Reactive oxygen species generation and mitochondrial dysfunction in the apoptotic cell death of human myeloid leukemia HL-60 cells by a dietary compound withaferin A with concomitant protection by N-acetyl cysteine.
Induction of apoptosis in cancer cells has become the major focus of anti-cancer therapeutics development. WithaferinA, a major chemical constituent of Withania somnifera, reportedly shows cytotoxicity in a variety of tumor cell lines while its molecular mechanisms of action are not fully understood. We observed that withaferinA primarily induces oxidative stress in human leukemia HL-60 cells and in several other cancer cell lines. The withanolide induced early ROS generation and mitochondrial membrane potential (Deltapsi(mt)) loss, which preceded release of cytochrome c, translocation of Bax to mitochondria and apoptosis inducing factor to cell nuclei. These events paralleled activation of caspases -9, -3 and PARP cleavage. WA also activated extrinsic pathway significantly as evidenced by time dependent increase in caspase-8 activity vis-à-vis TNFR-1 over expression. WA mediated decreased expression of Bid may be an important event for cross talk between intrinsic and extrinsic signaling. Furthermore, withaferinA inhibited DNA binding of NF-kappaB and caused nuclear cleavage of p65/Rel by activated caspase-3. N-acetyl-cysteine rescued all these events suggesting thereby a pro-oxidant effect of withaferinA. The results of our studies demonstrate that withaferinA induced early ROS generation and mitochondrial dysfunction in cancer cells trigger events responsible for mitochondrial -dependent and -independent apoptosis pathways.
Malik
Fayaz
F
Division of Pharmacology, Indian Institute of Integrative Medicine, Council of Scientific and Industrial Research, Canal Road, Jammu-Tawi 180001, India.
Kumar
Ajay
A
Bhushan
Shashi
S
Khan
Sheema
S
Bhatia
Aruna
A
Suri
Krishan Avtar
KA
Qazi
Ghulam Nabi
GN
Singh
Jaswant
J
D000111:Acetylcysteine / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D004875:Ergosterol / Q000031:analogs & derivatives* / Q000494:pharmacology; D018922:HL-60 Cells; D006367:HeLa Cells; D006801:Humans; D007951:Leukemia, Myeloid / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D008928:Mitochondria / Q000187:drug effects / Q000473:pathology*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D032341:Withania; D054358:Withanolides
D000970:Antineoplastic Agents; D017382:Reactive Oxygen Species; D054358:Withanolides; C009684:withaferin A; D000111:Acetylcysteine; D004875:Ergosterol