BIOLOGY OF SPATIOTEMPORAL NITRIC OXIDE GRADIENTS

这一建议要研究的中心原则是，一氧化氮(NO)是一种细胞信号分子，细胞毒性的介体不仅取决于给定时间点的瞬时浓度，而且还取决于浓度梯度。目前用于NO研究的方案不允许NO生物相关浓度梯度的空间释放，也不允许以高分辨率直接在时间和空间上检测NO。这项提议的一个重要目标是开发能够时空检测NO的新型荧光探针和供体，并开发新的明确的时空释放NO的方法。为了更好地了解NO梯度在调节细胞信号和细胞毒性中的作用，Cobre IDEA提案中进行的研究将垂直整合五个使用独特但互补的实验系统的医学相关研究项目。这些新的NO试剂将在整个提案中使用，并将专门用于该项目，以测试一氧化氮是精子趋化分子的假设。由于一氧化氮与老年人的肠道功能障碍有关，项目2将使用老年大鼠模型系统来研究肠道神经系统内神经和胃肠NO信号的变化是否参与肠道神经系统内神经和胃肠NO信号的变化。项目3将阐明特定的细胞生理学、病理学和信号通路，涉及上呼吸道上皮中NO的产生，以及它在加重和/或缓解呼吸道病原体引起的疾病中的作用。项目4将利用小鼠逆转录病毒模型研究一氧化氮在逆转录病毒诱导的神经退行性变中的作用。这项研究将为NO在HV诱导的神经疾病中的潜在作用提供洞察力。最后，项目5将研究NO在控制由专性细胞内细菌人类病原体感染中的作用。还将研究细菌对无应激的反应。这些研究不仅将导致具有广泛适用性的新型NO释放和检测试剂的开发，还将使人们更好地理解NO作为信号分子的作用，以及作为人类疾病进展中的保护或破坏因子。

The central tenet to be investigated in this proposal is that nitric oxide (NO) is a cell signaling molecule and mediator of cytotoxicity is not just dependent upon its instantaneous concentration at a given point, but rather it is also dependent upon its concentration gradient. The protocols that are currently in use for NO research do not allow the spatial release of biologically relevant concentration gradients of NO, nor do they allow the direct temporal and spatial detection of NO with fine resolution. An important goal of this proposal is to develop novel fluorescent probes and donors that afford spatiotemporal detection of NO, and to develop new methods for well defined spatiotemporal release of NO. To achieve a greater understanding of the role of NO gradients in mediating cell signaling and cytotoxicity, the research conducted in this COBRE IDeA proposal will vertically integrate five medically relevant research projects that employ unique but complementary experimental systems. These novel NO reagents will be used throughout this proposal and will be specifically used in this project to test the hypothesis that nitric oxide is a sperm chemotactic molecule. Because NO has been implicated in bowel dysfunction in the elderly, Project 2 will employ an aged rat model system to investigate whether alterations in neural and gastrointestinal NO signaling within the enteric nervous system of the bowel investigate whether alterations in neural and gastrointestinal NO signaling within the enteric nervous system of the bowel are involved. Project 3 will elucidate specific cell physiology, pathology, and signaling pathways involving NO production in the upper airway epithelium and its role in exacerbating and/or moderating disease caused by respiratory tract pathogens. Project 4 will investigate the role of NO in retrovirus-induced neurodegeneration using a murine retrovirus model. This investigation will provide insight into the potential role of NO in HV-induced neurological disease. Finally, Project 5 will examine the role of NO in controlling infection by obligate intracellular bacterial human pathogens. The bacterial response to NO stress will also be investigated. Not only will these studies lead to the development of novel NO release and detection reagents with broad scale applicability, they will also provide a greater understanding of the role of NO as a signaling molecule, and as protective or damaging factor in the progression of human disease.