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Fas-mediated apoptosis is modulated by intracellular glutathione in human T cells
1Department of Pathology 1, Sapporo Medical University School of Medicine, Japan.
Abstract:
Fas antigen is a member of the tumor necrosis factor receptor family that transduces a lethal signal to the Fas-sensitive cells. We previously established the Fas-resistant variant cell lines LAC2D1R and JKT2D1R from the parental Fas-sensitive cell lines, SUPT13 and Jurkat, respectively. Recently, we isolated the Fas-resistant variant CEM2D1R from CCRF-CEM. All of the variants were Fas+ but resistant to Fas-mediated apoptosis. Further biochemical analysis revealed that the intracellular glutathione (GSH) content of the Fas-resistant variants was higher than in the original cells. When the Fas-resistant variants were incubated with buthionine sulfoximine (BSO) or in GSH-free/cysteine-free medium to deplete GSH, Fas resistance was reversed. Incubation of the cells with cycloheximide also decreased intracellular GSH and reversed the Fas resistance. Furthermore, incubation of activated peripheral blood lymphocytes with BSO enhanced Fas-mediated apoptosis. When the Fas-sensitive cells were incubated with N-acetylcysteine (NAC), intracellular GSH was increased and Fas-mediated apoptosis was blocked. In contrast, Fas-resistant variants, as well as Fas-sensitive cells pre-treated with NAC remained susceptible to allogeneic lymphokine-activated killer cells, most likely due to perforin-dependent killing. The results suggest that Fas-mediated apoptosis, but not perforin-dependent killing, is modulated by intracellular GSH in human T lymphocytes.
Insights
Intracellular glutathione (GSH) levels regulate Fas-mediated apoptosis in human T lymphocytes. Depleting GSH reverses Fas resistance, while increasing GSH blocks this cell death pathway.
Area of Science:
- Immunology
- Cell Biology
- Biochemistry
Background:
- Fas antigen, a tumor necrosis factor receptor family member, initiates apoptosis in sensitive cells.
- Fas-resistant variants (LAC2D1R, JKT2D1R, CEM2D1R) were established from Fas-sensitive T lymphocyte lines.
- These variants express Fas but resist Fas-mediated programmed cell death.
Purpose of the Study:
- To investigate the role of intracellular glutathione (GSH) in Fas-mediated apoptosis.
- To determine if modulating GSH levels affects Fas sensitivity in human T lymphocytes.
Main Methods:
- Biochemical analysis of intracellular GSH content in Fas-sensitive and resistant cell lines.
- Treatment with buthionine sulfoximine (BSO) to deplete GSH.
- Treatment with N-acetylcysteine (NAC) to increase GSH.
- Assessment of apoptosis induction via Fas-mediated and perforin-dependent pathways.
Main Results:
- Fas-resistant variants exhibited higher intracellular GSH levels compared to parental cells.
- GSH depletion using BSO or GSH-free medium reversed Fas resistance.
- Cycloheximide treatment decreased intracellular GSH and reversed Fas resistance.
- BSO enhanced Fas-mediated apoptosis in activated peripheral blood lymphocytes.
- NAC treatment increased intracellular GSH, blocking Fas-mediated apoptosis.
- Cells remained susceptible to perforin-dependent killing irrespective of GSH levels or NAC treatment.
Conclusions:
- Intracellular glutathione (GSH) plays a critical role in modulating Fas-mediated apoptosis in human T lymphocytes.
- GSH levels can be manipulated to control sensitivity to Fas-induced cell death.
- Perforin-dependent cytotoxicity is independent of intracellular GSH modulation.