Nuria Arranz

Protective effects of isothiocyanates towards N-nitrosamine-induced DNA damage in the single-cell gel electrophoresis (SCGE)/HepG2 assay.

The aim of this study was to investigate the protective effect of isothiocyanates towards N‐nitrosamine‐induced DNA damage in the single‐cell gel electrophoresis (SCGE)/HepG2 assay. None of the isothiocyanates (ITCs) concentrations tested in the presence or absence of formamidopyrimidine‐DNA glycosylase (Fpg) caused DNA damage per se. Combined treatments of HepG2 cells with phenethyl isothiocyanate (PEITC), allyl isothiocyanate (AITC) or indol‐3‐carbinol (I3C) and N‐nitrosopyrrolidine (NPYR) or N‐nitrosodimethylamine (NDMA) reduced the genotoxic effects of the N‐nitrosamines in a dose‐dependent manner. The protective effect of the three ITCs tested was higher towards NPYR‐induced oxidative DNA damage than against NDMA. The greatest protective effect towards NPYR‐induced oxidative DNA damage was shown by I3C (1 µM, 79%) and by PEITC (1 µM, 67%) and I3C (1 µM, 61%) towards NDMA (in presence of Fpg enzyme). However, in absence of Fpg enzyme, AITC (1 µM, 72%) exerted the most drastic reduction towards NPYR‐induced oxidative DNA damage, and PEITC (1 µM, 55%) towards NDMA. Our results indicate that ITCs protect human‐derived cells against the DNA damaging effect of NPYR and NDMA, two carcinogenic compounds that occur in the environment. Copyright

Organosulfur compounds alone or in combination with vitamin C protect towards N-nitrosopiperidine- and N-nitrosodibutylamine-induced oxidative DNA damage in HepG2 cells.

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.

Inhibition by vitamin C of apoptosis induced by N-nitrosamines in HepG2 and HL-60 cells

The aim of this study was to evaluate the effect of vitamin C towards N‐nitrosopyrrolidine (NPYR)‐ and N‐nitrosodimethylamine (NDMA)‐induced apoptosis in human hepatoma (HepG2) and leukemia (HL‐60) cell lines using flow cytometry analysis and the terminal deoxynucleotidyl transferase‐mediated dUTP nick end labelling assay (TUNEL). None of the vitamin C concentrations tested (1–100 µm) caused cytotoxicity in HepG2 cells. However, there were significant losses of HL‐60 cells viability, measured by MTT assay, 72 h after treatment with 50 and 100 µm vitamin C (29 and 46%, respectively). Moreover, an increase of lactate dehydrogenase release was significant with 50 µm at 72 h (28%) and with 100 µm of vitamin C at 48 and 72 h (27 and 36%, respectively). Also, the percentage of apoptotic HL‐60 cells found in TUNEL assay increased to 21% when they were treated with 100 µm vitamin C for 72 h. Thus, in subsequent simultaneous treatments with NPYR (30 and 50 mm) or NDMA (27 and 68 mm) and vitamin C, concentrations of 5–50 µm vitamin C were used. Our results revealed that vitamin C, at all concentrations and times tested, reduced the apoptosis induced by NPYR and NDMA in both cell lines, showing a similar effect in HepG2 and HL‐60 cells towards NPYR (50 mm) — 65 and 63% of reduction, respectively — whereas towards NDMA (27 mm) the inhibition was higher in HL‐60 than in HepG2 cells — 75 and 57%, respectively. Therefore, our findings suggest that inhibition of apoptosis may be one of the mechanisms by which vitamin C exerts its protective effect. Copyright

Protective effect of a "Lactobacillus salivarius" strain of human origin

The protective effect of a Lactobacillus salivariusstrain from human faeces (HA8) was evaluated against the cytotoxicity of N-nitrosodimethylamine (NDMA), N-nitrosopyrrolidine (NPYR), N-nitrosodibutylamine (NDBA) and N-nitrosopiperidine (NPIP) by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. L. salivariusHA8 strain showed a moderate protective effect against NPYR and weak against NDBA and NPIP. No protective effect against cytotoxicity of NDMA was observed at the bacterial population used. To test the effect of L. salivariusHA8 on cytokine production (interleukin-1 ± (IL-1 ±), interleukin-8 (IL-8) and tumour necrosis factor alpha (TNF-≥)), the human macrophage cell line (THP-1) was cultured in the presence and absence of lipopolysaccharide (LPS). L. salivariusHA8 induced IL-1 ±, IL-8 and TNF-≥ releases when cells were stimulating with and without LPS.