The effects of skull malformations and intracranial hypertension on the glymphatic and meningeal lymphatic systems in craniosynostosis

摘要： 我们的研究表明，人类与颅缝早闭，第二个最普遍的颅面疾病， 导致颅内高压（即压力升高）的颅骨和脑血管畸形。的影响 颅内压升高对脑脊液（CSF）循环和中枢神经系统液体平衡的影响 系统在颅缝早闭中是未知的，这是该领域的一个显著空白。我们在颅缝早闭动物中的新发现 模型提示颅骨发育不良和压力升高损害CSF流动，脑血管周围废物清除， 淋巴系统）和脑膜淋巴管的发育。注入荧光 CSF中的示踪剂显示循环途径发生了显著变化，流入脑组织和流出量减少 到周围的脑膜淋巴网络。脑膜炎，生长在硬脑膜上与头骨相连， 发育不全也不复杂有趣的是，高度分支的部分专门用于CSF的吸收和废物 被称为“热点”的清除工作发展不良和/或缺失。这些新发现意义重大，因为 大脑的胶质淋巴系统和支持脑膜淋巴管是防止β-淀粉样蛋白的必要条件 斑块积聚和认知障碍。总的来说，这些数据形成了一个中心假设：颅缝早闭 引起颅内压升高和CSF流体动力学改变，损害发育和功能 胶质淋巴系统和脑膜淋巴系统。这反过来又会影响废物和β- 大脑中的淀粉样蛋白我们使用我们创新的系统生物学方法来测试这一假设， 颅骨发育与颅内压、CSF流量以及大脑的胶质淋巴系统和淋巴系统有关。在 目的1，我们使用两种颅缝早闭的遗传小鼠模型来测试受损的胶质淋巴（即CSF）流入和 外排阻碍注射到脑中的β-淀粉样蛋白寡聚体的清除。接下来，我们测试β- 通过产生颅缝早闭和淀粉样蛋白沉积的小鼠模型来减少淀粉样蛋白斑块， 在家族性阿尔茨海默病中发现的五种突变。我们还测试了胶质淋巴瘤的机制基础， 通过检测水通道蛋白-4水通道极化至神经胶质终足， 很不安这些神经胶质（即胶质淋巴）水通道促进CSF流过血管周围空间， 因此，CSF和脑间质液之间的废物交换。在目标2中，我们测试是否 脑膜淋巴缺陷影响废物引流到颈部淋巴结。我们还探索了一种分子 通过检测VEGF-C的表达和VEGFR 3的活化， 受体信号传导受到影响。总的来说，我们的新系统和方法可以剖析 正常颅骨发育控制CSF流量、脑废物清除途径和 脑膜淋巴引流这项工作具有深远的临床意义， 颅缝早闭患者的交换和脑膜炎可能意味着β-淀粉样斑块形成的风险增加 和神经认知障碍。

Abstract: Our work has shown that humans with craniosynostosis, the second most prevalent craniofacial disorder, have skull and cerebrovascular malformations that cause intracranial hypertension (i.e. raised pressure). The effects of raised intracranial pressure on cerebrospinal fluid (CSF) circulation and fluid balance in the central nervous system are unknown in craniosynostosis, a notable gap in the field. Our new findings in craniosynostosis animal models suggest that skull dysplasia and raised pressure impair CSF flow, brain perivascular waste clearance pathways (the glymphatic system), and the development of meningeal lymphatic vessels. Infusing fluorescent tracers into CSF reveals significant changes to circulation pathways, and less influx into brain tissue and outflow to surrounding meningeal lymphatic networks. Meningeal lymphatics, which grow in dura attached to the skull, are hypoplastic and less complex. Interestingly, highly branched segments specialized for CSF uptake and waste removal, known as “hotspots”, are poorly developed and/or missing. These novel findings are significant because the brain's glymphatic system and supporting meningeal lymphatic vessels are necessary to prevent β-amyloid plaque buildup and cognitive impairment. Collectively, these data form a central hypothesis; craniosynostosis causes raised intracranial pressure and altered CSF hydrodynamics that impair the development and functions of both the glymphatic and meningeal lymphatic systems. This, in turn, affects the clearance of waste and β- amyloid from the brain. We test this hypothesis using our innovative systems biology approach that integrates skull development with intracranial pressure, CSF flow, and the brain's glymphatic and lymphatic systems. In Aim 1, we use two genetic mouse models for craniosynostosis to test if impaired glymphatic (i.e. CSF) influx and efflux impede the clearance of β-amyloid oligomers injected into the brain. Next, we test if the clearance of β- amyloid plaques is reduced by generating mouse models for craniosynostosis and amyloid deposition that express five mutations found in familial Alzheimer's disease. We also test a mechanistic basis for glymphatic dysfunction in craniosynostosis, by examining if the polarization of Aquaporin-4 water channels to glial endfeet is perturbed. These glial (i.e. glymphatic) water channels facilitate CSF flow through perivascular spaces in the brain, and therefore waste exchange between CSF and brain interstitial fluid. In Aim 2, we test whether meningeal lymphatic deficits affect waste drainage to the cervical lymph nodes. We also explore a molecular mechanism for meningeal lymphatic deficits by testing if VEGF-C expression and the activation of VEGFR3 receptor signaling is affected. Collectively, our novel system and approaches can dissect interlocking mechanisms through which normal skull development controls CSF flow, brain waste clearance pathways, and meningeal lymphatic drainage. This work has profound clinical significance as deficits in glymphatic waste exchange and meningeal lymphatics in craniosynostosis may imply increased risks for β-amyloid plaque buildup and neurocognitive impairment.