SLCO/OATP-like Transport of Glutathione in FasL-induced Apoptosis

Abstract

Apoptosis is a ubiquitous genetically encoded pathway that enables cells to undergo highly regulated death in response to specific signals. It has also been observed to occur as a consequence of distinct pathologies, and its deregulation can lead to autoimmune diseases, cancer, and neurodegenerative diseases (1). It is characterized by the activation of precise pathways leading to specific biochemical and morphological alterations, including changes in the intracellular ionic homeostasis, cell shrinkage, phosphatidylserine externalization, chromatin condensation, DNA degradation, membrane blebbing, and apoptotic body formation (1, 2). Fas ligand (FasL) 2 -induced apoptosis by binding to its receptor Fas (CD95/Apo-1) has been reported to play a critical role in immune homeostasis. Fas receptormediated apoptosis has also been implicated in the pathogenesis of fulminant hepatitis, postischemic neuronal degeneration, traumatic brain injury, and inflammatory lung diseases. Its deregulation is also associated with progression and metastasis of tumors and autoimmune lymphoproliferative syndrome (3).GSH is the predominant low molecular weight thiol in animal cells. It participates in many cellular reactions, including antioxidant defense of the cell, drug detoxification, and cell signaling, and has been reported to be necessary for the proliferation of several cells, including lymphocytes (4 -6). A reduction in the intracellular GSH concentration ([GSH] i ) has been reported during cell death in response to different apoptotic stimuli, including death receptor-induced (7-10), stress-induced, (11), and drug-induced cell death (12). Several studies have shown a correlation between GSH efflux and the progression of apoptosis (13,14), and a recent report suggests that inhibition of GSH release in apoptosis is able to rescue cells from apoptosis (8); however, the precise mechanisms involved in intracellular GSH (GSH i ) loss have not been identified. Glutathione synthesis occurs intracellularly, and its catabolism occurs by a series of both enzymatic and plasma membrane export mechanisms. Glutathione degradation occurs extracellularly; therefore, the export of GSH and GSH adducts is also an important step in its turnover (15). To date, two GSH transporter families have been implicated in GSH efflux across the plasma membrane. These include the multidrug-resistant pro-

Glutathione Depletion Is Necessary for Apoptosis in Lymphoid Cells Independent of Reactive Oxygen Species Formation

“…Cell debris or fragments were gated out from the analysis on a forward scatter/ side scatter plot. The appearance of discrete cell populations was identified as described previously (13). Briefly, gates were set on dot plots of the respective dye-fluorescence versus forward scatter, and then represented accordingly.…”

“…Changes in the GSH i are reflected by the appearance of populations of cells with differences in mBCl fluorescence, reflecting changes in GSH i . Populations were gated as previously described (13). Briefly, contour plots of mBCl fluorescence versus forward scatter were used to identify populations of cells with different GSH i levels induced after FasL treatment.…”

“…Additionally, high GSH levels have also been associated with an apoptotic resistant phenotype in different cells (18 -21). We have recently shown that Fas ligand (FasL)-induced GSH efflux is mediated by its extrusion across the plasma membrane in lymphoid cells with characteristics of GSH/organic anion transporter (13); however, the mechanisms involved in the regulation of apoptosis by changes in intracellular GSH are not completely understood.…”

“…In addition, alterations in its concentration have been demonstrated as a common feature of many pathological situations including AIDS, neurodegenerative diseases, and cancer (9, 10). Intracellular GSH loss is an early hallmark in the progression of cell death in response to different apoptotic stimuli (11)(12)(13)(14), and has been associated with the activation of a plasma membrane transport mechanism rather than to its oxidation (11)(12)(13)(15)(16)(17). Furthermore, several studies have shown a correlation between cellular GSH depletion and the progression of apoptosis (12, 13).…”

“…Intracellular GSH loss is an early hallmark in the progression of cell death in response to different apoptotic stimuli (11)(12)(13)(14), and has been associated with the activation of a plasma membrane transport mechanism rather than to its oxidation (11)(12)(13)(15)(16)(17). Furthermore, several studies have shown a correlation between cellular GSH depletion and the progression of apoptosis (12,13). Additionally, high GSH levels have also been associated with an apoptotic resistant phenotype in different cells (18 -21).…”