title: Control of extracellular cysteine/cystine redox state by HT-29 cells is independent of cellular glutathione.
abstract: Human cell lines regulate the redox state (E(h)) of the cysteine/cystine (Cys/CySS) couple in culture medium to approximately -80 mV, a value similar to the average E(h) for Cys/CySS in human plasma. The mechanisms involved in regulation of extracellular E(h) of Cys/CySS are not known, but GSH is released from tissues at rates proportional to tissue GSH concentration, and this released GSH could react with CySS to contribute to maintenance of this balance. The present study was undertaken to determine whether depletion of cellular GSH alters regulation of extracellular Cys/CySS E(h). Decrease of GSH in HT-29 cells by inhibiting synthesis with l-buthionine-[S,R]-sulfoximine showed no effect on the rate of reduction of extracellular CySS to achieve a stable E(h) for Cys/CySS in the culture medium. Limiting Cys and CySS in the culture medium also substantially decreased cellular GSH but resulted in no significant effect on extracellular Cys/CySS E(h). Addition of CySS to these cells showed that extracellular Cys/CySS E(h) approached -80 mV at 4 h while cellular GSH and extracellular GSH/GSSG E(h) recovered more slowly. Together, these results show that HT-29 cells have the capacity to regulate the extracellular Cys/CySS E(h) by mechanisms that are independent of cellular GSH. The results suggest that transport systems for Cys and CySS and/or membranal oxidoreductases could be more important than cellular GSH in regulation of extracellular Cys/CySS E(h).
authors.affiliation: Department of Medicine/Pulmonary, Emory University School of Medicine, 615 Michael Street, Atlanta, GA 30322, USA.
authors.forename: Corinna L
authors.initials: CL
authors.lastname: Anderson
authors.forename: Smita S
authors.initials: SS
authors.lastname: Iyer
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Ziegler
authors.forename: Dean P
authors.initials: DP
authors.lastname: Jones
mesh_terms: D001693:Biological Transport, Active / Q000187:drug effects; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D003470:Culture Media; D003545:Cysteine / Q000172:deficiency / Q000378:metabolism*; D003553:Cystine / Q000172:deficiency / Q000378:metabolism*; D004220:Disulfides / Q000378:metabolism; D005110:Extracellular Space / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism*; D019073:HT29 Cells; D006801:Humans; D007700:Kinetics; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000187:drug effects; D013438:Sulfhydryl Compounds / Q000378:metabolism
chemical_list: D003470:Culture Media; D004220:Disulfides; D013438:Sulfhydryl Compounds; D003553:Cystine; D019328:Buthionine Sulfoximine; D005978:Glutathione; D003545:Cysteine