Role of bradykinin receptor 2 in tPA stroke therapy

项目摘要 组织纤溶酶原激活剂(TPA)的广泛使用，是FDA批准的唯一一种急性中风治疗方法， 由于其治疗时间窗口狭窄以及相关的脑出血风险，这种治疗仍然受到限制。缓激肽(BK)是 众所周知，它是一种内皮依赖性血管扩张剂和一种强大的促炎激动剂。一个 大量实验研究表明，卒中后BK受体上调是通过 卒中后早期的炎症效应，表明BK加重了脑损伤。在……里面 初步研究发现：1)tPA通过血浆激肽释放酶促进BK的形成。 激活；2)卒中后联合应用BK受体2拮抗剂和tPA治疗可减少 出血性转化；3)B2R激动剂在氧气和葡萄糖作用下不诱导内皮细胞死亡 剥夺；4)脾切除或巨噬细胞耗尽的小鼠使用tPA减少脑出血；5)BK 中风后输液增加出血转化，这在巨噬细胞耗竭的小鼠中被阻断。 拟议研究的目标是确定BK影响和激活的机制 TPA治疗后巨噬细胞，从而在血管形成过程中通过促炎过程导致脑出血 受伤。我们的最终目标是描述tPA治疗的机制基础，该治疗负责改善 并使tPA在中风患者中更安全和更广泛地使用。我们建议测试一下 以下具体目标：(I)确定条件性内皮细胞和巨噬细胞特异性的贡献 B2R缺失对tPA诱导的出血转化；(Ii)表征BK在活化中的作用 巨噬细胞在tPA处理后的聚集和募集；(Iii)阐明其分子机制 通过BK调节巨噬细胞的功能及其与脑内皮细胞的相互作用。我要用这个 指导职业发展奖(K01)，以实现一系列培训目标，这些目标建立在我之前的基础上 在神经科学方面的技能，但也在一些方式上扩展我的专业知识，这对我过渡到 独立调查员。这个应用程序的培训组件将基于我在免疫方面的专业知识 系统通过研究tPA治疗后脑出血所涉及的血管损伤。与培训一起 在分子免疫生物学方面，这将使我能够建立自己的中风尖端研究项目。 职业发展活动将由乔治·金博士与Magdy Selim博士合作指导， Kazuide Hawakaya和Stephan Kissler，因为他们的科学专长领域，以及技术和 我将从我们的指导关系中获得科学建议。我的职业发展活动将是 重点关注：1)实验室管理和组织方面的指导和指导；2)发展和 我的独立研究项目的发展；3)机构责任和满足以下要求 推广和扩大我的科学关系网；4)授予写作技能。

Project Summary The widespread use of tissue plasminogen activator (tPA), the only FDA-approved acute stroke treatment, remains limited by its narrow therapeutic time window and related risks of brain hemorrhage. Bradykinin (BK) is well known for its actions as an endothelial-dependent vasodilator and a powerful proinflammatory agonist. A number of experimental studies have show that BK receptors are upregulated after stroke by mediating inflammatory effects during early stages after stroke, indicating that BK aggravates cerebral injury. In preliminary studies, we found that: 1) BK formation is increased after tPA therapy through plasma kallikrein activation; 2) co-administration of a BK receptor 2 antagonist with tPA treatment after stroke reduces hemorrhage transformation; 3) B2R agonist does not induce endothelial cell death during oxygen and glucose deprivation; 4) splenectomized or macrophage-depleted mice with tPA decreased brain hemorrhage; 5) BK infusion after stroke increases hemorrhage transformation, which was blocked in macrophage-depleted mice. The objective of the proposed studies is to define the mechanisms by which BK affects and activates macrophages after tPA therapy, thus causing brain hemorrhage via proinflammatory processes during vascular injury. Our ultimate goal is to delineate the mechanistic underpinnings of tPA therapy responsible for improving the time window and to render tPA safer and more widely usable in patients with stroke. We propose to test the following specific aims: (i) to determine the contribution of conditional endothelium- and macrophage-specific B2R deletion to tPA-induced hemorrhage transformation; (ii) to characterize the role of BK in activated macrophages on accumulation and recruitment after tPA treatment; (iii) to delineate the molecular mechanisms by which BK regulates macrophage function and its interaction with brain endothelial cells. I will use this mentored career development award (K01) to fulfill a series of training objectives, which build upon my prior skills in neuroscience, but also expand my expertise in ways, which are essential to my transition to independent investigator. The training component of this application will build on my expertise in the immune system by investigating vascular injury involved in brain hemorrhage after tPA therapy. Together with training in molecular immunobiology, this will enable me to establish my own cutting-edge research program in stroke. The career development activities will be mentored by Dr. George King in collaboration with Drs. Magdy Selim, Kazuhide Hawakaya, and Stephan Kissler because of their areas of scientific expertise, and the technical and scientific advice that I stand to gain from our mentoring relationship. My career development activities will be focused on: 1) mentorship and guidance on laboratory management and organization; 2) the development and growth of my independent research program; 3) institutional responsibilities and fulfilling requirements for promotion and expanding my scientific network; 4) grant writing skills.