Calcineurin and inflammatory signaling processes in aging and Alzheimer's Disease

描述(申请人提供)：阿尔茨海默病(AD)是一种毁灭性的不可逆转的神经疾病，逐渐改变个性和行为，并破坏认知功能。阿尔茨海默病每年的费用超过1000亿美元，也是美国最昂贵的疾病之一。近年来，抗炎药已成为延缓AD症状发生的有希望的化合物。然而，我们对大脑中神经免疫/炎症(im/inf)信号的潜在机制的了解仍然不完整。钙调素依赖的蛋白磷酸酶钙调神经磷酸酶在淋巴细胞和其他细胞类型的im/inf信号级联中起关键作用，但在神经组织中类似的作用很少被考虑。在过去的一年里，我们发现在神经元-神经胶质细胞共培养中钙调神经磷酸酶的激活足以概括与衰老和AD一致的im/inf反应的几个组成部分，包括星形胶质细胞的激活和诱导许多im/inf相关基因的级联。我们还观察到，在AD模型小鼠淀粉样蛋白沉积周围的激活星形胶质细胞中，钙调神经磷酸酶显著和选择性地上调。因此，该项目的长期目标是验证钙调神经磷酸酶是激活与AD相关的im/inf信号过程的关键成分的假说。前两个目标将测试是否激活钙调神经磷酸酶和/或钙调神经磷酸酶依赖的转录因子，NFAT是星形胶质细胞im/inf信号转导所必需的。在存在或不存在有效的钙调神经磷酸酶和NFAT抑制剂的情况下，生物相关的炎症介质将被输送到星形胶质细胞特异性培养物中。Im/inf标记物的诱导将通过Western blots、RT-PCR和细胞因子阵列进行评估。还将研究星形胶质细胞中钙调神经磷酸酶激活的潜在钙来源，以及钙调神经磷酸酶通过上调这些钙来源放大自身活性的可能性。目的利用细胞分离、免疫沉淀、Western印迹、磷酸酶活性测定、免疫组织化学和原位杂交等方法，定性和定量地评价AD脑标本和/或AD啮齿动物模型中钙调神经磷酸酶信号的变化。这些研究的完成将大大增加我们对钙调神经磷酸酶在神经炎症和阿尔茨海默病中的作用的了解，并可能导致开发针对这种疾病和其他神经炎症突出的退行性疾病的新治疗策略。

DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is a devastating and irreversible neurological disease that progressively alters personality and behavior, and destroys cognitive function. With an annual price tag exceeding 100 billion dollars, AD is also one of the nation's most costly diseases. In recent years, anti-inflammatory agents have emerged as promising compounds for delaying the onset of AD symptoms. Yet, our understanding of the mechanisms underlying neuro-immune/inflammatory (im/inf) signaling in the brain remains incomplete. The Ca2+/calmodulin dependent protein phosphatase, calcineurin, plays a critical role in im/inf signaling cascades in lymphocytes and other cell types, but has received little consideration for a similar role in nervous tissue. In the past year, we found that calcineurin activation in neuron-glia co-cultures was sufficient for recapitulating several components of the im/inf response found consistently with aging and AD, including astrocyte activation and the induction of numerous im/inf-related gene cascades. We also observed a marked and selective upregulation of calcineurin in activated astrocytes surrounding amyloid deposits in AD model mice. The long-term goal of this project is to therefore test the hypothesis that calcineurin is a critical component for the activation of im/inf signaling processes associated with AD. The first two aims will test whether activation of calcineurin and/or the calcineurin-dependent transcription factor, NFAT is necessary for im/inf signaling in astrocytes. Biologically relevant inflammatory mediators will be delivered to astrocyte- specific cultures in the presence or absence of potent calcineurin and NFAT inhibitors. The induction of im/inf markers will be assessed with Western blots, RT-PCR, and cytokine arrays. Potential Ca2+ sources for calcineurin activation in astrocytes will also be examined, along with the possibility that calcineurin amplifies its own activity by up-regulating these Ca2+ sources. Aim three will use cell fractionation, immunoprecipitation, Western blot, phosphatase activity assays, immunohistochemistry, and in situ hybridization to qualitatively and quantitatively assess changes in calcineurin signaling in AD brain specimens and/or in a rodent model of AD. Completion of these studies will greatly increase our knowledge of the role of calcineurin in neuroinflammation and AD, and may lead to the development of novel treatment strategies for this disease and other degenerative conditions where neuroinflammation is prominent.