Alcohol-induced Impairment of Endothelial Cell Recovery

酒精引起的内皮细胞恢复损伤

• 批准号: 9238541
• 负责人: SULIE L. CHANG
• 金额: $ 21.59万
• 依托单位: SETON HALL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-01-03 至 2018-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9238541
• 关键词: Ablation; Adolescent and Young Adult; Affect; Alcohol abuse; Alcohol consumption; Alcohol-Induced Disorders; Alcoholic Beverages; Alcohols; Animal Model; Animals; Apoptosis; Beer; Behavior assessment; Binding; Biological; Biological Markers; Blood; Blood - brain barrier anatomy; Blood Vessels; Brain; Brain Injuries; Cell Membrane Proteins; Cells; Cessation of life; Coculture Techniques; Complex; Consumption; Data; Development; Disease; Dose; Endothelial Cells; Endothelium; Ensure; Ethanol; Experimental Animal Model; Exposure to; Functional disorder; HIV; HIV Infections; Head; Heavy Drinking; Hemorrhage; Hour; Human; Impairment; Individual; Injection of therapeutic agent; Investigation; Literature; Mediating; Modeling; Molecular; Mouse Strains; Mus; Necrosis; Neuraxis; Pathologic; Pattern; Proteins; Rattus; Recovery; Recovery of Function; Risk; Stem cells; Streptococcus intermedius; System; Techniques; Testing; Tight Junctions; Toxin; Transgenes; Transgenic Mice; Traumatic Brain Injury; alcohol effect; alcohol exposure; alcohol use disorder; alpha Toxin; base; binge drinking; brain health; brain repair; cell injury; clinically relevant; distilled alcoholic beverage; drinking; in vivo; injured; novel; novel therapeutic intervention; occludin; prevent; protein expression; repaired; restoration

Project Summary: Maintaining blood-brain barrier (BBB) integrity is essential for a healthy central nervous system. Many pathological conditions, including HIV infection, brain trauma, and alcohol use disorders (AUD), are known to compromise the integrity of the BBB. Binge drinking of hard liquor [>40% alcohol by volume (ABV)] is a popular activity among adolescents and young adults. Our preliminary studies indicate that binge drinking of alcoholic beverages with a high ethanol (EtOH) concentration can cause severe BBB damage. Thus, excessive alcohol consumption poses an increased risk for those individuals who are already susceptible to BBB dysfunction. Brain mircrovascular endothelial cells (BMVECs) are key cellular components of the BBB. BMVECs contain tight junction (TJ) complexes, consisting of occludin, claudins, and zonula occludens (ZO), that restrict paracellular passage of substances across the BBB and ensure the structural and functional integrity of the BBB. The death (apoptosis) of BMVECs is a biomarker of BBB damage, and can be caused by alcohol abuse. However, the mechanisms by which alcohol affects the recovery of BBB integrity are not fully understood, but may include alcohol-delayed repair of the BMVEC lining. While studies have shown that apoptosis of BMVECs can cause BBB damage, there has been little, if any, investigation into the repair of the BMVEC lining and the restoration of BBB integrity. This is due, mainly, to the lack of an appropriate experimental animal model in which BBB integrity is already compromised. Intermedilysin (ILY), a toxin secreted by Streptococcus intermedius, binds exclusively to the species- specific human cell membrane protein, CD59 (hCD59). We previously demonstrated that injection of ILY into a transgenic mouse strain that expresses hCD59 on endothelial cells induces rapid endothelial cell damage in a dose-dependent manner. We recently established a Cre-inducible floxedSTOP-hCD59 transgenic mouse line (ihCD59) and crossed the inducible hCD59 with Tek-cre to generate compound mice (ihCD59+/-/Tek-cre+/- (+/-, hemizygous for the transgene) in which Cre expression drives the expression of hCD59 specifically in endothelial cells. Thus, we can generate an endothelial cell-specific ablation model using ILY administration. Endothelial progenitor cells (EPCs) can differentiate into endothelial cells and repair an injured endothelial cell lining. Based on the literature and the data from our preliminary studies, we hypothesize that (a) alcohol- induced damage to the endothelial lining of an already compromised BBB is EtOH concentration- dependent; and (b) repair and restoration of the damaged endothelial cell lining is delayed by binge alcohol consumption. To test this hypothesis, we propose to utilize our novel ILY-mediated cell ablation technique to generate an already compromised BBB model. Then, we will examine EtOH concentration- dependent effects on the damaged BBB in that model, including the repair and restoration of the BBB endothelial cell lining. The two Specific Aims of this proposal are: Aim 1: To determine the mechanisms underlying damage to the endothelial cell lining of the BBB following binge exposure to various concentrations of EtOH. Aim 2: To determine the mechanisms underlying the repair of the endothelial cell lining of the BBB following binge exposure to various concentrations of EtOH. The findings from this highly significant and clinically relevant study will provide valuable information on the mechanisms by which alcohol abuse further damages an already compromised BBB, and the timeframe of repair and restoration of BBB integrity.

