ascorbic-acid and dibutylnitrosamine

The aim of this work was to determine the effect of vitamin C, diallyl disulfide (DADS) and dipropyl disulfide (DPDS) towards N-nitrosopiperidine (NPIP) and N-nitrosodibutylamine (NDBA)-induced apoptosis in human leukemia (HL-60) and hepatoma (HepG2) cell lines using the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay. None of the vitamin C (5-50 microm), DADS and DPDS (1-5 microm) concentrations selected induced a significant percentage of apoptosis. In simultaneous treatments, vitamin C, DADS and DPDS reduced the apoptosis induced by NPIP and NDBA in HL-60 and HepG2 cells (around 70% of reduction). We also investigated its scavenging activities towards reactive oxygen species (ROS) produced by NPIP and NDBA using 2',7'-dichlorodihydrofluorescein diacetate in both cell lines. ROS production induced by both N-nitrosamine was reduced to control levels by vitamin C (5-50 microm) in a dose-dependent manner. However, DADS (5 microm) increased ROS levels induced by NPIP and NDBA in HL-60 (40 and 20% increase, respectively) and HepG2 cells (18% increase), whereas DPDS was more efficient scavenger of ROS at the lowest concentration (1 microm) in both HL-60 (52 and 25% reduction, respectively) and HepG2 cells (24% reduction). The data demonstrated that the scavenging ability of vitamin C and DPDS could contribute to inhibition of the NPIP- and NDBA-induced apoptosis. However, more than one mechanism, such as inhibition of phase I and/or induction of phase II enzymes, could be implicated in the protective effect of dietary antioxidants towards NPIP- and NDBA-induced apoptosis in HL-60 and HepG2 cells.

Topics: Allyl Compounds; Antioxidants; Apoptosis; Ascorbic Acid; Carcinoma, Hepatocellular; Cell Culture Techniques; Dietary Supplements; Disulfides; Dose-Response Relationship, Drug; HL-60 Cells; Humans; In Situ Nick-End Labeling; Leukemia, Promyelocytic, Acute; Liver Neoplasms; Nitrosamines; Oxidative Stress; Reactive Oxygen Species

The aim of this study was to investigate the protective effect of isothiocyanates alone or in combination with vitamin C towards N-nitrosodibutylamine (NDBA) or N-nitrosopiperidine (NPIP)-induced oxidative DNA damage in the single cell gel electrophoresis (SCGE)/HepG2 assay. Phenethyl isothiocyanate (PEITC) and indole-3-carbinol (I3C) alone showed a weak protective effect towards NDBA (0.1 microm, 26-27%, respectively) or NPIP (1 microm, 26-28%, respectively)-induced oxidative DNA damage. Allyl isothiocyanate (AITC) alone did not attenuate the genotoxic effect provoked by NDBA or NPIP. In contrast, HepG2 cells simultaneously treated with PEITC, I3C and AITC in combination with vitamin C showed a stronger inhibition of oxidative DNA-damage induced by NDBA (0.1 microm, 67%, 42%, 32%, respectively) or NPIP (1 microm, 50%, 73%, 63%, respectively) than isothiocyanates (ITCs) alone. One feasible mechanism by which ITCs alone or in combination with vitamin C exert their protective effects towards N-nitrosamine-induced oxidative DNA damage could be by the inhibition of their cytochrome P450 dependent bioactivation. PEITC and I3C strongly inhibited the p-nitrophenol hydroxylation (CYP2E1) activity (0.1 microm, 66-50%, respectively), while the coumarin hydroxylase (CYP2A6) activity was slightly reduced (0.1 microm, 25-37%, respectively). However, the ethoxyresorufin O-deethylation (CYP1A1) activity was only inhibited by PEITC (1 microm, 55%). The results indicate that PEITC and I3C alone or PEITC, I3C and AITC in combination with vitamin C protects human-derived cells against the oxidative DNA damaging effects of NDBA and NPIP, two food carcinogenic compounds.

