OXIDATIVE NEUTROPHIL MICROBICIDAL MECHANISMS

长期目标是表征氧化型杀微生物剂 人中性粒细胞(PMN)内的机制 为感染患者制定合理的治疗干预措施 中性粒细胞数量或功能存在缺陷。因为伟大的 PMN杀微生物剂系统的数量和复杂性，建议 研究将集中在单个PMN的杀菌性能- 衍生酶髓过氧化物酶(MPO)。具体目标是 识别MPO在靶生物中引起的早期生化变化 并评估这些变化的新陈代谢后果。这个 目标是确定哪些生化变化相关性最好，以及 可能是复制能力丧失的原因 (生存能力)。初步研究表明，E. 与生存能力丧失密切相关的大肠杆菌呼吸作用 (r=0.97)，表明对微生物的损害与 呼吸链和杀微生物活性。按下列顺序排列 呼吸电子传递，从营养素到 脱氢物质，对苯二酚和末端氧化酶细胞色素至02， MPO介导的主要损伤似乎位于 脱氢酶基团和链的其他成分留下来 基本完好无损。正在进行的调查将集中在 对单个脱氢酶组分的损伤：铁-硫中心 和黄素辅助因子。电子受损的代谢后果 跨膜pH和电化学损失等转运 梯度(质子动力)，主动运输的损失，以及 将确定广泛的三磷酸腺苷水解。最后，尝试 将通过以下方式将有机体从MPO介导的毒性中拯救出来 提供可绕过以下产品需求的增长条件 MPO氧化体系。成功的营救将意味着损害 对被绕过的系统造成的损失 在正常生长条件下观察到的生存能力。更远的目标 是利用MPO介导的生化变化的知识来 将MPO与完整的其他抗菌活性区分开来 PMN。将PMN的杀微生物活性与MPO特异性进行比较 将寻求微生物结构和相关性的变化。 PMN介导的改变的MPO特异性将得到确认 使用MPO(遗传性MPO缺乏症)或H202缺乏的中性粒细胞 生产(慢性肉芽肿性疾病)。这些研究应该 确定PMN修饰导致的微生物结构 致命的伤害。他们还可以为新的设计提供线索 抗菌剂，并提出为什么某些微生物 能够躲避PMN介导的宿主防御系统。

Long-term objectives are to characterize oxidative microbicidal mechanisms within human neutrophils (PMN) with a view towards developing rational therapeutic interventions for infected patients with defects in PMN number or function. Because of the great number and complexity of PMN microbicidal systems, the proposed studies will focus on microbicidal properties of a single PMN- derived enzyme, myeloperoxidase (MPO). Specific aims are to identify early MPO-induced biochemical changes in target organisms and to evaluate the metabolic consequences of these changes. The goal is to determine which biochemical changes correlate best, and are perhaps responsible for, loss of replicative ability (viability). Preliminary studies have indicated a decrease in E. coli respiration that correlates well with loss of viability (r=0.97), suggesting a relationship between damage to the microbial respiratory chain and microbicidal activity. In the sequence of respiratory electron transport, from nutrients through dehydrogenates, quinones, and terminal oxidase cytochromes to 02, the principal MPO-mediated lesions appear to be in the dehydrogenase group with other components of the chain left substantially intact. Continuing investigations will focus on damage to individual dehydrogenase components: iron-sulfur centers and flavin cofactors. Metabolic consequences of impaired electron transport such as loss of transmembrane pH and electrochemical gradients (proton motive force), loss of active transport, and extensive ATP hydrolysis will be determined. Finally, attempts will be made to rescue organisms from MPO-mediated toxicity by providing growth conditions that bypass the need for products of MPO-oxidized systems. Successful rescue would suggest that damage to the bypassed systems was indeed responsible for the loss of viability observed under usual growth conditions. A further aim is to use the knowledge of MPO-mediated biochemical changes to distinguish MPO from other antimicrobial activity within intact PMNs. PMN microbicidal activity will be compared with MPO-specific changes in microbial structures and correlations will be sought. Specificity for MPO of the PMN-mediated changes will be confirmed using PMNs deficient in MPO (hereditary MPO-deficiency) or in H202 production (chronic granulomatous disease). These studies should identify microbial structures whose modification by PMN results in lethal injury. They may also offer clues for the design of new antimicrobial agents and suggest reasons why certain organisms are able to evade PMN-mediated host defense systems.