Heat shock proteins in brain ischemia and stroke

描述(由申请人提供)： 中风在退伍军人中是一种常见的疾病，治疗方法很少。脑缺血经常导致血脑屏障(BBB)的破坏，导致脑水肿和出血，这可能会使本已灾难性的疾病复杂化。最近的研究表明，基质金属蛋白酶(MMPs)，一种锌依赖的内肽酶家族，参与了细胞外基质和血脑屏障的分解，导致脑水肿和出血。我们的实验室和我们的合作者的工作重点是热休克蛋白的保护潜力，即高度诱导的70kD热休克蛋白(HSP70)。由于其伴侣功能，HSP70似乎具有细胞保护特性，可能导致新生蛋白质折叠的增强和防止蛋白质聚集。然而，相关领域的研究表明，热休克蛋白也可能参与MMPs的表达和加工。我们最近发现，热应激诱导的HSP70过表达会在转录和翻译水平上降低培养的星形胶质细胞中MMPs的表达。除了阻止基质金属蛋白酶的转录和翻译外，我们初步发现HSP70还可以阻止原基质金属蛋白酶的活性形式的加工，这表明了另一种保护作用。在这个应用中，我们建议进一步探索这些观察结果，即HSP70通过抑制MMPs的表达、翻译和处理来防止血脑屏障破坏、水肿和出血，从而保护大脑免受中风的伤害。目的1将使用脑源性内皮细胞和星形胶质细胞的细胞培养来确定在不同刺激下哪种细胞类型产生HSP70和MMPs，以及HSP70是否可以防止体外模型中血脑屏障的破坏。目标2将确定HSP70如何与MMPs相互作用。缺乏功能ATPase或底物结合结构域的HSP70突变体将被研究。目的3探讨热休克蛋白对血脑屏障完整性、脑水肿/出血的意义，并利用实验性卒中动物模型对基质金属蛋白酶进行研究。 公共卫生相关性： 中风在退伍军人中是一种常见的疾病，治疗方法很少。中风经常并发于血脑屏障(BBB)的破坏，导致脑肿胀和出血。在这个项目中，我们建议研究中风后大脑中增加的一种蛋白质HSP70的潜在益处。HSP70通常不在大脑中存在，除非在中风等压力条件下。然而，卒中期间进一步增加HSP70可以减少继发性脑损伤的数量，并预防卒中的并发症。我们建议探索HSP70如何预防脑肿胀和出血，以便增加HSP70或给药HSP70本身的药理策略有朝一日可能对人类中风有用。由于中风是退伍军人中常见的医疗问题，这项提案的工作可能会导致治疗造福于这一人群。

DESCRIPTION (provided by applicant): Stroke is a common affliction among veterans, and treatments are few. Brain ischemia frequently causes disruption of the blood brain barrier (BBB) leading to brain edema and hemorrhage which can complicate an already catastrophic illness. Recent work in the field has implicated matrix metalloproteinases (MMPs), a family of Zn-dependent endopeptidases, in the breakdown of the extracellular matrix and BBB leading to brain edema and hemorrhage. Work by our labs and those of our collaborators' have focused on the protective potential of heat shock proteins, namely, the highly inducible 70 kD heat shock protein (HSP70). HSP70 appears to have cytoprotective properties by nature of its chaperone functions, presumably leading to enhancement of nascent protein folding and prevention of protein aggregation. However, work in related fields has shown that HSPs may also be involved in the expression and processing of MMPs. We recently showed that overexpression of HSP70 or its induction by heat stress reduced expression of MMPs in cultured astrocytes at the transcriptional and translational level. In addition to preventing MMP transcription and translation, we have preliminarily found that HSP70 can also prevent processing of pro-MMPs to their active form, suggesting an additional action of protection. In this application, we propose to further explore these observations that HSP70 protects the brain against stroke by preventing BBB disruption, edema and hemorrhage by inhibiting the expression, translation and processing of MMPs. Aim 1 will use cell cultures of brain derived endothelial cells and astrocytes to establish which cell types generate HSP70 and MMPs following various stimuli, and if HSP70 can prevent disruption of the BBB using in vitro models. Aim 2 will determine how HSP70 interacts with MMPs. HSP70 mutants lacking either functional ATPase or substrate binding domains will be studied. Aim 3 will explore the significance of HSPs on BBB integrity, brain edema/hemorrhage and MMPs will be studied using in vivo models of experimental stroke. PUBLIC HEALTH RELEVANCE: Stroke is a common affliction among veterans, and treatments are few. Stroke is frequently complicated by the disruption of the blood brain barrier (BBB) causing brain swelling and hemorrhage. In this project, we propose to study the potential beneficial effect of a protein that is increased in the brain after stroke, HSP70. HSP70 is not normally present in the brain except under stressful conditions such as stroke. However, further increasing HSP70 during stroke can reduce the amount of ensuing brain injury and prevent complications of stroke as well. We propose to explore how HSP70 prevents brain swelling and hemorrhage, in order that pharmacological strategies to increase HSP70 or to administer HSP70 itself might be a useful treatment for stroke in humans some day. Since stroke is a common medical problem among veterans, work from this proposal could potentially lead to treatments benefiting this population.