Immunoregulation of herpes encephalitis by microglia

描述（由申请方提供）：小胶质细胞和其他脑巨噬细胞是CNS内神经炎症的主要调节因子。单纯疱疹病毒（HSV）-1是HIV-1感染患者中的重要机会致病菌，也是正常宿主中破坏性CNS感染的原因。众所周知，neuroAIDS期间观察到的脑损伤不是由神经元的直接HIV感染引起的，而是由巨噬细胞产生的神经炎症介质引起的。小胶质细胞产生这些相同的神经炎症介质响应于HSV，并且该提议利用HSV感染小鼠作为小动物模型来研究病毒脑感染期间活化的小胶质细胞的作用。待检验的中心假设是，病毒性脑炎期间发生的氧化应激可以被调节以防止免疫介导的组织损伤机制。为了验证这一假设，我们将（1）确定TLR 2是否介导疱疹性脑炎期间发生的氧化应激。这将通过使用从野生型与TLR 2敲除小鼠分离的培养的小胶质细胞比较病毒诱导的前氧化酶和反应性物质的产生来实现。其他实验将比较这些基因敲除动物感染病毒性脑炎期间发生的氧化应激诱导的脑损伤。然后我们将（2）确定抗氧化酶是否调节病毒性脑炎期间的氧化应激。这些研究将检查选择的抗氧化酶对HSV反应性物质和小胶质细胞凋亡的影响。我们将继续研究这些抗氧化酶在体内病毒性脑炎中控制氧化应激诱导的脑损伤的作用。最后，我们将（3）确定病毒诱导的氧化性脑损伤是否可以控制。这将通过在iNOS-荧光素酶转基因小鼠的脑中过表达抗炎细胞因子并使用体内实时生物发光成像评估表达来实现。在最后的研究中，我们将调节疱疹性脑炎期间发生的氧化应激反应。这将通过构建过表达破坏TLR信号传导的牛痘病毒蛋白以及抗氧化酶的重组疱疹病毒，并评估感染动物中产生的组织损伤来实现。通过本提案中描述的研究，我们希望确定治疗疱疹性脑炎和神经艾滋病以及其他形式的病毒性脑炎的新方法，这些方法可以减少免疫介导的脑损伤的程度。公共卫生相关性已经确定，在neuroAIDS期间观察到的脑损伤不是由神经元的直接HIV感染引起的，而是由小胶质细胞产生的神经炎性介质（例如活性氧和活性氮物质）诱导的。通过使用小胶质细胞活化和抗氧化酶的抑制剂，有可能调节病毒性脑炎期间发生的氧化应激反应。在这些实验中，我们希望确定治疗疱疹性脑炎和神经艾滋病以及其他形式的病毒性脑炎的新方法，这些方法可以降低免疫介导的脑损伤机制的程度。

DESCRIPTION (provided by applicant): Microglia and other brain macrophages are the principal regulators of neuroinflammation within the CNS. Herpes simplex virus (HSV)-1 is an important opportunistic pathogen in HIV-1-infected patients as well as the cause of a devastating CNS infection in normal hosts. It is well established that the brain damage seen during neuroAIDS is not caused by direct HIV infection of neurons, but rather is induced by macrophage-produced neuroinflammatory mediators. Microglial cells produce these same neuroinflammatory mediators in response to HSV, and this proposal utilizes HSV infection of mice as a small animal model to investigate the role of activated microglia during viral brain infection. The central hypothesis to be tested is that oxidative stress occurring during viral encephalitis can be modulated to prevent immune-mediated mechanisms of tissue damage. To test this hypothesis, we will (1) determine whether TLR2 mediates oxidative stress occurring during herpes encephalitis. This will be achieved by comparing virus-induced production of pro-oxidative enzymes and reactive species using cultured microglia isolated from wild-type versus TLR2 knockout mice. Additional experiments will compare oxidative stress-induced brain damage occurring during viral encephalitis following infection of these knockout animals. We will then (2) determine whether antioxidant enzymes regulate oxidative stress during viral encephalitis. These studies will examine the effect of select antioxidant enzymes on the production of reactive species and microglial cell apoptosis in response to HSV. We will go on to examine the role of these antioxidant enzymes in controlling oxidative stress-induced brain damage during viral encephalitis in vivo. Finally, we will (3) determine if virus-induced oxidative brain damage can be controlled. This will be achieved through overexpressing anti-inflammatory cytokines in the brains of iNOS-luciferase transgenic mice and assessing expression using in vivo real-time bioluminescence imaging. In the final studies, we will modulate the oxidative stress response occurring during herpes encephalitis. This will be achieved by constructing recombinant herpesviruses overexpressing vaccinia virus proteins that disrupt TLR signaling, as well as antioxidant enzymes, and assessing the resulting tissue damage in infected animals. Through studies described in this proposal, we hope to identify new approaches for treatment of herpes encephalitis and neuroAIDS, as well as other forms of viral encephalitis, which reduce the extent of immune-mediated brain damage. PUBLIC HEALTH RELEVANCE It is well established that brain damage seen during neuroAIDS is not caused by direct HIV infection of neurons, but rather is induced by microglial cell- produced neuroinflammatory mediators, such as reactive oxygen and reactive nitrogen species. Through the use of inhibitors of microglial cell activation and antioxidant enzymes, it may be possible to modulate the oxidative stress response occurring during viral encephalitis. In these experiments, we hope to identify new approaches to the treatment of herpes encephalitis and neuroAIDS, as well as other forms of viral encephalitis, which reduce the extent of immune- mediated mechanisms of brain damage.