beta-Phenylethyl isothiocyanate-mediated apoptosis in hepatoma HepG2 cells.
beta-Phenylethyl isothiocyanate (PEITC) is a promising chemoprotective compound that is routinely consumed in the diet as its glucosinolate precursor. Previous studies have shown that PEITC can inhibit phase I enzymes and induce phase II detoxification enzymes along with apoptosis in vitro. The detailed mechanisms involved in the apoptotic cascade, however, have not been elucidated. In the present study, we demonstrate that PEITC can induce apoptosis in hepatoma HepG2 cells in a concentration- and time-dependant manner as determined by TUNEL positive and SubG1 population analysis. Caspase-3-like activity and poly(ADP-ribosyl)polymerase cleavage increased during treatment with 20 microM PEITC; high concentrations, however, induced necrosis. Pre-treatment with Z-VAD-FMK and the caspase-3-specific inhibitor Ac-DEVD-CHO prevented PEITC-induced apoptosis, as determined by caspase-3-like activity and DNA fragmentation. Additional investigations also showed that at concentrations of 5-10 microM PEITC, DNA synthesis was inhibited and G2/M phase cell cycle arrest occurred, correlating with an alteration in cyclin B1 and p34(cdc2) protein levels. Furthermore, we also demonstrate a concentration- and time-dependant burst of superoxide (O2*-) in PEITC-treated cells. However, pre- and co-treatment with the free radical scavengers Trolox, ascorbate, mannitol, uric acid and the superoxide mimetic manganese (III) tetrakis (N-methyl-2-pyridyl) porphyrin failed to prevent PEITC-mediated apoptosis. Taken together, these results suggest that PEITC potently induces apoptosis and cell cycle arrest in HepG2 cells and that the generation of reactive oxygen species appears to be a secondary effect.
Rose
P
P
Department of Community, Occupational and Family Medicine, MD3, National University of Singapore, 16 Medical Drive, 117597 Singapore. cofpcr@nus.edu.sg
Whiteman
M
M
Huang
S H
SH
Halliwell
B
B
Ong
C N
CN
D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001973:Bromodeoxyuridine / Q000493:pharmacokinetics; D006528:Carcinoma, Hepatocellular; D002453:Cell Cycle / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004261:DNA Replication / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D006801:Humans; D020287:In Situ Nick-End Labeling; D008658:Inactivation, Metabolic; D017879:Isothiocyanates / Q000494:pharmacology*; D007700:Kinetics; D008113:Liver Neoplasms; D013481:Superoxides / Q000378:metabolism; D014407:Tumor Cells, Cultured
D016588:Anticarcinogenic Agents; D017879:Isothiocyanates; D013481:Superoxides; C058305:phenethyl isothiocyanate; D001973:Bromodeoxyuridine