Glutathione determination by the Tietze enzymatic recycling assay and its relationship to cellular radiation response.
Large fluctuations in glutathione content were observed on a daily basis using the Tietze enzyme recycling assay in a panel of six human cell lines of varying radiosensitivity. Glutathione content tended to increase to a maximum during exponential cell proliferation, and then decreased at different rates as the cells approached plateau phase. By reference to high-performance liquid chromatography and flow cytometry of the fluorescent bimane derivative we were able to verify that these changes were real. However, the Tietze assay was occasionally unable to detect glutathione in two of our cell lines (MGH-U1 and AT5BIVA), although the other methods indicated its presence. The existence of an inhibitory activity responsible for these anomalies was confirmed through spiking our samples with known amounts of glutathione. We were unable to detect a direct relationship between cellular glutathione concentration and aerobic radiosensitivity in our panel of cell lines.
Eady
J J
JJ
Radiotherapy Research Unit, Institute of Cancer Research, Sutton, Surrey, UK.
Orta
T
T
Dennis
M F
MF
Stratford
M R
MR
Peacock
J H
JH
D000332:Aerobiosis; D001260:Ataxia Telangiectasia / Q000473:pathology*; D002295:Carcinoma, Transitional Cell / Q000473:pathology; D002461:Cell Line, Transformed; D002851:Chromatography, High Pressure Liquid; D004228:Dithionitrobenzoic Acid / Q000378:metabolism; D005347:Fibroblasts / Q000737:chemistry / Q000201:enzymology / Q000528:radiation effects; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D005978:Glutathione / Q000031:analogs & derivatives / Q000032:analysis* / Q000172:deficiency; D005980:Glutathione Reductase / Q000378:metabolism*; D006801:Humans; D008545:Melanoma / Q000473:pathology; D009249:NADP / Q000378:metabolism; D009363:Neoplasm Proteins / Q000378:metabolism*; D014411:Neoplastic Stem Cells / Q000737:chemistry* / Q000201:enzymology / Q000528:radiation effects; D009447:Neuroblastoma / Q000473:pathology; D009579:Nitrobenzoates / Q000378:metabolism*; D010084:Oxidation-Reduction; D011836:Radiation Tolerance; D013438:Sulfhydryl Compounds; D014407:Tumor Cells, Cultured; D001749:Urinary Bladder Neoplasms / Q000473:pathology
D005456:Fluorescent Dyes; D009363:Neoplasm Proteins; D009579:Nitrobenzoates; D013438:Sulfhydryl Compounds; C011136:thionitrobenzoic acid; D009249:NADP; D004228:Dithionitrobenzoic Acid; D005980:Glutathione Reductase; D005978:Glutathione