Pathways for the Inflammatory Damage of Biomolecules

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DESCRIPTION (provided by applicant): Oxidants generated by activated white blood cells are critical to host defenses against microorganisms. However, overproduction of reactive species can damage host tissue. Indeed, white blood cells represent the cellular hallmark of inflammation, and oxidants have been implicated in tissue injury in inflammatory diseases ranging from atherosclerosis to neurodegenerative disorders to cancer.We have investigated four phagocyte-dependent pathways that oxidatively damage proteins in vitro. The pathways and their characteristic products are: myeloperoxidase and 3-chlorotyrosine; tyrosyl radical and o,o-dityrosine; hydroxyl radical and ortho-tyrosine; and reactive nitrogen species and 3-nitrotyrosine.Using two clinically relevant models of inflammation, we will study genetically engineered mice whose phagocytes are unable to produce specific oxidants. In the proposed research, we will ask three related questions. First, we will identify the pathways that generate chlorotyrosine, dityrosine, ortho-tyrosine, and nitrotyrosine in viva. The experiments will reveal whether a genetic deficiency of any of these oxidant-generating systems inhibits production of any of the chemical markers, thereby determining which pathway generates a particular marker in viva.Second, we will determine whether tissue, plasma, and urinary levels of the oxidized amino acids change in parallel. We will also investigate the absorption, metabolism, and urinary excretion of the oxidized amino acids. These experiments will determine whether plasma and urinary levels of these well-characterized products can be used as noninvasive markers of oxidative stress.Third, we plan to determine whether two proposed antioxidants-vitamin C or vitamin E- inhibit oxidative stress in our models of inflammation. Collectively, the proposed experiments will identify the oxidative pathways that cause phagocytes to damage tissues and will test the hypothesis that levels of oxidized amino acids in urine and plasma indicate levels of oxidative stress in vivo.