项目概要： 维持血脑屏障（BBB）的完整性对于健康的中枢神经系统至关重要。许多 已知包括HIV感染、脑创伤和酒精使用障碍（AUD）在内的病理状况， 危及BBB的完整性。狂饮烈性酒（酒精含量超过40%）是一种流行的 青少年和年轻人的活动。我们的初步研究表明， 具有高乙醇（EtOH）浓度的饮料可导致严重的BBB损伤。因此，过量的酒精 因此，对于那些已经对BBB功能障碍易感的个体来说，食用这种食物会增加风险。 脑微血管内皮细胞（BMVECs）是血脑屏障的关键细胞成分。BMVEC含有 紧密连接（TJ）复合体，由闭合蛋白、闭合蛋白和闭合小带（ZO）组成，限制 这一过程可促进物质的细胞旁通道穿过血脑屏障，并确保血脑屏障的结构和功能完整性。 BBB. BMVECs的死亡（凋亡）是BBB损伤的生物标志物，并且可以由酒精滥用引起。 然而，酒精影响血脑屏障完整性恢复的机制还不完全清楚， 可能包括BMVEC内衬的酒精延迟修复。虽然研究表明BMVECs的凋亡 可能会导致BBB损伤，很少有，如果有的话，调查BMVEC衬里的修复和 恢复BBB的完整性。这主要是由于缺乏适当的实验动物模型， 血脑屏障的完整性已经受损 中间体溶素（ILY）是由中间链球菌分泌的一种毒素，仅与该物种结合- 特异性人细胞膜蛋白，CD 59（hCD 59）。我们以前证明了将ILY注射到 在内皮细胞上表达hCD 59的转基因小鼠品系诱导快速内皮细胞损伤， 剂量依赖性我们最近建立了一个Cre诱导的hCD 59转基因小鼠系 将诱导型hCD 59与Tek-cre杂交以产生复合小鼠（ihCD 59 +/-/Tek-cre+/-（+/-， 其中Cre表达驱动hCD 59的表达，特异性地在 内皮细胞因此，我们可以使用ILY施用产生内皮细胞特异性消融模型。 内皮祖细胞（Endothelial progenitor cells，EPCs）可以分化为内皮细胞并修复损伤的内皮细胞 细胞内衬根据文献和我们初步研究的数据，我们假设（a）酒精- 对已经受损的BBB的内皮衬里的诱导损伤是EtOH浓度- 依赖性的;和（B）损伤的内皮细胞衬里的修复和恢复被暴食延迟 酒精消费。为了验证这一假设，我们建议利用我们的新ILY介导的细胞消融 技术来生成已经受损的BBB模型。然后，我们将检查EtOH浓度- 对该模型中受损BBB的依赖性作用，包括BBB的修复和恢复 内皮细胞衬里该提案的两个具体目标是： 目的1：探讨血脑屏障内皮细胞损伤的机制 在大量暴露于不同浓度的乙醇后 目的2：探讨血脑屏障内皮细胞衬里修复的机制 在大量暴露于不同浓度的乙醇后 这项具有高度重要意义和临床相关性的研究结果将为以下方面提供有价值的信息 酒精滥用进一步损害已经受损的BBB的机制，以及 修复和恢复BBB的完整性。