Immunoregulatory role of mast cells in infections of the cerebrospinal fluid compartment: Investigations on a mouse model of pneumococcal meningitis

细菌性脑膜炎是全世界感染相关死亡的十大原因之一。在欧洲最常见的病原体是肺炎链球菌。充满脑脊液（CSF）的蛛网膜下腔的肺炎球菌感染产生医学上已知的最强大的炎症反应之一。虽然这种反应的分子机制已在很大程度上的特点，仍然是不确定的脑膜炎的细胞启动。在旨在首次深入了解肥大细胞在肺炎球菌性脑膜炎中的作用的初步实验中，我们观察到肥大细胞缺陷的W/W-v小鼠CSF细胞增多症意外显著增加，这是通过肥大细胞重建可逆的。脑细胞因子表达分析显示白细胞介素（IL）-1 β水平平行增加。用IL-1受体拮抗剂治疗W/W-v小鼠使CSF白细胞计数恢复至野生型小鼠中观察到的水平。这些数据表明，在肺炎球菌脑膜炎中，肥大细胞可能通过控制IL-1 β的生物活性来限制嗜酸性炎症。然而，脑膜肥大细胞限制IL-1 β水平的潜在机制需要澄清。本研究项目的目的是明确肺炎球菌脑膜炎中肥大细胞的功能。首先，我们计划评估一个额外的肥大细胞缺陷型小鼠品系，W-SH小鼠，肺炎球菌脑膜炎的表型。我们预计，W-sh小鼠将显示疾病的W/W-v小鼠的表型，提供进一步的证据表明，表型是由于肥大细胞缺陷，而不是其他小鼠品系相关的因素。此外，这将使我们能够在进一步的实验中使用W-sh小鼠。W-sh小鼠是可生育的，并且在充分研究的C57 BL/6遗传背景上，便于将转基因小鼠系用于计划的肥大细胞重建实验。在下一步中，我们计划确定负责肺炎球菌脑膜炎中肥大细胞抗炎活性的机制，从而关注肥大细胞衍生的抗炎细胞因子的作用（例如，IL-10）和丝氨酸肽酶（如类胰蛋白酶或糜蛋白酶），后者最近已被证明能够将几种细胞因子降解成无活性分子。最后，我们计划研究培养的肥大细胞对肺炎球菌攻击的反应。在此过程中，应在模拟CSF填充蛛网膜下腔特征的条件下培养骨髓源性肥大细胞。这些细胞计划用于研究[i]它们对肺炎球菌诱导的小鼠巨噬细胞释放IL-1 β的影响，以及[ii]它们降解成熟IL-1 β的潜力。我们认为，这项研究项目将显著提高我们对中枢神经系统内免疫调节机制的认识。

Bacterial meningitis is among the top 10 causes of infection-related deaths worldwide. The most frequent etiologic agent in Europe is Streptococcus pneumoniae. Pneumococcal infection of the cerebrospinal fluid (CSF)-filled subarachnoid space generates one of the most powerful inflammatory responses known in medicine. Although the molecular mechanisms underlying this response have been largely characterized, there is still uncertainty about the cellular initiators of meningeal inflammation. In pilot experiments aimed at getting first insight into the role of mast cells in pneumococcal meningitis, we observed an unexpected, marked increase in CSF pleocytosis in mast cell deficient W/W-v mice which was reversible by mast cell reconstitution. Analyses of brain cytokine expression revealed a parallel increase in interleukin (IL)-1ß levels. Treatment of W/W-v mice with an IL-1 receptor antagonist restored CSF leukocyte counts to levels seen in wild type mice. These data suggest that, in pneumococcal meningitis, mast cells may limit neutrophilic inflammation through control of IL-1ß bioactivity. The mechanism(s) underlying the restriction of IL-1ß levels by meningeal mast cells, however, need(s) to be clarified. The aim of this research project is to definitively characterize the function of mast cells in pneumococcal meningitis. In the first place, we plan to evaluate the phenotype of an additional mast cell-deficient mouse strain, the W-sh mice, with pneumococcal meningitis. We expect that W-sh mice will display the disease phenotype of W/W-v mice, providing further evidence that the phenotype is due to mast cell deficiency rather than other mouse strain-related factors. Moreover, it would enable us to use W-sh mice in further experiments. W-sh mice are fertile and on the well-studied C57BL/6 genetic background, facilitating the use of transgenic mouse lines for the planned mast cell reconstitution experiments. In the next step, we plan to identify the mechanisms responsible for the anti-inflammatory activity of mast cells in pneumococcal meningitis, thereby focussing on the role of mast cell-derived anti-inflammatory cytokines (e.g., IL-10) and serine peptidases (like tryptase or chymase), the latter of which have recently been shown to be able to degrade several cytokines into inactive molecules. Finally, we plan to investigate the response of cultured mast cells upon pneumococcal challenge. In doing so, bone marrow-derived mast cells shall be cultured under conditions mimicking features of the CSF-filled subarachnoid space. These cells are planned to be used for investigations of [i] their impact on the pneumococci-induced IL-1ß release by murine macrophages, as well as [ii] their potential to degrade mature IL-1ß. In our opinion, this research project will markedly improve our knowledge about mechanisms of immunoregulation within the central nervous system.