Effects of Neuroinflammation on Gap Junction Communication in Glia

描述（由申请人提供）：间隙连接代表接触细胞之间的直接细胞间通道，允许小分子（bbb10 1kda）通过，包括离子、代谢前体和第二信使。在中枢神经系统（CNS）中观察到广泛的细胞间偶联和大量的间隙连接，提示神经胶质室的合胞样组织。一种对神经胶质间隙连接通讯（GJC）的影响尚不清楚的中枢神经系统感染性疾病是由化脓性细菌引起的实质感染，导致脑脓肿的形成。最近的研究表明，在发展中的脑脓肿中检测到的几种促炎介质，由金黄色葡萄球菌激活的胶质细胞产生，包括白细胞介素-1 （IL-1）、肿瘤坏死因子- α (TNF-a)和一氧化氮（NO），能够调节星形胶质细胞和小胶质细胞的GJC。具体来说，这些分子减弱了星形胶质细胞中的GJC，而激活的小胶质细胞则成为功能偶联的。我们将这种现象称为“合胞开关”，并提出在脑脓肿过程中形成的炎症环境可能对重塑常驻胶质细胞之间的相互作用类型很重要，并且偏离生理偶联可能影响远离感染主要焦点的大脑区域的完整性。这些变化可能是由脓肿内Cx表达的区域差异决定的。本研究的目的是研究IL-1、TNF-a和NO在调节神经胶质合胞开关中的功能重要性，以及Cx43在脑脓肿发病中的作用。为了实现这一目标，以下具体目标将被解决：(1)评估金黄色葡萄球菌及其细胞壁产物PGN对初级星形胶质细胞和小胶质细胞的同质细胞GJC的影响以及负责合胞开关的信号通路；(2)研究促炎介质IL-1、TNF-a和NO在金黄色葡萄球菌刺激下对神经胶质细胞GJC的调节作用；(3)利用遗传KO模型研究促炎介质对金黄色葡萄球菌诱导的实验性脑脓肿小鼠模型中连接蛋白表达的影响以及Cx43在疾病发病机制中的功能重要性。由于正常中枢神经系统中神经胶质细胞群的广泛间隙连接偶联，正常神经胶质合胞网络的神经炎症破坏可能在一定程度上导致脑脓肿消退后患者观察到的一些长期影响，包括癫痫发作和认知缺陷。这些实验将为了解促炎介质如何影响脑脓肿胶质细胞GJC的程度提供有意义的见解。

DESCRIPTION (provided by applicant): Gap junctions represent direct intercellular conduits between contacting cells that permit the passage of small molecules (> 1 kDa) including ions, metabolic precursors, and second messengers. The observation of extensive intercellular coupling and large numbers of gap junctions in the central nervous system (CNS) suggests a syncytium-like organization of glial compartments. One CNS infectious disease in which nothing is known regarding its impact on glial gap junction communication (GJC) is parenchymal infection with pyogenic bacteria leading to the establishment of brain abscess. Recent studies have revealed that several proinflammatory mediators detected in developing brain abscesses and produced by S. aureus activated glia, including interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF-a), and nitric oxide (NO) are capable of modulating GJC in astrocytes and microglia. Specifically, these molecules attenuate GJC in astrocytes whereas activated microglia become functionally coupled. We have coined this phenomenon a "syncytial switch" and propose that the inflammatory milieu that develops during the course of brain abscess may be important for remodeling the types of interactions between resident glia and that deviation from physiological coupling may impact the integrity of brain regions distant from the primary focus of infection. These changes may be dictated by regional variations in Cx expression within the abscess. The objective of the proposed work is to investigate the functional importance of IL-1, TNF-a, and NO in regulating the glial syncytial switch and the role of Cx43 in brain abscess pathogenesis. To address this objective the following Specific Aims will be addressed: (1), to evaluate the consequences of S. aureus and its cell wall product PGN on homocellular GJC in primary astrocytes and microglia and the signaling pathways responsible for the syncytial switch; (2), to establish the functional importance of the proinflammatory mediators IL-1, TNF-a, and NO on modulating glial GJC in response to S. aureus stimulation using primary glia from knockout (KO) mice; and (3), to investigate the role of proinflammatory mediators on connexin expression and the functional importance of Cx43 in disease pathogenesis in a mouse model of S. aureus-induced experimental brain abscess using genetic KO models. Due to the extensive gap junctional coupling of glial cell populations in the normal CNS, neuroinflammatory disruption of normal glial syncytial networks could contribute, in part, to some of the long-term effects observed in patients following brain abscess resolution including seizures and cognitive deficits. These experiments will provide meaningful insights into how proinflammatory mediators influence the extent of glial GJC in brain abscess.