampus. Choline and SAM, by restoring DNMT activity, prevent the effects of ethanol on DNMT activity, uPA and tPA expression, ECM levels, and neuritogenesis/dendritic arborization. Aim 1 will characterize of the effects of ethanol treatments , with or without choline and SAM, and of DNMT inhibition in astrocytes on DNMT activity and expression, uPA and tPA expression, release, activity, promoter DNA methylation, and PC levels in astrocytes and on neuritogenesis. Aim 2 will investigate the effect of in vivo ethanol exposure from postnatal day (PD) 4 to PD 9 and choline treatments on hippocampal DNMT activity and expression, tPA and uPA expression and promoter DNA methylation, fibronectin and laminin protein levels, and dendritic arborization and spine density. These endpoints will be measured on PD 9 and on PD 30. The expression of fibronectin, laminin-1, tPA, uPA, and DNMT isoforms in astrocytes and neurons will be verified by immunohistochemistry. This study may unveil novel glia-mediated effects of ethanol on neuronal development and provide mechanisms by which choline protects hippocampal neuron development from the effects of ethanol by modulating astrocytic functions. 1

描述（由申请人提供）： 在子宫内接触酒精可能会导致胎儿酒精谱系障碍 (FASD)，从而导致学习障碍。酗酒是退伍军人中一个众所周知的问题，其中包括越来越多的育龄妇女。乙醇对发育中的大脑的一些影响归因于神经元可塑性的过早丧失。星形胶质细胞在介导神经元可塑性和发育过程中神经元连接的形成中发挥着重要作用。我们最近发现，体外星形胶质细胞的胆碱能刺激会增加细胞外基质（ECM）蛋白层粘连蛋白和纤连蛋白的水平，从而触发海马神经元的神经突发生，而乙醇则降低星形胶质细胞中的神经突原性ECM和星形胶质细胞诱导的神经突发生的水平。神经源性 ECM 蛋白可以通过其降解速率的变化进行调节。在初步研究中，我们发现参与 ECM 降解的组织纤溶酶原激活剂 (tPA) 和尿激酶纤溶酶原激活剂 (uPA) 被乙醇上调。我们的研究还表明，DNA 甲基化的表观遗传机制可能与乙醇的这种作用有关，因为乙醇会抑制星形胶质细胞中的 DNA 甲基转移酶 (DNMT) 和 tPA 启动子 DNA 甲基化，这些事件与基因表达增加相关。我们假设乙醇诱导的星形胶质细胞中 DNMT 活性的抑制会增加参与 ECM 降解的蛋白质的表达并抑制海马可塑性。与我们的研究结果一致，DNA 甲基化在发育过程中被乙醇延迟。乙醇诱导的星形胶质细胞中的 DNMT 活性和 DNA 甲基化变化及其对神经元的影响尚未被探索。 FASD 最有希望的治疗方法是胆碱，它可以转化为磷脂酰胆碱 (PC)（一种主要的膜磷脂），并且仅在肝脏和肾脏中转化为蛋氨酸，然后转化为 s-腺苷甲硫氨酸 (SAM)，这是胎儿的主要甲基供体。 DNMT酶催化DNA甲基化的酶促反应。补充胆碱改善受产前酒精暴露影响的海马功能的神经化学机制仍然难以捉摸。初步研究表明，在体外，用胆碱和 SAM 处理星形胶质细胞可防止乙醇诱导的 DNMT 抑制。本研究的总体假设是乙醇抑制 DNMT 活性，减少 tPA 和 uPA 启动子 DNA 甲基化，增加 uPA 和 tPA 的表达和释放，减少神经突 ECM，并抑制发育中的海马神经突发生/树突分枝。胆碱和 SAM 通过恢复 DNMT 活性，防止乙醇对 DNMT 活性、uPA 和 tPA 表达、ECM 水平以及神经发生/树突分枝化的影响。目标 1 将表征乙醇处理（有或没有胆碱和 SAM）以及星形胶质细胞中 DNMT 抑制对星形胶质细胞中 DNMT 活性和表达、uPA 和 tPA 表达、释放、活性、启动子 DNA 甲基化和 PC 水平的影响。神经发生。目标 2 将研究出生后第 4 天至 PD 9 的体内乙醇暴露和胆碱治疗对海马 DNMT 活性和表达、tPA 和 uPA 表达以及启动子 DNA 甲基化、纤连蛋白和层粘连蛋白水平以及树突分枝和脊柱密度。这些终点将在 PD 9 和 PD 30 上测量。星形胶质细胞和神经元中纤连蛋白、层粘连蛋白-1、tPA、uPA 和 DNMT 亚型的表达将通过免疫组织化学进行验证。这项研究可能揭示乙醇对神经元发育的神经胶质介导的新作用，并提供胆碱通过调节星形胶质细胞功能来保护海马神经元发育免受乙醇影响的机制。 1