Meningeal Mast Cells: Key effectors of stroke pathology

描述（由申请人提供）：中风是美国严重长期残疾的主要原因，需要减少损伤和促进康复的治疗方法。脑卒中后炎症是损伤和恢复的关键决定因素，因此是一个有希望的治疗靶点。肥大细胞（MCs）在其他病理炎症过程的发展中起着关键作用，最近被认为在中风后脑炎症和损伤的加剧中起作用。这表明MCs可能是中风诱导炎症的关键决定因素，因此是一个有吸引力的治疗靶点。本提案的主要重点是阐明肥大细胞在加剧卒中病理中的作用机制，以确定卒中的新治疗策略为长期目标。我们的总体假设是，脑膜中的肥大细胞是中风病理的关键效应因子。这在Aim 1中使用两种不同的方法进行了测试。功能获得方法（Aim 1a）使用“肥大细胞敲入小鼠”模型，通过植入体外培养的肥大细胞，选择性地修复遗传MC缺陷小鼠的MC缺陷。将MCs直接植入这些mc缺陷小鼠的脑膜将确定位于脑膜的MCs在卒中病理中的参与程度。这是功能丧失方法（Aim1b）的补充，其中通过脑膜注射白喉毒素（DT）到一种新的小鼠模型中，在MCs上选择性表达DT受体，从而耗尽脑膜MCs。总之，这些小鼠模型的创新使用使我们能够确定脑膜MCs对中风后MCs的有害影响是否必要和充分。目的2利用肥大细胞敲入小鼠模型探讨MCs作用的分子机制。通过移植来自不同受体敲除小鼠的MCs，我们将研究脑膜MC激活的机制（Aim 2a），通过移植来自细胞因子敲除小鼠的MCs，我们将确定肥大细胞分泌因子对MCs对中风病理的下游影响的重要作用（Aim 2b）。确定脑卒中后脑膜MCs的关键作用将突出脑膜在调节脑病理中的重要性。由于脑膜相对容易接近（例如，通过鞘内注射），这一概念可能为中风治疗提供一种新的策略，可以克服将药物靶向损伤大脑的障碍，减少全身免疫调节的不良副作用。此外，通过建立MCs的作用机制，我们开始描绘参与调节中风反应的分子途径，这是寻找新的治疗靶点的重要一步。总之，我们提出的研究解决了对卒中后炎症事件的理解的重大空白，这些炎症事件有助于卒中病理学，并可能确定卒中治疗的新策略。

DESCRIPTION (provided by applicant): Stroke, a leading cause of serious long-term disability in the US, is in need of therapeutics that reduce damage and promote recovery. Post-stroke inflammation is a critical determinant of damage and recovery and is thus a promising therapeutic target. Mast cells (MCs), which play critical roles in the development of inflammatory processes in other pathologies, were recently ascribed a role in the exacerbation of post-stroke brain inflammation and damage. This suggests that MCs could be key determinants of stroke-induced inflammation and hence are an attractive therapeutic target. The primary focus of this proposal is to elucidate the mechanism of action of mast cells in exacerbating stroke pathology, with the long-term goal of identifying novel therapeutic strategies for stroke. Our overall hypothesis is that mast cells residing in the meninges are key effectors of stroke pathology. This is tested in Aim 1 using two different approaches. The gain-of-function approach (Aim 1a) uses a "mast cell knock-in mouse" model in which the MC-deficiency of genetically MC- deficient mice is selectively repaired by engraftment of in vitro grown mast cells. Direct engraftment of MCs into the meninges of these MC-deficient mice will determine the involvement of meningeal-located MCs in stroke pathology. This is complemented by the loss-of-function approach (Aim1b) where meningeal MCs are depleted by meningeal injection of diphtheria toxin (DT) into a novel mouse model that selectively expresses the DT receptor on MCs. Together, innovative use of these mouse models enables us to determine if meningeal MCs are necessary and sufficient for the detrimental effects of MCs after stroke. Aim 2 addresses the molecular mechanism of action of MCs using the mast cell knock-in mouse model. Through engraftment of MCs from various receptor knock-out mice we will investigate the mechanism of meningeal MC activation (Aim 2a), and by engraftment of MCs from cytokine knock-out mice we will identify mast cell-secreted factors important for the downstream effects of MCs on stroke pathology (Aim 2b). Identifying a crucial role for meningeal MCs after stroke will highlight the importance o the meninges in modulating brain pathology. As the meninges are relatively accessible (e.g., by intrathecal injection) this concept could potentially present a new strategy for stroke therapeutic that may overcome the hurdle of targeting drugs to the injured brain and reduce unwanted side effects of systemic immunomodulation. Furthermore, by establishing the mechanism of action of MCs we begin to delineate the molecular pathways involved in modulating the response to stroke, an essential step to finding novel therapeutic targets. Together, our proposed studies address significant gaps in the understanding of post-stroke inflammatory events that contribute to stroke pathology and may identify new strategies for stroke therapeutics.