N-Acetyl-L-cysteine enhances apoptosis through inhibition of nuclear factor-kappaB in hypoxic murine embryonic fibroblasts.
In this study, we investigated the role of reduced glutathione (GSH) and nuclear factor-kappaB (NFkappaB) in hypoxia-induced apoptosis. Hypoxia caused p53-dependent apoptosis in murine embryonic fibroblasts transfected with Ras and E1A. N-Acetyl-l-cysteine (NAC) but not other antioxidants, such as the vitamin E analog trolox and epigallocatechin-3-gallate, enhanced hypoxia-induced caspase-3 activation and apoptosis. NAC also enhanced hypoxia-induced apoptosis in two human cancer cell lines, MIA PaCa-2 pancreatic cancer cells and A549 lung carcinoma cells. In murine embryonic fibroblasts, all three antioxidants blocked hypoxia-induced reactive oxygen species formation. NAC did not enhance hypoxia-induced cytochrome c release but did enhance poly-(ADP ribose) polymerase cleavage, indicating that NAC acted at a post-mitochondrial level. NAC-mediated enhancement of apoptosis was mimicked by incubating cells with GSH monoester, which increased intracellular GSH similarly to NAC. Hypoxia promoted degradation of an inhibitor of kappaB(IkappaBalpha), NFkappaB-p65 translocation into the nucleus, NFkappaB binding to DNA, and subsequent transactivation of NFkappaB, which increased X chromosome-linked inhibitor of apoptosis protein levels. NAC failed to block degradation by IkappaBalpha and sequestration of the p65 subunit of NFkappaB to the nucleus. However, NAC did abrogate hypoxia-induced NFkappaB binding to DNA, NFkappaB-dependent gene expression, and induction of X chromosome-linked inhibitor of apoptosis protein. In conclusion, NAC enhanced hypoxic apoptosis by a mechanism apparently involving GSH-dependent suppression of NFkappaB transactivation.
Qanungo
Suparna
S
Department of Anatomy and Case Comprehensive Cancer Center, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106, USA.
Wang
Mi
M
Nieminen
Anna-Liisa
AL
D000111:Acetylcysteine / Q000378:metabolism*; D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002470:Cell Survival / Q000502:physiology; D045304:Cytochromes c / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D000860:Hypoxia / Q000378:metabolism*; D020672:I-kappa B Proteins / Q000378:metabolism; D051379:Mice; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D051636:X-Linked Inhibitor of Apoptosis Protein
D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; C000606205:Nfkbia protein, mouse; D011506:Proteins; D017382:Reactive Oxygen Species; D051636:X-Linked Inhibitor of Apoptosis Protein; D000072000:NF-KappaB Inhibitor alpha; D045304:Cytochromes c; D005978:Glutathione; D000111:Acetylcysteine