title: The role of glutathione in chronic adaptation to oxidative stress: studies in a normal rat kidney epithelial (NRK52E) cell model of sustained upregulation of glutathione biosynthesis.
abstract: Reduced glutathione (GSH) is considered to play a central role in protection of cells from oxidant injury. However, the question remains as to whether sustained elevation of intracellular GSH levels, as compared with the ability to rapidly upregulate GSH synthesis, is more important with respect to protection of cell constituents from oxidative stress. To address this question, we conducted studies to evaluate the direct influence of chronically increased endogenous GSH content on chemically induced intracellular free radical formation and oxidative stress using a kidney epithelial cell model adapted to sustain intracellular GSH concentrations in excess of eightfold that observed in unadapted parent kidney cells. Elevated GSH levels in adapted cells were found to be attributable, at least in part, to coordinately increased amounts of both the regulatory and catalytic subunits of gamma-glutamylcysteine synthetase (GCS), the rate-limiting enzyme in GSH synthesis. Studies using electron spin resonance (ESR) spectroscopy and scanning laser cytometry demonstrated that cells having sustained elevation of GSH levels did not attenuate free radical formation and associated oxidative stress compared with parent cells when treated with the prooxidant chemicals, menadione or potassium dichromate. In contrast, nonadapted kidney parent cells treated 18 h after initial prooxidant challenge displayed significantly attenuated free radical signals. Additionally, cells adapted to sustain excess GSH were somewhat more sensitive than parent cells in terms of resistance to prooxidant (chromate) toxicity, as determined by cell viability assays. These findings suggest that the capacity of cells to rapidly upregulate GSH synthesis, rather the ability to chronically sustain elevated intracellular GSH levels, may play a more important role in terms of protection from cytotoxicity associated with prooxidant chemical exposures.
authors.affiliation: Department of Environmental Health, University of Washington, Suite 100, Seattle, Washington, 98105, USA. jwoods@u.washington.edu
authors.forename: J S
authors.initials: JS
authors.lastname: Woods
authors.forename: T J
authors.initials: TJ
authors.lastname: Kavanagh
authors.forename: J
authors.initials: J
authors.lastname: Corral
authors.forename: A W
authors.initials: AW
authors.lastname: Reese
authors.forename: D
authors.initials: D
authors.lastname: Diaz
authors.forename: M E
authors.initials: ME
authors.lastname: Ellis
mesh_terms: D000222:Adaptation, Physiological; D000818:Animals; D015152:Blotting, Northern; D002460:Cell Line; D002469:Cell Separation; D002470:Cell Survival; D004578:Electron Spin Resonance Spectroscopy; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D005434:Flow Cytometry; D005609:Free Radicals / Q000378:metabolism; D005721:Glutamate-Cysteine Ligase / Q000235:genetics / Q000378:metabolism; D005978:Glutathione / Q000096:biosynthesis*; D007668:Kidney / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008298:Maleates / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D011192:Potassium Dichromate / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D019097:Spin Trapping; D015854:Up-Regulation; D014812:Vitamin K / Q000494:pharmacology
chemical_list: D005609:Free Radicals; D008298:Maleates; D012333:RNA, Messenger; D014812:Vitamin K; D005721:Glutamate-Cysteine Ligase; C014476:diethyl maleate; D005978:Glutathione; D011192:Potassium Dichromate