Heat shock proteins and neuroprotection in cerebral ischemia

热休克蛋白与脑缺血的神经保护

• 批准号: 8731306
• 负责人: Jun Chen
• 金额: --
• 依托单位: VETERANS HEALTH ADMINISTRATION
• 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 2019-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8731306
• 关键词: Actins; Adult; Apoptotic; Blood - brain barrier anatomy; Brain; Brain Injuries; Cells; Cellular Structures; Central Nervous System Diseases; Cerebral Ischemia; Cerebrum; Clinical; Cytoskeletal Modeling; Cytoskeleton; Data; Development; Disabled Persons; Disease; Edema; Endothelial Cells; Endothelium; Event; Exhibits; Extravasation; Future; Gelatinase A; Gene Targeting; Genes; Glucose; Heat Shock Protein 27; Heat shock proteins; Immune; Impairment; In Vitro; Infarction; Infiltration; Inflammation; Inflammation Mediators; Inflammatory; Ischemia; Ischemic Stroke; Lentivirus Vector; Life; Liquid substance; MMP2 gene; Mediating; Medical; Middle Cerebral Artery Occlusion; Military Personnel; Modeling; Molecular Chaperones; Mus; NF-kappa B; Nervous System Physiology; Neurologic; Neurological outcome; Outcome; Oxygen; Pathogenesis; Patients; Permeability; Plasma Proteins; Production; Protective Agents; Protein Family; Proteins; Quality of life; Reperfusion Injury; Reperfusion Therapy; Rodent; Role; Signaling Molecule; Stroke; Survivors; Testing; Therapeutic; Therapeutic Agents; Thrombolytic Therapy; Time; Transfection; Transgenic Mice; Transgenic Organisms; United States; Veterans; abstracting; aged; base; cell type; depolymerization; deprivation; disability; improved; in vivo; innovation; intravenous administration; intravenous injection; member; nervous system disorder; neuroprotection; novel; novel therapeutics; overexpression; polymerization; post stroke; prevent; protective effect; public health relevance; small molecule; stroke therapy; treatment strategy; vpr Genes; white matter injury; young adult

DESCRIPTION (provided by applicant): Abstract Stroke is a major medical concern for United States military veterans. Disruption of the blood-brain barrier (BBB) is a catastrophic event in the pathogenesis of ischemic/reperfusion (I/R) brain injury. Our recent studies suggested that structural alterations in brain endothelial cells (EC), including abnormal actin polymerization and the resulting redistribution of junctional proteins, is a novel mechanism responsible for early BBB leakage after I/R (30 min-3h); while matrix metalloproteinase (MMP) 2/9 activation, a predominant mechanism thought to contribute to post-stroke BBB disruption, contribute to the BBB leakage to larger molecules (>40kDa) in a relatively delayed manner. Therefore, restoring EC structure/function may offer an innovative therapeutic strategy for early BBB protection against I/R, while blocking MMP2/9 may provide a relatively delayed protection to BBB. Heat shock protein 27 (HSP27), a member of the small heat shock protein family, confers neuroprotection in several models of CNS diseases, including I/R brain injury. In addition to its well-known protein chaperone and anti-apoptotic functions, HSP27 may act as a potent actin depolymerization factor in certain cell types such as EC, thus potentially inhibiting actin polymerization-mediated BBB disruption. Further, HSP27 may inhibit the activation of NF-kappaB, a central signaling molecule for the production of MMP2/9 and pro-inflammatory mediators. We recently observed that HSP27 is transiently upregulated in brain EC after I/R. However, the precise role of HSP27 in the functional integrity of EC following I/R and the underlying mechanism remain unknown. Using transgenic mice overexpressing HSP27 and the lentiviral gene-transfection approach, we have obtained novel pilot data showing that HSP27 overexpression protects the endothelium from I/R-induced hyperpermeability in vitro and in vivo; that HSP27 inhibited oxygen glucose deprivation-induced actin polymerization and redistribution of junctional proteins in EC; and that intravenous administration of HSP27 containing a cell permeable transduction domain (TAT-HSP27) enabled rapid delivery of the protein into brain micro-vasculatures, reduced BBB damage and inhibited MMP2/9 activity after I/R. This proposal will further explore the BBB protective effect of HSP27 on I/R brain injury and elucidate the underlying mechanisms. The central hypothesis to be tested is that HSP27 protects BBB against I/R injury by stabilizing EC cytoskeletal organization in microvasculature early after I/R and inhibiting the production of MMP2/9 as well as pro-inflammatory proteins. The following specific aims are proposed: Aim 1: To determine whether endothelial targeted overexpression of HSP27 is sufficient to provide early protection on BBB integrity and to confer long term protection against cerebral I/R. Aim 2: Test the hypothesis that HSP27 protects endothelial integrity following I/R by dualistic mechanisms: stabilizing the actin cytoskeleton and inhibiting NFkB-dependent MMP2/9 production and inflammation. Aim 3: Test the hypothesis that post-stroke delivery of TAT-HSP27 to EC protects against BBB damage and improves outcomes in both young adult and aged mice. The proposed study attempts to develop HSP27 into a novel, clinically feasible therapeutic strategy to ameliorate post-stroke BBB damage, brake down the progression of brain damage, and improve long-term neurological functions in stroke victims. The successful completion of this proposed study will help improve the quality of life for veterans suffering from stroke.

