In vivo imaging of astrocytes to investigate their functional and structural interactions with the blood brain barrier, immune cells and the demyelinating axon in the animal model of Multiple Sclerosis.

多发性硬化症（MS）是最常见的中枢神经系统（CNS）慢性炎症性疾病，在北美和欧洲的年轻人中导致长期和严重的残疾。神经轴突结构的早期损伤已被认为是多发性硬化症病理的一个主要方面，并与后来的残疾密切相关。然而，目前尚不清楚为什么随着时间的推移，残疾的累积程度在个体患者之间有所不同。最近的报道表明，残疾积累的差异可能源于保护中枢神经系统谷氨酸稳态的遗传变异。我们最近证明了CD4+ T细胞对神经元损伤形成的直接影响。在我们的研究中，免疫细胞介导的神经元损伤的一个重要机制是谷氨酸介导的兴奋毒性。谷氨酸稳态是星形胶质细胞的一项主要任务，它具有从细胞外空间清除兴奋毒性物质的能力，并负责局部限制递质释放。迄今为止，星形胶质细胞是多发性硬化症发病机制中最被忽视的参与者之一，被认为是疤痕形成细胞的旁观者角色。我们的目的是更好地了解星形胶质细胞在神经炎症条件下的病理生理参与。我们将研究星形胶质细胞在EAE病变中对神经元和髓鞘损伤的作用。我们主要对星形胶质细胞在炎症中枢神经系统中轴突-胶质细胞单位的代谢和解毒作用感兴趣。我们将使用不同的转基因模型来分析神经元/轴突和/或星形胶质细胞中的Ca2+动力学。此外，我们将使用不同的药理学和转基因方法来干扰星形胶质细胞生物学，以分析对轴突完整性的影响。我们将采用常规分子生物学方法对EAE诱导后不同天数的星形胶质细胞进行分析。我们将在遗传MS易感性和疾病进展研究结果的背景下，对动物模型的结果进行批判性评估。此外，我们将在自己的患者队列中证实这一数据。此外，我们计划在体内描述星形胶质细胞在发展中的EAE病变中的作用，重点研究血脑屏障（BBB）及其与免疫细胞的相互作用。我们将利用双光子显微镜成像方法研究EAE病变血脑屏障处星形胶质细胞与免疫细胞的相互作用。在这里，重点将放在发展中的病变上，即在发病前和临床体征出现时，以研究星形胶质细胞在这些病变的病理生理中的早期参与。这些实验将突出星形胶质细胞在炎症中枢神经系统病变演变、损伤、保护和/或修复中的整体作用。这些发现可能为阻止多发性硬化症患者的神经变性和进展性残疾开辟新的治疗靶点。

Multiple sclerosis (MS) is the most common chronic inflammatory disease of the central nervous system (CNS) that causes prolonged and severe disability in young adults in North America and Europe. Early damage of neuroaxonal structures has been recognized as a major aspect of MS pathology and correlates best with later disability. However, it is not clear why the extent of accumulation of disability over time varies between individual patients. Recent reports suggest that differences in disability accumulation might derive from genetic variations in safeguarding CNS glutamate homeostasis. We have recently demonstrated a direct effect of CD4+ T cells on neuronal damage formation. One important mechanism of immune cell mediated neuronal damage in our study was (glutamate-mediated) excitotoxicity. Glutamate homeostasis is a major task of astrocytes, which have the capacity to remove excitotoxic substances from the extracellular space and are responsible for local confinement of transmitter release. Up to date astrocytes range among the most neglected players in MS pathogenesis with an alleged bystander role as scar forming cells. We aim to better understand the involvement of astrocytes in the pathophysiology in neuroinflammatory conditions. We will investigate the role of astrocytes in established EAE lesions in respect to neuronal and myelin damage. We are primarily interested in the metabolic and detoxification role of astrocytes for the axono-glial unit in the inflamed CNS. We will use different transgenic models to analyze Ca2+ dynamics in neurons/axons and/or astrocytes. Furthermore we will interfere with astrocyte biology using different pharmacological and transgenic approaches to analyze the effect on axonal integrity. We will analyze FACS-sorted astrocytes from different days after EAE induction by conventional molecular biology methods. We will critically evaluate the results from the animal model on the background of the results from genetic MS susceptibility and disease progression studies. In addition, we will confirm this data in an own patient cohort. Furthermore, we plan to characterize in vivo the role of astrocytes in the developing EAE lesion with focus on the blood brain barrier (BBB) and their interaction with immune cells. We will use our two-photon microscopy imaging method to investigate the interaction of astrocytes with immune cells at the blood brain barrier of EAE lesions. Here, the focus will be on the developing lesion, i.e. before onset and in the onset of clinical signs to investigate the early involvement of astrocytes in the pathophysiology of these lesions. These experiments will highlight the overall role of astrocytes for lesion evolution, damage, protection and/or repair in the inflamed CNS. These findings might open up new therapeutic targets to stop neurodegeneration and progressing disability in MS patients.