OXYGEN AND OXY-RADICALS IN PHAGOCYTIC KILLING

这项研究的目的是考察小、高度 人体杀菌活性中的活性分子 中性粒细胞(PMNL)，我们最重要的 对感染的防御。这是一项旨在 生成有关含氧自由基或类似物的基本信息 PMNL产生强烈的氧化剂。化学探头， 少量引入到乳胶珠上，将被 用于检测吞噬小体中形成的各种氧化剂。 这些探针是：1)亚油醇，因为1，4-二烯部分 对各种引发的自由基自氧化反应敏感 含氧自由基；2)将使用4-苯基丁酸十二酯来探测 羟基自由基；3)乙基6-(3‘-5’-二甲氧基苯氧基) 己酸将被用来探测和量化HOCl的形成； 和4)将使用6-(苯硫基)己酸乙酯来探测 吞噬小体的一般氧化状态。来自 这些化合物的反应将以质量为特征 光谱分析。将综合真实的标准以确认 产品的身份。 初步实验表明，铁是人体必需的营养物质 观察其中一个探针分子亚油酰基的过氧化 酒精。铁的实验给出了一种不同寻常的分布 氧化产物的分布--而不是 对于羟基自由基引发的过程是可以预期的， 但分布更像是如果 单线态氧也参与了氧化过程。 因此，铁的作用对于理解 PMNL吞噬小体中的氧化过程。个人探测器 分子被设计用来评估各种氧化剂的性质和 使我们能够确定参与的氧化剂的性质 以及这种氧化剂是如何形成的。 探针分子将在之后被引入PMNL 掺入到可吞噬的颗粒上。我们一直在使用 包覆在乳胶珠上的探针分子，我们发现 递送系统激活PMNL并使它们在所有情况下 通常与细菌杀灭有关的过程。这个 在缓冲液中使用包覆有探针分子的乳胶粒 只含有葡萄糖会消除许多潜在的 与更复杂的系统相关的模棱两可。

The purpose of this study is to examine the role of small, highly reactive molecules in the bactericidal activity of human polymorphonuclear leukocyte (PMNL), one of our most important defense against infection. This is a study that is designed to generate fundamental information as to how oxy-radicals or similar strong oxidants are generated by the PMNL. Chemical probes, introduced in small quantities coated onto latex beads, will be used to test for various oxidants that are formed in the phagosome. These probes are: 1) linoleyl alcohol because the 1,4-diene moiety is sensitive to free radical autooxidation initiated by a variety of oxy-radicals; 2) dedecyl 4-phenylbutyrate will be used to probe for the hydroxyl radical; 3) ethyl 6-(3'-5'-dimethoxyphenoxyl) hexanoate will be used to probe for and to quantify HOCl formation; and 4) ethyl 6-(phenylthio)hexanoate will be used to probe the general oxidation state of the phagosome. Products from the reaction of these compounds will be characterized by mass spectrometry. Authentic standards will be synthesized to confirm the identity of the products. Preliminary experiments have shown that iron is essential for observing peroxidation of one of the probe molecules, linoleyl alcohol. The experiments with iron gave an unusual distribution of oxidation products -- not the distribution of products that would be expected for a process initiated by the hydroxyl radical, but a distribution more like what would have been predicated if singlet oxygen were also participating in the oxidative process. Therefore, the role of iron is critical to understanding the oxidative processes in the PMNL phagosome. The individual probe molecules are designed to assess various oxidant properties and allow us to determine the nature of the oxidant that participates in bacterial killing and how this oxidant is formed. The probe molecules will be introduced to the PMNL after incorporation onto phagocytosable particles. We have been using probe molecules coated onto latex beads and we find that this delivery system activates the PMNLs and causes them to under go all of the processes normally associated with bacterial killing. The use of latex particles coated with probe molecules in a buffer containing only glucose eliminates many of the potential ambiguities associated with more complex systems.