Glutathione stimulates A549 cell proliferation in glutamine-deficient culture: the effect of glutamate supplementation.

Extracellular glutathione (GSH) is degraded by an external cell-surface enzyme, gamma-glutamyltranspeptidase (gamma-GT). The products are transported into cells to participate in important cellular processes. In the present study, we tested the hypothesis that extracellular GSH is a source of glutamic acid for cells that express gamma-GT. Under a glutamine-deficient culture condition, the extracellular GSH-supplemented glutamic acid would enhance intracellular glutamine synthesis, thereby stimulating cell proliferation. Human lung carcinoma A549 cells were cultured in glutamine-deficient Dulbecco's modified Eagle medium, and they did not proliferate unless glutamine was supplemented. Extracellular GSH, however, provoked a partial proliferation. The GSH effect correlated with a high level of gamma-GT activity and an increased intracellular level of glutamic acid. A constituent amino acid of GSH, glutamic acid but not cysteine, produced the same growth-stimulatory effect as GSH. Furthermore, neither oxothiazolidine-4-carboxylate (OTC), a cellular cysteine-delivery compound, nor cysteinylglycine, a dipeptide released from the gamma-GT reaction, stimulated cell proliferation. Moreover, buthionine sulfoximine (BSO), a selective inhibitor of gamma-glutamylcysteine synthetase, enhanced the GSH growth stimulatory effect, suggesting that increased cellular GSH synthesis does not correlate with cell growth stimulation. The results obtained demonstrated that glutamine is required for A549 cell proliferation and exogenous GSH partially substitutes for the growth stimulatory action of glutamine. It also suggests that the glutamic acid rather than the cysteine released from the GSH is responsible for the cell proliferation.