Alcohol, Glial Gene Expression and the Heat Shock Pathway

酒精、神经胶质基因表达和热休克途径

• 批准号: 7940991
• 负责人: NEIL L. HARRISON
• 金额: $ 23.91万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7940991
• 关键词: Acute; Alcohol consumption; Alcoholism; Alcohols; Astrocytes; Base Pairing; Behavioral; Binding; Bioinformatics; Blood; Brain; Brain Injuries; Cell Line; Cell Lineage; Cell Nucleus; Cells; Chemicals; Chronic; Cytokine Receptors; Cytoplasm; Defense Mechanisms; Elements; Environment; Exposure to; Gelshift Analysis; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Health; Heat shock proteins; Heat-Shock Response; Human; Immune; Impaired cognition; Lead; Maintenance; Measures; Methods; Microglia; Mus; Myelin; Myelin Associated Glycoprotein; Myelin Basic Proteins; Myelin Proteolipid Protein; Nerve; Neuraxis; Neuroglia; Neurons; Oligodendroglia; Oxidative Stress; PTGS2 gene; Pathway interactions; Physical Dependence; Physiological; Physiological Adaptation; Process; Proteins; Public Health; Quercetin; RNA Interference; Rattus; Reactive Oxygen Species; Regulation; Reporting; Research; Research Personnel; Response Elements; Reverse Transcriptase Polymerase Chain Reaction; Role; Secondary to; Signal Transduction; Speed; TLR4 gene; Western Blotting; Work; alcohol effect; alcohol exposure; alcohol measurement; alcohol response; alcohol sensitivity; brain cell; cis acting element; cytokine; drinking; glioma cell line; heat-shock factor 1; human NOS2A protein; immunocytochemistry; inhibitor/antagonist; neuron loss; neurotoxicity; novel; oligodendrocyte-myelin glycoprotein; problem drinker; public health relevance; research study; triptolide; white matter

DESCRIPTION (provided by applicant): Drinking alcohol results in a variety of short-term behavioral changes and long-term physiological adaptations that can lead to tolerance and physical dependence. Chronic exposure to alcohol has been reported to produce loss of neurons and white matter within the brain. Alcohol is known to trigger the release of cytokines and reactive oxygen species (ROS) from astrocytes and may have similar effects in microglia (MCG), as well as decreasing the synthesis of myelin by oligodendrocytes (OLGs). Alcohol regulates gene expression, and distinct sets of alcohol-responsive genes (ARGs) are activated by acute alcohol. We recently discovered, in the Gabra4 gene, an 11 base pair sequence element essential for the induction of gene transcription by alcohol - the alcohol response element (ARE). Stimulation of Gabra4 by alcohol involves the heat shock pathway, causing heat shock factor 1 (HSF1) to bind to the ARE (Pignataro et al., 2007). The ARE is present in a large number of other neuronal ARGs. Alcohol also activates the heat shock pathway in astrocytes. We now propose to extend these studies to OLG and MCG cells. Our overall hypothesis is that alcohol alters the expression of genes critical for the function of glial cells via activation of the heat shock pathway. The specific aims are: Aim 1) To study the activation of the heat shock pathway by alcohol in astrocytes. We will investigate activation of the heat shock pathway by alcohol in a mouse astrocyte cell line and in primary cultures of mouse astrocytes. We will measure the effects of alcohol on the expression of heat shock proteins (Hsps) and study the translocation of HSF1, and the interaction between HSF1 and the ARE in astrocyte nuclei. Aim 2) To identify alcohol-responsive genes in astrocytes. We will use a combination of bioinformatics and gene microarrays to identify novel ARGs in astrocytes, and then examine the subset of ARGs dependent on the ARE via the heat shock pathway. We will use pharmacological inhibitors of the heat shock pathway and RNAi to determine the involvement of HSF1. Aim 3) To study the effects of alcohol on gene expression in oligodendrocytes. We will study effects of alcohol on the expression of myelin- associated genes in an OLG cell line, investigate the effects of alcohol on expression of Hsps in OLGs and use pharmacological inhibitors to study the role of of HSF1 in the regulation of ARGs in OLGs. We will use bioinformatics, microarrays and Q-PCR to identify ARGs in OLGs. Aim 4) To study the effects of alcohol on gene expression in microglia. We will study the effects of alcohol on the expression of cytokine IL-1b (Il1b), the cytokine receptors, IL-1R1 (Il1r1) and TLR4 (Tlr4) and the enzymes iNOS (Nos2) and COX-2 (COX2) in a MCG cell line. We will investigate the effects of alcohol on the expression of Hsps in MCG and use pharmacological inhibitors to assess the role of HSF1. PUBLIC HEALTH RELEVANCE: Alcoholism is a major public health problem in the US. In the long-term, chronic drinking results in physiological changes that are detrimental to human health and have societal consequences. The brains of alcoholics become "rewired", and a part of this process results from changes in gene expression that occur in response to alcohol. We have shown that a cellular defense mechanism known as the heat shock pathway is activated by alcohol. In this proposal we plan to extend this work to study the changes within cells known as glia, the brain cells that support the activity of nerves within the brain. Astrocytes control the chemical environment of the nerve cells, and form a barrier between the brain and the blood. Oligodendrocytes form a sheath around nerve cells that speeds signal conduction within the brain. Microglia are the immune component of the brain and eliminate foreign invaders. Alcohol may alter the activity of all three types of glial cell by changing gene expression, and in doing so may harm nerve cells.

