The observation that
antioxidant vitamins fail to confer protective benefits in large, well-designed randomized clinical trials has led many to question the role of oxidative stress in the pathogenesis of disease. However, there is little evidence that proposed
antioxidants actually scavenge reactive intermediates in vivo. Ascorbate reacts rapidly with
oxidants produced by activated neutrophils in vitro, and neutrophils markedly increase their
oxidant production when mice are infected intraperitoneally with the gram-negative bacterium Klebsiella pneumoniae. To explore the
antioxidant properties of ascorbate in vivo, we therefore used K. pneumoniae
infection as a model of oxidative stress. When mice deficient in
L-gulono-gamma-lactone oxidase (Gulo(-/-)), the rate-limiting
enzyme in ascorbate synthesis, were depleted of ascorbate and infected with K. pneumoniae, they were three times as likely as ascorbate-replete Gulo(-/-)mice to die from
infection. Mass spectrometric analysis of peritoneal lavage fluid revealed a marked increase in the levels of oxidized
amino acids and of
F2-isoprostanes (sensitive and specific markers of
lipid oxidation) in infected animals. Surprisingly, there were no significant differences in the levels of the oxidation products in the ascorbate-deficient and -replete Gulo(-/-)mice. Our observations suggest that ascorbate plays a previously unappreciated role in host defense mechanisms against invading pathogens but that the
vitamin does not protect
amino acids and
lipids from oxidative damage during acute
inflammation. To examine the oxidation hypothesis of disease, optimal
antioxidant regimens that block oxidative reactions in animals and humans need to be identified.