Table_1_Mast Cells Mediate Inflammatory Injury and Aggravate Neurological Impairment in Experimental Subarachnoid Hemorrhage Through Microglial PAR-2 Pathway.DOCX

Subarachnoid hemorrhage (SAH) is a devastating cerebrovascular disease with high mortality and disability. Aberrant neuroinflammation has been identified as a critical factor accounting for the poor prognosis of SAH patients. Mast cells (MCs), the sentinel cells of the immune system, play a critical in the early immune reactions and participate in multiple pathophysiological process. However, the exact role of MCs on the pathophysiological process after SAH has not been fully understood. The current study was conducted to determine the role of MCs and MC stabilization in the context of SAH. Mouse SAH model was established by endovascular perforation process. Mice received saline or cromolyn (MC stabilizer) or compound 48/80 (MCs degranulator). Post-SAH evaluation included neurobehavioral test, western blot, immunofluorescence, and toluidine blue staining. We demonstrated that SAH induced MCs activation/degranulation. Administration of MC stabilizer cromolyn conferred a better neurologic outcome and decreased brain edema when compared with SAH+vehicle group. Furthermore, cromolyn significantly inhibited neuroinflammatory response and alleviated neuronal damage after SAH. However, pharmacological activation of MCs with compound 48/80 dramatically aggravated SAH-induced brain injury and exacerbated neurologic outcomes. Notably, pharmacological inhibition of microglial PAR-2 significantly reversed MCs-induced inflammatory response and neurological impairment. Additionally, the effect of MCs-derived tryptase in mediating neuroinflammation was also abolished by the microglial PAR-2 blockage in vitro. Taken together, MCs yielded inflammatory injury through activating microglia-related neuroinflammation after SAH. These data shed light on the notion that MCs might be a novel and promising therapeutic target for SAH.

蛛网膜下腔出血（Subarachnoid hemorrhage, SAH）是一种极具破坏性的脑血管疾病，具有较高的病死率与致残率。异常神经炎症被证实是影响蛛网膜下腔出血患者预后不良的关键因素。肥大细胞（Mast cells, MCs）作为免疫系统的哨兵细胞，在早期免疫反应中发挥关键作用，并参与多种病理生理过程。然而，目前尚未完全阐明肥大细胞在蛛网膜下腔出血后病理生理过程中的具体作用。本研究旨在探讨肥大细胞及肥大细胞稳定化在蛛网膜下腔出血中的作用。研究采用血管内穿刺法构建小鼠蛛网膜下腔出血模型，小鼠分别接受生理盐水、色甘酸钠（MC稳定剂）或化合物48/80（MC脱颗粒剂）干预。蛛网膜下腔出血后评估指标包括神经行为学检测、蛋白质印迹（Western blot）、免疫荧光染色及甲苯胺蓝染色。本研究证实，蛛网膜下腔出血可诱导肥大细胞活化/脱颗粒。与蛛网膜下腔出血+溶剂对照组相比，给予肥大细胞稳定剂色甘酸钠可改善神经功能结局并减轻脑水肿。此外，色甘酸钠可显著抑制蛛网膜下腔出血后的神经炎症反应并减轻神经元损伤。然而，采用化合物48/80对肥大细胞进行药理学活化后，可显著加重蛛网膜下腔出血诱导的脑损伤并恶化神经功能结局。值得注意的是，对小胶质细胞蛋白酶激活受体2（PAR-2）进行药理学抑制，可显著逆转肥大细胞诱导的炎症反应及神经功能损伤。此外，体外实验中，小胶质细胞PAR-2阻断可消除肥大细胞源性类胰蛋白酶介导的神经炎症效应。综上，蛛网膜下腔出血后，肥大细胞通过激活小胶质细胞相关神经炎症通路引发炎症性损伤。本研究结果表明，肥大细胞有望成为蛛网膜下腔出血的新型潜在治疗靶点。