Contribution of mitochondrial GSH transport to matrix GSH status and colonic epithelial cell apoptosis.
Previously, we showed that cellular glutathione/glutathione disulfide (GSH/GSSG) play an important role in apoptotic signaling, and early studies linked mitochondrial GSH (mtGSH) loss to enhanced cytotoxicity. The current study focuses on the contribution of mitochondrial GSH transport and mitochondrial GSH/GSSG status to apoptosis initiation in a nontransformed colonic epithelial cell line, NCM460, using menadione (MQ), a quinone with redox cycling bioreactivity, as a model of oxidative challenge. Our results implicate the semiquinone radical in MQ-mediated apoptosis, which was associated with marked oxidation of the mitochondrial soluble GSH and protein-bound thiol pools, mitochondria-to-cytosol translocation of cytochrome c, and activation of caspase-9. MQ-induced apoptosis was potentiated by inhibition of mtGSH uptake in accordance with exacerbated mitochondrial GSSG (mtGSSG) and protein-SSG and compromised mitochondrial respiratory activity. Moreover, cell apoptosis was prevented by N-acetyl-L-cysteine (NAC) pretreatment, which restored cellular redox homeostasis. Importantly, mtGSH transport inhibition effectively blocked NAC-mediated protection in accordance with its failure to attenuate mtGSSG. These results support the importance of mitochondrial GSH transport and the mtGSH status in oxidative cell killing.
Circu
Magdalena L
ML
Department of Molecular and Cellular Physiology, Louisiana State University Health Sciences Center, Shreveport, LA 71130, USA.
Rodriguez
Cynthia
C
Maloney
Ronald
R
Moyer
Mary Pat
MP
Aw
Tak Yee
TY
D000111:Acetylcysteine / Q000494:pharmacology; D000933:Antifibrinolytic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D001692:Biological Transport; D015153:Blotting, Western; D020169:Caspases / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D002478:Cells, Cultured; D003106:Colon / Q000187:drug effects / Q000378:metabolism*; D045304:Cytochromes c / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D001728:Dicumarol / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D005434:Flow Cytometry; D016166:Free Radical Scavengers / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism*; D019803:Glutathione Disulfide / Q000378:metabolism; D006801:Humans; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D010084:Oxidation-Reduction; D010101:Oxygen Consumption; D021381:Protein Transport; D024483:Vitamin K 3 / Q000494:pharmacology
D000933:Antifibrinolytic Agents; D004791:Enzyme Inhibitors; D016166:Free Radical Scavengers; D024483:Vitamin K 3; D001728:Dicumarol; D045304:Cytochromes c; D020169:Caspases; D005978:Glutathione; D019803:Glutathione Disulfide; D000111:Acetylcysteine