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，患有颅缝早闭的人，第二种最常见的头面部疾病， 导致颅内高压(即压力升高)的头骨和脑血管畸形。其影响 颅内压升高对中枢神经系统脑脊液循环和液体平衡的影响 在颅缝融合系统尚不清楚，这是该领域的一个显著空白。我们在颅缝早闭动物身上的新发现 模型表明，颅骨发育不良和压力升高损害了脑脊液的流动，脑血管周围废物的清除 途径(淋巴系统)和脑膜淋巴管的发育。注入荧光 进入脑脊液的示踪剂显示循环途径发生了显著变化，流入和流出脑组织的量减少 到周围的脑膜淋巴网络。脑膜淋巴管生长在与头骨相连的硬脑膜中， 发育不全且不那么复杂。有趣的是，高度分枝的节段专门用于脑脊液的吸收和排泄 清除，即所谓的“热点”，不发达和/或缺失。这些新发现意义重大，因为 大脑的淋巴系统和支持脑膜的淋巴管是预防β-淀粉样蛋白所必需的 斑块堆积和认知障碍。总而言之，这些数据形成了一个中心假设：颅缝早闭 导致颅内压升高和脑脊液流体动力学改变，从而损害发育和功能 淋巴系统和脑膜淋巴系统。这反过来又会影响废物和β的清除- 大脑中的淀粉样蛋白。我们使用创新的系统生物学方法来验证这一假设，该方法将 颅骨发育与颅内压、脑脊液流量以及大脑的淋巴系统和淋巴系统有关。在……里面 目的1，我们使用两种遗传性的小鼠颅缝早闭模型来测试淋巴(即脑脊液)内流受损和 外流阻碍了注射到大脑中的β-淀粉样寡聚体的清除。接下来，我们测试β的清关是否- 通过建立颅缝闭合和淀粉样蛋白沉积的小鼠模型来减少淀粉样斑块 表达在家族性阿尔茨海默病中发现的五种突变。我们还测试了语音学的一个力学基础 通过检查水通道蛋白-4的水通道是否极化到神经胶质终足，研究颅缝融合症的功能障碍 感到心烦意乱。这些神经胶质(即淋巴管)水通道促进脑脊液通过血管周围间隙。 因此，脑脊液和脑间质液体之间的废物交换。在目标2中，我们测试 脑膜淋巴缺陷会影响排泄物向颈部淋巴结的引流。我们还探索了一种分子 检测血管内皮生长因子C表达和血管内皮生长因子受体3激活的脑膜淋巴功能缺陷机制 受体信号会受到影响。总体而言，我们的新系统和方法可以剖析联锁 正常颅骨发育控制脑脊液流动、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，因为淋巴排泄物的缺陷 颅缝融合症患者的交换和脑膜淋巴管可能意味着β-淀粉样斑块堆积的风险增加 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.