Topics: Anticarcinogenic Agents; Aryl Hydrocarbon Hydroxylases; Ascorbic Acid; Carcinogens; Carcinoma, Hepatocellular; Cell Line, Tumor; Comet Assay; Cytochrome P-450 CYP1A1; Cytochrome P-450 CYP2A6; Cytochrome P-450 CYP2E1; Cytochrome P-450 CYP2E1 Inhibitors; DNA Breaks; DNA-Formamidopyrimidine Glycosylase; Dose-Response Relationship, Drug; Enzyme Inhibitors; Humans; Indoles; Isothiocyanates; Liver Neoplasms; Microsomes, Liver; Mixed Function Oxygenases; Nitrosamines; Oxidative Stress

The aim of this study was to investigate the protective effects of organosulfur compounds (OSCs) alone or in combination with vitamin C towards N-nitrosopiperidine (NPIP) and N-nitrosodibutylamine (NDBA)-induced oxidative DNA damage in the single cell gel electrophoresis (SCGE)/HepG2 assay. Diallyl sulfide (DAS) did not protect against NDBA-induced oxidized purines, but it reduced the oxidized purines induced by NPIP (1 microM, 29%). The formation of formamidopyridine-DNA glycosylase (Fpg) sensitive sites induced by NPIP or NDBA was prevented by dipropyl sulfide (DPS) at concentrations of 1-10 microM (55-24% and 66-15%, respectively). The maximum reduction of the formation of Fpg sensitive sites induced by NPIP was observed at the highest concentration of diallyl disulfide (DADS) (2.5 microM, 38%). However, the oxidative DNA damage induced by NDBA was strongly reduced by DADS at the lowest concentration tested (0.1 microM, 92%). The oxidative DNA damage induced by NPIP or NDBA was prevented by all the concentrations of dipropyl disulfide (DPDS) (0.1-2.5 microM, 59-80% and 51-64%, respectively). DADS and DPDS, in combination with vitamin C showed an overall protective effect towards the formation of Fpg sensitive sites induced by NPIP and NDBA. However, the contribution of OSCs to the protective effect found in combined experiments might not be relevant, because it could be caused by vitamin C alone. One feasible mechanism by which OSCs exert their protective effects towards N-nitrosamine-induced oxidative DNA damage could be by modulation of phase I and II enzyme activities. DADS and DPDS (0.1-2.5 microM) exerted greater inhibition on CYP2E1 and CYP2A6 activity than DAS and DPS (1-50 microM). However, DAS and DADS (1 microM) exerted greater inhibition on CYP1A1 activity than DPS and DPDS (1 microM). DAS/DPS (50 microM) and DADS (2.5 microM) exerted a moderate increase of UDP-glucuronyltransferase (UGT1A4) activity, whereas DPDS (2.5 microM) had the most pronounced effect.

Topics: Ascorbic Acid; Cell Line, Tumor; Cytochrome P-450 Enzyme System; DNA Damage; Humans; Nitrosamines; Oxidative Stress; Recombinant Proteins; Sulfur Compounds

The aim of this study was to investigate the protective effect of vitamin C towards N-nitrosamine-induced DNA damage in the single-cell gel electrophoresis (SCGE)/HepG2 assay. None of the vitamin C concentrations tested (1-10 microM) in presence or absence of formamidopyrimidine-DNA glycosylase (Fpg enzyme) caused DNA damage per se. HepG2 cells simultaneously treated with vitamin C and N-nitrosodimethylamine (NDMA), N-nitrosopyrrolidine (NPYR), N-nitrosodibutylamine (NDBA) or N-nitrosopiperidine (NPIP) reduced the genotoxic effects of the N-nitrosamines in a dose-dependent manner. At concentrations of 1-5 microM vitamin C, the protective effect was higher towards NPYR-induced oxidative DNA damage (78-79%) than against NDMA (39-55%), NDBA (12-14%) and NPIP (3-55%), in presence of Fpg enzyme. However, a concentration of 10 microM vitamin C led to a maximum reduction in NDBA (94%), NPYR (81%), NPIP (80%) and NDMA (61%)-induced oxidative DNA damage, in presence of Fpg enzyme. The greatest protective effect of vitamin C (10 microM) was higher towards NDBA-induced oxidative DNA damage. One feasible mechanism by which vitamin C exerted its protective effect is that may interact with the enzyme systems catalyzing the metabolic activation of the N-nitrosamines, blocking the production of genotoxic intermediates. Vitamin C (10 microM) strongly reduced the coumarin hydroxylase (82%) activity. However, the p-nitrophenol hydroxylase and the ethoxyresorufine O-deethylation activities were slightly and weakly reduced (32-19%), respectively.

Topics: Antioxidants; Aryl Hydrocarbon Hydroxylases; Ascorbic Acid; Carcinoma, Hepatocellular; Cell Line, Tumor; Comet Assay; Cytochrome P-450 CYP1A1; Cytochrome P-450 CYP2A6; Cytochrome P-450 CYP2E1; Dimethylnitrosamine; DNA Damage; DNA-Formamidopyrimidine Glycosylase; Dose-Response Relationship, Drug; Humans; Mixed Function Oxygenases; N-Nitrosopyrrolidine; Nitrosamines; Oxidative Stress