NITRITE/COLLAGEN REACTIONS IN AGING AND SMOKING

衰老和吸烟中的亚硝酸盐/胶原蛋白反应

• 批准号: 6044277
• 负责人: DAVID C PAIK
• 金额: $ 10.33万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-03-01 至 2004-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6044277
• 关键词: aging; ascorbate; biomarker; chemical reaction; collagen; crosslink; cytotoxicity; environmental toxicology; free radical scavengers; high performance liquid chromatography; human tissue; nitric oxide; nitrites; postmortem; protein protein interaction; protein structure; retinoids; smoking; statistics /biometry; tobacco abuse; tocopherols; tyrosine analog

Although nitrite reactions have been studied in the past, the specific reaction with collagen and its role in aging has never been examined. We have recently observed that in vitro nitrite-treated collagen shows similarities with aged human collagen. Shared properties include the formation of a unique tyrosine derivative we call compound N, a decease in unmodified tyrosine residues, the accumulation of a yellowish chromophore absorbing at 350 nm, and an increase in collagen cross-linking. Increased cross-linking, evidenced by a decreased susceptibility to enzymatic digestion, results in the stiffening of collagen and may account for the stiffening of human blood vessels, lung, and skin which occurs with aging. The majority of human exposure to nitrite results from endogenous nitric oxide production and exogenous sources which include cigarette smoking and cured meat ingestion. In order to establish the in vivo significance of the nitrite/collagen reaction to age-related changes in human collagen we will develop methods to detect biomarkers of the reaction (especially tyrosine derivatives such as compound N, 3-nitrotyrosine, dityrosine, diazotyrosine and deaminated diazotyrosine). Kinetics on the reactions between nitrite and potential reaction intermediates will be conduced in order to more precisely define the mechanism of reaction involved. The biomarkers and reaction intermediates will be identified by reverse phase HPLC amino acid analysis with photodiode array, fluorescence, and electrochemical in-line detection systems. The methods for biomarker detection will be used to compare collagen extracted from the skin, aorta, and lung of young and old individuals. Degree of cross-linking and tyrosine content will also be determined for each collagen sample. Statistical analyses will include multiple logistic regressions with age and smoking as independent variables. Finally, we will examine the effectiveness of free radical scavengers such as vitamins A, C and E to modulate these reactions in vitro. Because cigarette smoking is a major source of an individual's exogenous nitrite exposure, the nitrite/collagen reaction may provide important clues to the pathogenesis of disease processes associated with both aging and cigarette smoking. These include a variety of atherosclerotic vascular diseases, chronic obstructive pulmonary disease, premature sign wrinkling, and age-related lens cataracts.

尽管过去已经研究了亚硝酸盐反应，但是从未研究过与胶原蛋白的特定反应及其在衰老中的作用。 我们最近观察到，经过体外亚硝酸盐处理的胶原蛋白显示出与老化的人类胶原蛋白的相似性。 共享特性包括我们称之为化合物N的独特酪氨酸衍生物的形成，未修饰的酪氨酸残基的脱发，在350 nm处吸收的淡黄色色团的积累以及胶原蛋白交叉链接的增加。交联的增加，证明了酶消化的敏感性降低，导致胶原蛋白的僵硬，可能是随着衰老而发生的人体血管，肺和皮肤的僵硬。 大多数人暴露于亚硝酸盐是由内源性一氧化氮的产生和外源性来源引起的，包括吸烟和腌制肉类。 为了确定亚硝酸盐/胶原蛋白反应对人类胶原蛋白与年龄相关的变化的体内意义，我们将开发方法来检测反应的生物标志物（尤其是酪氨酸衍生物，例如复合N，3-硝基酪氨酸，二甲苯蛋白，dintilrosine，dintyrosine，diinzotyrosine，nizzotyrosine and Diquyarosine and diacamine diaczotyrosine）。 为了更精确地定义涉及的反应机理，将抑制亚硝酸盐和潜在反应中间体之间反应的动力学。 生物标志物和反应中间体将通过使用光电二极管阵列，荧光和电化学内检测系统的反相HPLC氨基酸分析来鉴定。 生物标志物检测的方法将用于比较年轻人和老年人从皮肤，主动脉和肺中提取的胶原蛋白。 每个胶原蛋白样品还将确定交联的程度和酪氨酸含量。 统计分析将包括多种逻辑回归，年龄和吸烟作为自变量。 最后，我们将研究自由基清除剂（例如维生素A，C和E）在体外调节这些反应的有效性。由于吸烟是个人外源亚硝酸盐暴露的主要来源，因此亚硝酸盐/胶原蛋白反应可能为与衰老和吸烟相关的疾病过程的发病机理提供重要线索。 其中包括各种动脉粥样硬化血管疾病，慢性阻塞性肺部疾病，过早的症状皱纹和与年龄相关的透镜白内障。