Cytotoxicity and apoptosis induction by butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT).
The cytotoxicity and apoptosis-inducing activity of butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), and 2-tert-butyl-4-methylphenol (BMP) and the mixture of BHA and BHT (BHA/BHT) (1:1, molar ratio) were investigated, using human promeylocytic leukemia cell lines (HL-60) and human squamous cell carcinoma cell lines (HSC-2). The 50% cytotoxic concentration (CC50) declined in the order of BHA, BHT (0.2-0.3 mM) > BHA/BHT (0.04-0.07 mM) > BMP (0.02-0.05 mM). The addition of antioxidants (N-acetyl-Lcysteine, sodium ascorbate, catalase) reduced the cytotoxicity of BHA/BHT or BMP against HSC-2 cells, but not that of BHA or BHT, whereas the addition of NADH, a quinone reductase to BMP, enhanced the cytotoxicity. These findings suggested that the cytotoxicity of BHA/BHT and BMP might be caused by reactive intermediates. BHA-induced cytotoxicity was enhanced by horseradish peroxidases, suggesting that BHA was oxidizable and produced cytotoxic BHA radicals. Internucleosomal DNA fragmentation of HL-60 cells was preferably induced by BHA/BHT and BMP, followed by BHA. The MnSOD mRNA expression in HL-60 cells assayed by reverse transcriptase-polymerase chain reaction was highly inhibited by BHA/BHT or BMP, accompanied by the change in the electrophoretic mobility of MnSOD on polyacryamide gel. These compounds activated caspase-3, 8 and 9 in HL-60 cells. Activations of caspases, particularly caspase-3, declined in the order of BHA/BHT > BHA > BMP > BHT. The most cytotoxic BMP activated caspase-3 activity to the least extent, possibly in part due to the occurrence of necrosis. The great cytotoxicity and apoptosis induction by BHA/BHT may be due to reactive intermediates derived from the interaction between BHA phenoxyl radical and BHT or BHT phenoxyl radical.
Department of Oral Diagnosis, Meikai University School of Dentistry, Saitama 350-0283, Japan.
Masatoshi
M
Saito
Hiroshi
H
Sakagami
Seiichiro
S
Fujisawa
D000975:Antioxidants / Q000633:toxicity; D017209:Apoptosis / Q000187:drug effects*; D002083:Butylated Hydroxyanisole / Q000633:toxicity*; D002084:Butylated Hydroxytoluene / Q000633:toxicity*; D002273:Carcinogens / Q000633:toxicity*; D002294:Carcinoma, Squamous Cell; D002470:Cell Survival / Q000187:drug effects*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D018922:HL-60 Cells; D006801:Humans; D009243:NAD / Q000494:pharmacology*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D013482:Superoxide Dismutase / Q000235:genetics; D014407:Tumor Cells, Cultured
D000975:Antioxidants; D002273:Carcinogens; D012333:RNA, Messenger; D009243:NAD; D002084:Butylated Hydroxytoluene; D002083:Butylated Hydroxyanisole; D013482:Superoxide Dismutase