E-Prostanoid Receptor Subtype 2 (EP2) Regulation of Microglial Activation

描述（由申请人提供）：小胶质细胞先天免疫反应的激活与几种神经退行性疾病的发生或进展有关。也许在阿尔茨海默病（AD）模型中研究得最好，先天免疫激活已被证明对神经元既有有益的影响，也有有害的影响。有益作用与清除Abeta肽的神经毒性物质有关，而对神经元的旁分泌损伤则是由包括活性氧和活性氮在内的多种有毒物质引起的。许多学术和制药实验室的主要目标是确定在抑制旁分泌神经毒性的同时增强或维持先天免疫激活的有益作用的方法。最近，我们证明了原代小鼠小胶质细胞EP2受体的基因消融导致了Abeta物种吞噬增加和旁分泌神经毒性完全阻断的高度理想的双重表型。其他人用不同的方法验证了我们的发现。综上所述，这些数据有力地支持了EP2受体在阿尔茨海默病和其他神经退行性疾病中作为一个非常有希望的靶点来操纵小胶质先天免疫反应。ep2介导的信号传导与β吞噬和旁分泌神经毒性相关的机制尚不清楚。我们的小鼠初级小胶质细胞表达的初步数据确定了一个与EP2紧密相关的候选基因。据我们所知，没有关于该基因在任何脑细胞（包括小胶质细胞）中的作用的报道。然而，鉴于新出现的数据，我们假设它的表达和活性与EP2信号传导有机制联系，并且它可能是EP2消融产生增强Abeta吞噬和减少旁分泌神经毒性的高度理想的双重表型的关键因素。我们将通过以下具体目的来验证这一假设：1)在体外验证我们的候选mRNA和蛋白在Abeta治疗前后在野生型和EP2-/-原代小鼠小胶质细胞中的表达。2)绘制野生型小鼠和转基因AD小鼠骨髓移植前后的体内区域和细胞脑分布图。3)在敲除小鼠和小胶质细胞系中，通过shRNA敲除确定其在EP2-/-双表型背景下的表达功能。

DESCRIPTION (provided by applicant): Activation of innate immune response in microglia is associated with the initiation or progression of several neurodegenerative diseases. Perhaps best studied in models of Alzheimer's disease (AD), innate immune activation has been shown to have both beneficial and deleterious effects on neurons. Beneficial effects are related to the clearance of nerurotoxic species of Abeta peptides, while paracrine damage to neurons results from the elaboration of a variety of toxic substances including reactive oxygen and nitrogen species. A major goal of many academic and pharmaceutical laboratories is to identify means to augment or maintain the beneficial actions of innate immune activation while suppressing paracrine neurotoxicity. Recently we demonstrated that genetic ablation of the EP2 receptor from primary mouse microglia resulted in the highly desirable dual phenotype of an increase in phagocytosis of Abeta species and complete blockade of paracrine neurotoxicity. Others have validated our findings using different methods. Taken together, these data strongly support EP2 receptor as a highly promising target for manipulating microglial innate immune response in AD and perhaps other neurodegenerative diseases. The mechanisms by which EP2-mediated signaling is related to Abeta phagocytosis and paracrine neurotoxicity are not known. Our Preliminary Data of mouse primary microglia expression identified a candidate gene that is tightly associated to EP2. We are aware of no report on the actions of this gene in any brain cells, including microglia. However, given emerging data we hypothesize that its expression and activity are mechanistically linked to EP2 signaling and that it may be a key element by which ablation of EP2 generated the highly desirable dual phenotype of enhanced Abeta phagocytosis and reduced paracrine neurotoxicity. We will test this hypothesis through the following Specific Aims: 1) Perform in vitro validation of our candidate mRNA and protein expression in wild type and EP2-/- primary mouse microglia before and after Abeta treatment. 2) Map the in vivo regional and cellular brain distribution in wild type mice and a transgenic mouse model of AD before and after following bone-marrow transplantation from either wild type or EP2-/- mice. 3) Determine the functionality of its expression in the context of the EP2-/- dual phenotype in both knockout mice and a microglial cell line using shRNA knockdown.