Astrocyte dysfunction in EAE

描述（由申请人提供）：星形胶质细胞对于维持中枢神经系统（CNS）内的稳态至关重要。这些细胞产生血小板反应蛋白（TSP），大的寡聚体基质细胞蛋白，在细胞附着，细胞迁移，细胞骨架动力学和血管生成中具有重要作用。TSP-1和TSP-2最近也被确定为主要的星形胶质细胞衍生的因子，促进兴奋性突触形成在体外和在体内发育过程中。这种活性已经被映射到它们的表皮生长因子样重复序列，通过结合到神经元a2 d-1电压门控钙通道，也是药物加巴喷丁（GBP）的受体。GBP在体外和体内发育过程中有效抑制TSP介导的突触形成。虽然TSP水平在成年CNS中普遍下降，但我们在正常断奶小鼠的脊髓中发现了可测量的TSP-1。此外，脊髓TSP-1水平迅速下降，然后恢复平行于后肢瘫痪的出现和消失，在复发性实验性自身免疫性脑脊髓炎（EAE），一个建立的啮齿动物模型的人类脱髓鞘疾病，多发性硬化症（MS）。我们和其他人已经表明，可逆的突触病理学可以在复发性EAE小鼠的腰脊髓中发现，我们发现，全身给予GBP的EAE动物在高峰瘫痪开始，当脊髓TSP-1水平处于最低点，刚刚开始恢复，显着延迟临床改善和重建运动突触在腰脊髓灰质。综上所述，这些发现使我们假设星形胶质细胞在成年脊髓中产生TSP-1以维持运动通路中的兴奋性突触，但复发性EAE期间的局部炎症环境导致该介质变得失调，从而导致短暂的突触变化和可逆的后肢瘫痪。如果得到证实，这将使星形胶质细胞成为代表动物CNS脱髓鞘疾病的可逆性神经功能缺损的主要贡献者，并且通过扩展，成为人类治疗干预的新靶点。在此，提出了以下目标：1）为了证实星形胶质细胞衍生的TSP- 1/TSP-2信号通过神经元a2 d-1电压门控钙通道在通过作用于脊髓运动通路中的兴奋性突触介导复发性EAE的可逆性麻痹中的重要性，和2）为了表征在体外和在体内EAE期间引起星形胶质细胞下调TSP-1表达的机制，并确定当EAE缓解发生时脊髓TSP-1表达恢复和临床恢复的因素。这些研究将阐明在已建立的CNS脱髓鞘疾病动物模型中可逆性神经功能缺损的新细胞和分子机制。由于临床复发导致人类MS的慢性疾病进展，因此必须更好地了解这些可逆事件并开发阻断它们的新方法。目前的MS疗法靶向炎症反应，但还没有直接干预神经元或神经胶质细胞水平。 公共卫生相关性：这些研究将阐明可逆性瘫痪发展的新机制，这种可逆性瘫痪发生在人类脱髓鞘疾病复发缓解型多发性硬化症（MS）的成熟动物模型中。由于暂时的临床复发导致人类MS的慢性不可逆疾病进展，因此必须更好地了解这些事件并开发阻断它们的新治疗方法。目前的MS疗法靶向免疫反应，但迄今为止还没有一种疗法能像我们打算做的那样直接干预神经细胞或它们的连接。

DESCRIPTION (provided by applicant): Astrocytes are essential for the maintenance of homeostasis within the central nervous system (CNS). These cells produce thrombospondins (TSPs), large oligomeric matricellular proteins with important roles in cell attachment, cell migration, cytoskeletal dynamics and angiogenesis. TSP-1 and TSP-2 have also recently been identified as the main astrocyte-derived factors that promote excitatory synapse formation in vitro and during development in vivo. This activity has been mapped to their epidermal growth factor-like repeats, via binding to the neuronal a2d-1 voltage-gated calcium channel that is also a receptor for the drug, gabapentin (GBP). GBP potently inhibits TSP-mediated synapse formation in vitro and during development in vivo. While TSP levels generally decline in the adult CNS, we find measurable amounts of TSP-1 in the spinal cords of normal weanling mice. Furthermore, spinal cord TSP-1 levels fall rapidly and then recover in parallel to the appearance and disappearance of hind limb paralysis during relapsing experimental autoimmune encephalomyelitis (EAE), an established rodent model of the human demyelinating disease, multiple sclerosis (MS). We and others have shown that reversible synaptic pathology can be found in the lumbar spinal cords of mice with relapsing EAE, and we find that systemic administration of GBP to animals with EAE starting at peak paralysis, when spinal cord TSP-1 levels are at their nadir and just beginning to recover, significantly delays both clinical improvement and the reestablishment of motor synapses in lumbar spinal grey matter. Taken together, these findings lead us to hypothesize that astrocytes produce TSP-1 in the adult spinal cord to maintain excitatory synapses in the motor pathway, but the local inflammatory milieu during relapsing EAE causes this mediator to become dysregulated resulting in transient synaptic changes and reversible hind limb paralysis. If confirmed, this would make astrocytes central contributors to the reversible neurological deficits that typify animal CNS demyelinating diseases, and by extension, novel targets for therapeutic intervention in humans. Here, the following aims are proposed: 1) To confirm the importance of astrocyte-derived TSP- 1/TSP-2 signaling through neuronal a2d-1 voltage-gated calcium channels in mediating the reversible paralysis of relapsing EAE by acting on excitatory synapses in motor pathways of the spinal cord, and 2) To characterize the mechanisms that cause astrocytes to down-regulate TSP-1 expression in vitro and in the spinal cord during EAE in vivo, and determine the factors responsible for the return of spinal cord TSP-1 expression and clinical recovery as EAE remission occurs. These studies will clarify novel cellular and molecular mechanisms underlying reversible neurological deficits in an established animal model of CNS demyelinating disease. Since clinical relapses beget chronic disease progression in human MS, it is essential to better understand these reversible events and to develop novel approaches that block them. Current MS therapies target the inflammatory response, but none as of yet directly intervene at either the neuronal or the glial cell level. PUBLIC HEALTH RELEVANCE: These studies will clarify novel mechanisms underlying the development of reversible paralysis that occurs in a well-established animal model of the human demyelinating disease, relapsing-remitting multiple sclerosis (MS). Since temporary clinical relapses lead to chronic irreversible disease progression in human MS, it is essential to better understand these events and to develop novel treatment approaches that block them. Current MS therapies target the immune response, but none as of yet can directly intervene at the level of nerve cells or their connections as we intend to do.