Abstract
The therapeutic immunopharmacological potential of glutathione in the alveolar epithelium is not well characterized. We developed an in vitro model of fetal alveolar type II epithelial cells to investigate the effect of redox disequilibrium on chemioxyexcitation (ΔpO2/ROS) induced up-regulation of pro-inflammatory cytokines. Buthionine sulfoximine, an irreversible inhibitor of γ-glutamylcysteine synthetase, the rate-limiting enzyme in glutathione (GSH) biosynthesis, induced intracellular reactive oxygen species (ROS) and the release of interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α. Chloroethyl nitrosourea, which blocks the NADPH-dependent recycling of oxidized glutathione (GSSG), reduced ROS-induced cytokine production, similar to pyrrolidine dithiocarbamate, an antioxidant/pro-oxidant thiuram, which elevates GSSG. The antioxidant and GSH precursor, acetylcysteine, abrogated cytokine release concomitant with suppression of ROS, an effect mimicked by γ-glutamylcysteinyl-ethyl ester, a cell permeant GSH. Cysteine, the rate-limiting amino acid in the de novo synthesis of GSH, administered as oxothiazolidine carboxylate and adenosylmethionine, mitigated the chemioxyexcitation-dependent cytokine release. Ebselen, an anti-inflammatory antioxidant, which mimics the effect of glutathione peroxidase, neutralized ROS by the GSH-peroxidase-coupled reaction, thereby blocking the pathway to cytokine enhancement. Our results indicate that modulating redox equilibrium by pharmacological thiols exhibits differential regulation on pro-inflammatory cytokines, thus bearing clinical consequences for the therapeutic treatment of pediatric distresses in pathophysiology.
Footnotes
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Send reprint requests to: Dr. John J. E. Haddad, Oxygen Signaling Group, Center for Research into Human Development, Tayside Institute of Child Health, Faculty of Medicine, Ninewells Hospital and Medical School, University of Dundee, Dundee DD1 9SY, Scotland, UK. E-mail: j.j.haddad{at}dundee.ac.uk
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This work was supported by grants from the Medical Research Council, Anonymous Trust and Tenovus-Scotland (S.C.L.). J.J.H. is a recipient of the George John Livanos prize (London). Part of this work was presented at Experimental Biology-2000, San Diego, CA (Haddad et al., 2000b).
- Abbreviations:
- GSH
- l-γ-glutamyl-l-cysteinyl-glycine
- NAC
- N-acetyl-l-cysteine
- SAM
- S-adenosyl-l-methionine
- BCNU
- 1,3-bis-(2-chloroethyl)-1-nitrosourea
- BSO
- l-buthionine-(S,R)-sulfoximine
- DHR
- dihydrorhodamine
- γ-GCE
- γ-glutamyl-l-cysteinyl-ethyl ester
- GSSG
- glutathione disulfide oxidized
- GSH-PX
- glutathione peroxidase
- GSSG-RD
- glutathione reductase
- γ-GCS
- γ-glutamylcysteine synthetase
- GS
- glutathione synthase
- IL
- interleukin
- OTC
- 2-oxothiazolidine-4-carboxylate
- ebselen
- 2-phenyl-1,2-benzisoselenazol-3(2H)-one
- PDTC
- pyrrolidine dithiocarbamate
- ROS
- reactive oxygen species
- redox
- reduction-oxidation
- TNF-α
- tumor necrosis factor-α
- X/XO
- xanthine/xanthine oxidase
- DMEM
- Dulbecco's modified Eagle's medium
- FCS
- fetal calf serum
- EU
- enzyme unit activity
- HIF1-α
- hypoxia-inducible factor 1α
- NF-κB
- nuclear factor-κB
- Received August 25, 2000.
- Accepted November 17, 2000.
- The American Society for Pharmacology and Experimental Therapeutics
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