描述（由申请人提供）： 摘要 中风是美国退伍军人面临的主要医疗问题。血脑屏障（BBB）的破坏是缺血/再灌注（I/R）脑损伤发病机制中的灾难性事件。我们最近的研究表明，脑内皮细胞 (EC) 的结构改变，包括异常的肌动蛋白聚合和由此产生的连接蛋白的重新分布，是导致 I/R（30 分钟至 3 小时）后早期 BBB 渗漏的新机制；而基质金属蛋白酶 (MMP) 2/9 激活（被认为是导致中风后 BBB 破坏的主要机制）会以相对延迟的方式导致 BBB 渗漏至较大分子 (>40kDa)。因此，恢复EC结构/功能可能为早期BBB针对I/R的保护提供创新的治疗策略，而阻断MMP2/9可能为BBB提供相对延迟的保护。 热休克蛋白 27 (HSP27) 是小热休克蛋白家族的成员，可在多种中枢神经系统疾病模型（包括 I/R 脑损伤）中提供神经保护作用。除了其众所周知的蛋白质伴侣和抗凋亡功能外，HSP27 还可以在某些细胞类型（如 EC）中充当有效的肌动蛋白解聚因子，从而潜在地抑制肌动蛋白聚合介导的 BBB 破坏。此外，HSP27 可能会抑制 NF-κB 的激活，NF-κB 是产生 MMP2/9 和促炎介质的中心信号分子。我们最近观察到 I/R 后脑 EC 中 HSP27 短暂上调。然而，HSP27 在 I/R 后 EC 功能完整性中的确切作用及其潜在机制仍不清楚。 使用过表达 HSP27 的转基因小鼠和慢病毒基因转染方法，我们获得了新的初步数据，表明 HSP27 过表达可以保护内皮细胞免受 I/R 诱导的体外和体内通透性过高的影响； HSP27 抑制氧葡萄糖剥夺诱导的肌动蛋白聚合和 EC 中连接蛋白的重新分布；静脉注射含有细胞通透性转导结构域 (TAT-HSP27) 的 HSP27 能够将蛋白质快速输送到脑微血管系统中，减少 I/R 后的 BBB 损伤并抑制 MMP2/9 活性。该提案将进一步探讨HSP27对I/R脑损伤的BBB保护作用并阐明其潜在机制。待测试的中心假设是，HSP27 通过在缺血再灌注后早期稳定微血管系统中的 EC 细胞骨架组织并抑制 MMP2/9 以及促炎蛋白的产生，从而保护 BBB 免受缺血再灌注损伤。提出以下具体目标： 目标 1：确定 HSP27 的内皮靶向过度表达是否足以为 BBB 完整性提供早期保护并针对脑 I/R 提供长期保护。 目标 2：检验 HSP27 通过二元机制保护 I/R 后内皮完整性的假设：稳定肌动蛋白细胞骨架和 抑制 NFkB 依赖性 MMP2/9 的产生和炎症。 目标 3：测试以下假设：中风后将 TAT-HSP27 递送至 EC 可防止年轻成年和老年小鼠的 BBB 损伤并改善结果。 拟议的研究试图将 HSP27 开发成一种新型的、临床上可行的治疗策略，以改善中风后 BBB 损伤，阻止脑损伤的进展，并改善中风患者的长期神经功能。这项拟议研究的成功完成将有助于改善患有中风的退伍军人的生活质量。