描述（由申请人提供）：饮酒会导致各种短期行为变化和长期的生理适应，这会导致耐受性和身体依赖性。据报道，慢性暴露于酒精会导致大脑内神经元和白质的丧失。已知酒精会引发星形胶质细胞中细胞因子和活性氧（ROS）的释放，并且可能在小胶质细胞（MCG）中具有相似的作用，并降低了少突胶质细胞（OLGS）对髓磷脂的合成。酒精调节基因表达，而不同的酒精反应基因（ARGS）被急性酒精激活。最近，我们在Gabra4基因中发现了一个11个基对序列元素对于通过酒精诱导基因转录至关重要的基本对序列元件 - 酒精反应元件（AS）。酒精对Gabra4的刺激涉及热休克途径，导致热休克因子1（HSF1）与AS结合（Pignataro等，2007）。其中存在于许多其他神经元中。酒精还激活星形胶质细胞中的热休克途径。现在，我们建议将这些研究扩展到OLG和MCG细胞。我们的总体假设是，酒精通过激活热冲击途径来改变对神经胶质细胞功能至关重要的基因的表达。具体目的是：目标1）研究星形胶质细胞中酒精对热冲击通路的激活。我们将研究小鼠星形胶质细胞系和小鼠星形胶质细胞的原发性培养物中酒精的热休克途径的激活。我们将测量酒精对热休克蛋白（HSP）表达的影响，并研究HSF1的易位，以及H​​SF1与植物之间的相互作用在星形胶质细胞核中。目标2）鉴定星形胶质细胞中的酒精反应性基因。我们将使用生物信息学和基因微阵列的组合来识别星形胶质细胞中的新颖ARG，然后通过热休克途径检查依赖于该的ARG的子集。我们将使用热休克途径和RNAi的药理抑制剂来确定HSF1的参与。目标3）研究酒精对少突胶质细胞基因表达的影响。我们将研究酒精对OLG细胞系中髓磷脂相关基因表达的影响，研究酒精对HSP在OLG中的表达的影响，并使用药理抑制剂研究HSF1在OLGS中ARGS调节中的作用。我们将使用生物信息学，微阵列和Q-PCR来识别OLG中的ARG。目标4）研究酒精对小胶质细胞基因表达的影响。我们将研究酒精对细胞因子IL-1B（IL1B），细胞因子受体，IL-1R1（IL1R1）和TLR4（TLR4）（TLR4）以及MCG细胞系中的COX-2（COX2）的影响。我们将研究酒精对HSP在MCG中表达的影响，并使用药理抑制剂评估HSF1的作用。 公共卫生相关性：酒精中毒是美国的主要公共卫生问题。从长远来看，长期饮酒会导致对人类健康有害并产生社会后果的生理变化。酗酒者的大脑成为“重新连接”，这一过程的一部分是由于响应酒精而发生的基因表达的变化而导致的。我们已经表明，一种称为热休克途径的细胞防御机制被酒精激活。在此提案中，我们计划扩展这项工作，以研究称为神经胶质的细胞内的变化，这是支持大脑内神经活性的脑细胞。星形胶质细胞控制神经细胞的化学环境，并在大脑和血液之间形成障碍。少突胶质细胞形成神经细胞周围的鞘，加速大脑内信号传导。小胶质细胞是大脑的免疫成分，消除了外国入侵者。酒精可能通过改变基因表达来改变所有三种类型的神经胶质细胞的活性，而这样做可能会损害神经细胞。

项目成果

Alcohol induces synaptotagmin 1 expression in neurons via activation of heat shock factor 1.

• DOI: 10.1016/j.neuroscience.2011.07.035
• 发表时间: 2011-10-13
• 期刊: NEUROSCIENCE
• 影响因子: 3.3
• 作者: Varodayan, F. P.;Pignataro, L.;Harrison, N. L.
• 通讯作者: Harrison, N. L.

The regulation of neuronal gene expression by alcohol.

• DOI: 10.1016/j.pharmthera.2009.09.002
• 发表时间: 2009-12
• 期刊: PHARMACOLOGY & THERAPEUTICS
• 影响因子: 13.5
• 作者: Pignataro, Leonardo;Varodayan, Florence P.;Tannenholz, Lindsay E.;Harrison, Neil L.
• 通讯作者: Harrison, Neil L.