| Oxidants have long been recognized as critical components of our host defense system. These highly reactive toxins, however, may inflict host injury if not properly controlled. Tissue and plasma antioxidants are present to prevent such injury. When these defenses are overcome, biological oxidation may contribute to the pathogenesis of a multitude of disorders.;Neutrophils are a critical component of our innate host defenses. They generate a battery of oxidants to destroy invading pathogens. Myeloperoxidase (MPO) is the most abundant protein of neutrophils, catalyzing the production of halogenating and nitrating intermediates from hydrogen peroxide, chloride (Cl-), bromide (Br-), and nitrite (NO2-), the nitric oxide (NO·) autoxidation product.;While there is extensive evidence demonstrating involvement of oxidants in host defense and disease, there is little information regarding their physiologic source. Furthermore, the role of antoxidants in preventing oxidant-mediated host injury is poorly understood. The studies described in this thesis explore the role of MPO and vitamin C in this complex scenario of oxidants, antioxidants, host defense, and host injury.;We developed gas chromatography/mass spectrometry (GC/MS) methods to quantify 3-chlorotyrosine, 3-bromotyrosine, and 3-nitrotyrosine, markers of chlorination, bromination, and nitration, respectively, from biological samples. Using this sensitive and specific methodology, we investigated the physiologic role of MPO in producing reactive chlorinating, brominating, and nitrating intermediates in vivo. We subjected wild-type and MPO-deficient mice to two models of sepsis: cecal ligation and puncture and intraperitoneal injection of Klebsiella pneumoniae. Our results demonstrate that MPO is a physiologically relevant source of halogenating and nitrating intermediates in vivo. MPO-deficient mice exhibit higher mortality when subjected to both models of infection. Br- and NO2- significantly alter the bactericidal properties of MPO. Thus, MPO-derived halogenating and nitrating intermediates are likely to exert potent antimicrobial action during sepsis.;The role of vitamin C, a proposed antioxidant, in host defense and host injury was also explored. Mice were generated that had non-functional L-gulono-gamma-lactone oxidase (gulo-/-), a key vitamin C synthetic enzyme. Gulo-/- mice that were not supplemented with vitamin C exhibited a striking survival defect when injected with K. pneumoniae . Surprisingly, GC/MS analysis revealed no differences in levels of several different oxidation products. These observations suggest a previously unsuspected role for vitamin C in neutrophil function and raise questions about its proposed role as an antioxidant. |
