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G Hoppe, Y-C Chai, JJ Mieyal, J Sears; Glutaredoxin Expression in Human Retinal Pigment Epithelial Cell Lines Inversely Correlates with the Levels of Protein Glutathiolation . Invest. Ophthalmol. Vis. Sci. 2002;43(13):712.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To compare expression of glutaredoxin and thioredoxin vis-à-vis the level of protein-glutathione (GSH) adducts, i.e., glutathiolation, in two human RPE cell lines, ARPE-19 and D407. Methods: Polymerase chain reaction was used to detect mRNA glutaredoxin in cDNA libraries of ARPE-19 and D407 cell lines. Antisera to glutaredoxin, thioredoxin, and protein-GSH adduct were used to probe lysates from ARPE-19 and D407. GSH levels were measured by high performance liquid chromatography. Results: Glutaredoxin transcripts from ARPE-19 and D407 were each 321 base pairs in length and DNA sequence was identical to human neutrophil glutaredoxin mRNA. Western blot demonstrated 7 fold more glutaredoxin protein expression in D407 compared to ARPE-19. Thioredoxin expression was equivalent in both cell lines. ARPE-19 had overall reduced amounts of GSH, as well as an inversion of GSH/glutathione disulfide (GSH/GSSG) ratio. Protein modification by glutathiolation was greater in ARPE-19 than in D407, both in naïve cells as well as after an oxidative challenge by diamide. Conclusion: ARPE-19 and D407 show differential expression of glutaredoxin, levels of cytosolic GSH, and protein glutathiolation. These data suggest that maintainence of unbound GSH is regulated by glutaredoxin in RPE cell lines ARPE-19 and D407.
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