Inhibition and Inactivation of NO Synthase by Tobacco

烟草对 NO 合酶的抑制和灭活

• 批准号: 8033226
• 负责人: YOICHI OSAWA
• 金额: $ 28.61万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-05-01 至 2012-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8033226
• 关键词: Active Sites; Address; Arginine; Bioavailable; Biological; Biological Assay; Biological Markers; Biological Models; Blood; CYP2E1 gene; Cell model; Charcoal; Chemicals; Cigarette; Clinical; Complex Mixtures; Crude Extracts; Cytochrome P450; Disease; Enzymes; Event; Exposure to; Family; Functional disorder; Goals; Hand; Heating; Heme; Individual; Lead; Liver; Mediating; Metabolic; Metabolism; Methods; Molecular Weight; Mus; Nature; Neurons; Nicotine; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Oral Administration; Oral Tobacco; Physiological; Play; Process; Production; Prosthesis; Proteins; Pterins; Reaction; Regulation; Research; Research Personnel; Risk Factors; Role; Smoke; Smoker; Smoking; Stress; Techniques; Technology; Testing; Time; Tobacco; Tobacco smoke; Tobacco smoking; Tobacco use; Water; base; biological systems; cigarette smoking; cigarette smoking; clinical effect; drug of abuse; human NOS3 protein; inhibitor/antagonist; metabolomics; novel; oxidation; programs; tetrahydrobiopterin; therapy development; tool

DESCRIPTION (provided by applicant): The objective of the proposed research is to characterize the tobacco-mediated inhibition and inactivation of NO-synthase (NOS), a cytochrome P450-like enzyme. In the course of these studies, we aim to isolate and identify the chemicals in tobacco that are responsible for these effects. This will be addressed, in part, by a differential metabolic profiling approach, as well as the use of high-throughput bioassays and state of the art LC-MS/MS techniques. Exposure to chemicals in tobacco is known to cause a dysfunction in NO production and availability. Cigarette smoking is associated with a variety of diseases and, since NO plays a central role in a myriad of physiological functions, it is likely that such interactions may be clinically important. We have established, with the use of crude extracts derived from tobacco and tobacco smoke, that there is an immediate, reversible inhibition of NOS, as well as a metabolism-based, time-dependent, irreversible inactivation of NOS. Further studies on the extracts indicate that small, organic, water-soluble compounds are responsible for causing NOS dysfunction. Moreover, these extracts specifically inactivated endothelial NOS (eNOS) when L-arginine was present, whereas specific inactivation of neuronal NOS (nNOS) was observed when tetrahydrobiopterin was in excess. The following specific aims are proposed: 1. To characterize the inactivation of nNOS caused by tobacco components. This includes identification of the chemicals responsible for inactivation. 2. To characterize the inactivation of eNOS caused by tobacco components. The mechanism of tetrahydrobiopterin oxidation, as well as the identity of the chemicals responsible for inactivation will be addressed. 3. To characterize the mechanism of inhibition of eNOS and nNOS caused by tobacco smoke, as well as determine the identity of the components responsible. The successful completion of these aims will identify potentially novel inhibitors and inactivators of NOS as pharmacological tools, as well as allow for a more detailed study of the effects of these chemicals in smokers. Since over 40 million people are exposed to these chemicals on a daily basis, information on the nature of these chemicals and their interaction with important biological systems would be useful. These studies should also lead to identification of biomarkers for tobacco-related diseases involving NOS and facilitate development of therapies for NOS dysfunction.

描述（由申请人提供）：拟议研究的目的是表征烟草介导的一氧化氮合酶（NOS）（一种细胞色素 P450 样酶）的抑制和失活。在这些研究过程中，我们的目标是分离和鉴定烟草中造成这些影响的化学物质。这将通过差异代谢分析方法以及使用高通量生物测定和最先进的 LC-MS/MS 技术来部分解决。众所周知，接触烟草中的化学物质会导致一氧化氮的产生和利用功能障碍。吸烟与多种疾病有关，并且由于一氧化氮在多种生理功能中发挥着核心作用，因此这种相互作用可能具有重要的临床意义。通过使用从烟草和烟草烟雾中提取的粗提物，我们已经确定，NOS 具有立即、可逆的抑制作用，以及基于代谢的、时间依赖性、不可逆的 NOS 失活作用。对提取物的进一步研究表明，小的有机水溶性化合物是导致 NOS 功能障碍的原因。此外，当存在L-精氨酸时，这些提取物特异性地灭活内皮NOS（eNOS），而当四氢生物蝶呤过量时，观察到神经元NOS（nNOS）的特异性灭活。提出以下具体目标： 1. 表征烟草成分引起的 nNOS 失活。这包括鉴定导致失活的化学物质。 2. 表征烟草成分引起的 eNOS 失活。将讨论四氢生物蝶呤氧化的机制以及负责失活的化学物质的身份。 3. 表征烟草烟雾抑制 eNOS 和 nNOS 的机制，并确定其作用成分。这些目标的成功完成将确定潜在的新型 NOS 抑制剂和灭活剂作为药理学工具，并允许更详细地研究这些化学物质对吸烟者的影响。由于每天有超过 4000 万人接触这些化学物质，因此有关这些化学物质的性质及其与重要生物系统相互作用的信息将非常有用。这些研究还应有助于识别涉及 NOS 的烟草相关疾病的生物标志物，并促进 NOS 功能障碍疗法的开发。