Decrease in Intracellular Glutathione Level Alters Expressions of B-cell CLL/Lymphoma 2 Family Members in the Mouse Retina

Abstract

Oxidative stress affects all intracellular macromolecules, and leads cells to death under unfavorable conditions. Glutathione (GSH) is known to play a critical role in the cellular defense against unregulated oxidative stress in mammalian cells including neurons. We previously demonstrated that GSH depletion induces cell death in the retina, but the mechanism of cell defense by GSH is still unclear. Thus, we here examined the effect of GSH depletion on expression of members of B-cell CLL/lymphoma 2 (Bcl-2) family (bcl-2, bcl-X_L, bax, bak, n-bak and bad) known to play key roles in determining cell viability. In order to deplete intracellular GSH, we systemically administrated buthionine sulphoximine (BSO), an inhibitor of γ-glutamylcysteine synthetase to mice. After 0, 1, 4, and 7 days of BSO injection, total RNAs from retina of each animals were isolated and subjected to real-time reverse transcription (RT)-polymerase chain reaction (PCR) analysis. Expression of bcl-X_L increased one day after BSO injection, but was back to the basal level on day 4. Its expression decreased on day 7. Expressions of bcl-2 and bax were significantly decreased from 4 days after BSO injection, whereas expression of bad was not changed. An anti-apoptotic molecule, bak displayed a significant decrease 7 days after BSO injection, whereas neuronal cell specific Bak (Bcl-2 homologous antagonist/killer), N-Bak was not altered in its gene expression. Taken together, we demonstrated that decrease in intracellular glutathione level altered expressions of Bcl-2 family members in distinct manners. Our study implies a defensive mechanism of GSH against oxidative stress for retinal neuronal survival which may involve alteration of expression of Bcl-2 family members, especially Bcl-2 and Bcl-X_L. Thus, over-expression of Bcl-2 and Bcl-X_L may not the only but a critical factor in GSH-dependent cellular protection, and which implies Bcl-2 and Bcl-X_L may provide a potent therapeutic tool for cures against oxidative stress induced retinal degenerative diseases such as glaucoma, retinopathy, and age-related macular degeneration.