Activation of plasma membrane reduced glutathione transport in death receptor apoptosis of HepG2 cells.

Cells undergoing apoptosis release reduced glutathione (GSH) into the extracellular space; however, the physiological significance and the mechanism behind the GSH export remain unclear. The present study demonstrates that GSH is released by HepG2 cells undergoing Fas, tumor necrosis factor alpha (TNFalpha), or tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-stimulated cell death. GSH release was observed at times when extracellular lactate dehydrogenase (LDH) activity and propidium iodide (PI) incorporation were low, suggesting that the GSH release does not occur because of nonspecific cell damage, but is occurring through a specific transport system. Caspase 3-like proteases were activated before GSH was released, indicating that protease may be involved in signaling GSH release. To investigate the mechanism of GSH release, studies were performed in the presence of GSH transport inhibitors, as well as 25 mM GSH in the media. Two organic anion transporter inhibitors, probenecid and dibromosulfophthalein (DBSP), were effective in inhibiting Fas-stimulated GSH release. The addition of 25 mM GSH to the extracellular media also prevented the loss of intracellular GSH and delayed cell death. These findings suggest that an organic anion transporter is involved in GSH release during apoptosis, and that maintenance of intracellular GSH levels during apoptosis provides protection for the cell.