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.

项目摘要： 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 病理状况，包括HIV感染，脑外伤和酒精使用障碍（AUD），已知 妥协BBB的完整性。饮酒烈酒[> 40％的酒精量（ABV）]很受欢迎 青少年和年轻人的活动。我们的初步研究表明酒精饮酒暴饮暴食 具有较高乙醇（ETOH）浓度的饮料会导致严重的BBB损害。因此，酒精过多 对于那些已经容易患有BBB功能障碍的人，消费构成了增加的风险。 脑米尔血管内皮细胞（BMVEC）是BBB的关键细胞成分。 BMVEC包含 紧密连接（TJ）配合物，由封闭式，claudins和Zonula occludens（ZO）组成，限制了 物质跨BBB的细胞细胞传递，并确保结构和功能完整性 BBB。 BMVEC的死亡（凋亡）是BBB损害的生物标志物，可能是由于酗酒而引起的。 但是，酒精影响BBB完整性恢复的机制尚未完全理解，但是 可能包括含酒精的BMVEC衬里的维修​​。虽然研究表明BMVEC的凋亡 可能造成BBB损坏，对BMVEC衬里的维修​​和 恢复BBB完整性。这主要是由于缺乏适当的实验动物模型 哪些BBB完整性已经被损害。 由链球菌分泌的毒素Intermedilysin（Ily）仅结合物种 - 特定的人类细胞膜蛋白，CD59（HCD59）。我们以前证明了将Ily注射到 在内皮细胞上表达HCD59的转基因小鼠菌株在A 剂量依赖性方式。我们最近建立了CRE诱导的FloxedStop-HCD59转基因小鼠系 （IHCD59）并用Tek-Cre越过诱导HCD59以生成复合小鼠（ihcd59+/ - /tek-cre +/-（+/-（+/-，，） 对于转基因的半合基），其中CRE表达驱动HCD59的表达专门在 内皮细胞。因此，我们可以使用ILY给药生成内皮细胞特异性消融模型。 内皮祖细胞（EPC）可以区分内皮细胞并修复受伤的内皮细胞 细胞衬里。根据文献和初步研究的数据，我们假设（a）酒精 - 诱导对已经受损的BBB内皮内皮壁的损害是EtOH浓度 - 依赖； （b）受损的内皮细胞衬里的修复和恢复被暴饮暴食延迟 饮酒。为了检验这一假设，我们提议利用我们的新型ILY介导的细胞消融 生成已经受损的BBB模型的技术。然后，我们将检查EtOH浓度 - 该模型中对损坏的BBB的依赖性影响，包括BBB的修复和恢复 内皮细胞衬里。该提案的两个具体目的是： 目标1：确定BBB内皮细胞衬里的损伤的机制 暴饮暴食之后。 目标2：确定BBB内皮细胞衬里修复的基础机制 暴饮暴食之后。 这项非常重要和临床相关的研究的发现将提供有关该研究的宝贵信息 酗酒进一步损害已经受到威胁的BBB的机制，以及 维修和恢复BBB完整性。