Cellular effects of photogenerated oxidants and long-lived, reactive, hydroperoxide photoproducts.
Reaction of radicals and singlet oxygen ((1)O(2)) with proteins results in both direct damage and the formation of long-lived reactive hydroperoxides. Elevated levels of protein hydroperoxide-derived products have been detected in multiple human pathologies, suggesting that these secondary oxidants contribute to tissue damage. Previous studies have provided evidence for protein hydroperoxide-mediated inhibition of thiol-dependent enzymes and modulation of signaling processes in isolated systems. In this study (1)O(2) and hydroperoxides have been generated in J774A.1 macrophage-like cells using visible light and the photosensitizer rose bengal, with the consequences of oxidant formation examined both immediately and after subsequent (dark-phase) incubation. Significant losses of GSH (≤50%), total thiols (≤20%), and activity of thiol-dependent proteins (GAPDH, thioredoxin, protein tyrosine phosphatases, creatine kinase, and cathepsins B and L; 10-50% inhibition) were detected after 1 or 2 min photo-oxidation. Non-thiol-dependent enzymes were not affected. In contrast, NADPH levels increased, together with the activity of glutathione reductase, glutathione peroxidase, and thioredoxin reductase; these increases may be components of a rapid global cytoprotective cellular response to stress. Neither oxidized thioredoxin nor radical-mediated protein oxidation products were detected at significant levels. Further decreases in thiol levels and enzyme activity occurred during dark-phase incubation, with this accompanied by decreased cell viability. These secondary events are ascribed to the reactions of long-lived hydroperoxides, generated by (1)O(2)-mediated reactions. Overall, this study provides novel insights into early cellular responses to photo-oxidative damage and indicates that long-lived hydroperoxides can play a significant role in cellular damage.
Rahmanto
Aldwin Suryo
AS
The Heart Research Institute, Newtown, Sydney, NSW 2042, Australia.
Morgan
Philip E
PE
Hawkins
Clare L
CL
Davies
Michael J
MJ
D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D005978:Glutathione / Q000378:metabolism; D005979:Glutathione Peroxidase / Q000378:metabolism; D005980:Glutathione Reductase / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D016877:Oxidants / Q000378:metabolism*; D055668:Photochemical Processes; D011506:Proteins / Q000378:metabolism; D012395:Rose Bengal / Q000378:metabolism; D026082:Singlet Oxygen / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000378:metabolism; D013879:Thioredoxins / Q000378:metabolism
D000975:Antioxidants; D016877:Oxidants; D011506:Proteins; D013438:Sulfhydryl Compounds; D026082:Singlet Oxygen; D012395:Rose Bengal; D013879:Thioredoxins; D006861:Hydrogen Peroxide; D005979:Glutathione Peroxidase; D005980:Glutathione Reductase; D005978:Glutathione