Microglia/macrophage polarization after intracerebral hemorrhage

项目总结： 脑出血后小胶质细胞/巨噬细胞极化 自发性脑出血(ICH)是一种致死率和致残率较高的疾病，但目前对其研究还不够深入。 与缺血性中风相比，缺乏有效的治疗方法。较大的血肿体积和扩大是 独立地与不良的患者预后相关；因此，快速清除有毒血液可以限制脑出血- 诱发性脑损伤。此外，小胶质细胞和巨噬细胞(M/MΦ)在脑出血后改变活动状态以清除 有毒的血液，并可能保护大脑。然而，过度激活的M/MΦ通过以下方式导致继发性脑损伤 释放细胞毒性物质。这些相反的影响可能是由不同的M/MΦ子集造成的，这些子集是 分为经典激活促炎(M1)和交替激活的抗炎(M2) 细胞。另一种激活的M2巨噬细胞表现出对凋亡细胞的吞噬作用增加，这可能 涉及清道夫受体CD36的激活和其负性调节因子Toll样受体(TLR)的抑制 4.重要的是，白介素10(IL-10)，一种抗炎细胞因子，抑制炎症，极化 巨噬细胞转化为M2c亚型，并增强吞噬功能。脑出血患者IL-10水平升高 在血液和脑组织中。然而，M/MΦ极化的范围和时间以及IL-10的确切作用 脑出血后M2极化的信号转导尚不清楚。我们研究的长期目标是限制脑出血的伤害和 改善功能结果。本R01的总体目标是调查M/MΦ的调制是否 IL-10的表型和功能减少了脑出血的损伤，改善了组织学和功能预后。我们的 初步研究表明，在脑片培养中，暴露于血红蛋白和 在体脑出血模型中IL-10促进脑片小胶质细胞吞噬和CD36表达 培养；IL-10缺陷小鼠血肿溶解受损，CD36和TLR4表达改变 与C57BL/6野生型小鼠相比，外源性IL-10成功地减少了血肿体积。这些 研究结果提示，IL-10将M/MΦ极化为M2表型可减轻脑出血损伤并改善 脑出血后组织学和功能恢复。在三个具体目标中，我们将确定M2小胶质细胞 IL-10的极化是暴露于血液成分的体外脑片培养中吞噬功能的原因 (目标1)：IL-10对M2 M/MΦ极化是否改善脑出血后的组织学和功能预后 (目标2)；IL-10诱导M/MΦM2极化是否需要激活CD36和抑制TLR4 (目标3)。这项研究获得的信息将为我们提供对M/MΦ极化的新见解 ICH及其IL-10信号诱导M/MΦM2极化的细胞和分子机制 减少脑出血的伤害。基于多学科的方法，我们的发现可能会导致一种新的治疗方法 不仅针对脑出血，而且针对其他脑部疾病。这一新的概念验证工作是为了研究调制 M/MΦ极化是最近的NINDS-SPRG确定的一个关键优先事项。

PROJECT SUMMARY: Microglia/macrophage polarization after intracerebral hemorrhage Spontaneous intracerebral hemorrhage (ICH) causes high mortality and morbidity, but it is understudied compared to ischemic stroke and lacks effective treatment. Greater hematoma volume and expansion are independently associated with poor patient outcomes; therefore, rapid removal of toxic blood could limit ICH- induced brain injury. Additionally, microglia and macrophages (M/MΦ) shift activity states after ICH to remove toxic blood and may protect the brain. However, over-activated M/MΦ cause secondary brain damage by releasing cytotoxic substances. These opposing effects may result from distinct M/MΦ subsets, which are categorized into classically activated proinflammatory (M1) and alternatively activated anti-inflammatory (M2) cells. Alternatively activated M2 macrophages exhibit increased phagocytosis of apoptotic cells, which could involve activation of the scavenger receptor CD36 and inhibition of its negative regulator, toll-like receptor (TLR) 4. Importantly, interleukin-10 (IL-10), an anti-inflammatory cytokine, represses inflammation, polarizes macrophages to an M2c subtype, and enhances phagocytosis. Patients with ICH have elevated levels of IL-10 in blood and brain tissue. However, the extent and timing of M/MΦ polarization and the exact role of IL-10 signaling in M2 polarization after ICH are unknown. The long-term goal of our research is to limit ICH injury and improve functional outcomes. The overall objective of this R01 is to investigate whether modulation of M/MΦ phenotype and function by IL-10 reduces ICH injury and improves histologic and functional outcomes. Our preliminary studies showed that IL-10 expression increases in brain slice cultures exposed to hemoglobin and in an in vivo model of ICH; that IL-10 increases microglial phagocytosis and CD36 expression in brain slice cultures; that IL-10-deficient mice have impaired hematoma resolution and altered CD36 and TLR4 expression compared to C57BL/6 wild-type mice; and that exogenous IL-10 successfully reduces hematoma volume. These findings prompt the hypothesis that polarizing M/MΦ to M2 phenotype by IL-10 reduces ICH injury and improves histologic and functional recovery after ICH. In three specific aims, we will determine whether M2 microglial polarization by IL-10 is responsible for phagocytosis in ex vivo brain slice cultures exposed to blood components (Aim 1); whether M2 M/MΦ polarization by IL-10 improves histologic and functional outcomes after ICH in vivo (Aim 2); and whether IL-10 induction of M/MΦ M2 polarization requires activation of CD36 and inhibition of TLR4 (Aim 3). The information gained from this study will provide us with novel insight into the M/MΦ polarization after ICH and the cellular and molecular mechanisms by which IL-10 signaling–induced M/MΦ M2 polarization reduces ICH injury. Based on multidisciplinary approaches, our findings will potentially lead to a new therapeutic strategy not only for ICH but also for other brain disorders. This novel proof-of-concept work to study modulation of M/MΦ polarization is a critical priority identified by the recent NINDS-SPRG.