Glutathione depletion by DL-buthionine-SR-sulfoximine (BSO) potentiates X-ray-induced chromosome lesions after liquid holding recovery

The impact of intracellular glutathione depletion on chromosome damage induced by X irradiation under aerobic conditions was investigated in two different cell lines, Ehrlich ascites tumor cells (EATC) and Chinese hamster ovary cells (CHO-K1). Thiol-depleted cell cultures in plateau phase were obtained by prolonged incubation in growth medium containing DL-buthionine-SR-sulfoximine (BSO), a specific inhibitor of gamma-glutamyl-cysteine synthetase. Cells were then assayed using the procedures of G. L. Ellmann and J. Sedlack and R.H. Lindsay for non-protein bound SH (NPSH), glutathione (GSH), and total SH (TSH). In both cell lines GSH was reduced to less than 10% of controls at higher BSO concentrations around 1 mM, whereas TSH and NPSH were affected to only 40-60%. In EATC pretreated with up to 1 mM BSO for 72 h, increased levels of spontaneously occurring micronuclei were found. At BSO concentrations above 200 microM, both cell lines showed a potentiation of chromosome lesions scored as micronuclei and induced under aerobic X irradiation when liquid holding recovery in the original nutrient-depleted medium was performed; the extent of chromosome damage eventually reached that which could be obtained by application of beta-arabinofuranosyladenine (beta-araA), known to inhibit DNA repair processes by blocking DNA polymerases. It is therefore suggested that GSH depletion causes impairment of repair of lesions leading to chromosome deletions and subsequently to micronuclei. In contrast to CHO cell cultures, EATC showed a reversion of the potentiation effect as indicated by a decrease in the micronucleus content during prolonged incubation in the presence of BSO in the millimolar range