Claudine Bezençon

Ochratoxin A-mediated DNA and protein damage: roles of nitrosative and oxidative stresses.

Ochratoxin A (OTA) is a mycotoxin occurring in a variety of foods. OTA is nephrotoxic and nephrocarcinogenic in rodents. An OTA-mediated increase of the inducible nitric oxide synthase (iNOS) expression was observed in normal rat kidney renal cell line and in rat hepatocyte cultures, suggesting the induction of nitrosative stress. This was associated with an increased nuclear factor kappa-light chain enhancer of activated B cells activity. The potential consequences of iNOS induction were further investigated. A significant increase in the levels of protein nitrotyrosine residues was observed with OTA. In addition, OTA was found to increase the level of DNA abasic sites in both cell cultures system. This end point was used as an indirect measure of 8-nitroguanine formation. Treatment of the cells with L-N(6)-(1-iminoethyl) lysine, a specific inhibitor of iNOS activity, inhibited the OTA-mediated overnitration of proteins but did not reduce the level of DNA abasic sites. It was found previously that nuclear factor-erythroid 2 p45-related factor 2 (Nrf2) activators were able to restore the cellular defense against oxidative stress and could prevent DNA abasic sites in cell cultures. In the present study, pretreatment of the cells with activators of Nrf2 prevented OTA-mediated increase in lipid peroxidation, confirming the potential of Nrf2 activators to confer protection against OTA-mediated oxidative stress. In addition, it was found that Nrf2 activators could also prevent OTA-induced protein nitration and cytotoxicity. In conclusion, the present data further confirm oxidative stress as a key source of OTA-induced DNA damage and provide additional evidence for a role of this mechanism in OTA carcinogenicity. The exact role of nitrosative stress still remains to be established.

Coffee diterpenes prevent benzo[a]pyrene genotoxicity in rat and human culture systems.

The coffee-specific diterpenes cafestol and kahweol (C+K) have been identified as two important chemoprotective agents in coffee. In the present study, the potential preventive effects of C+K against the genotoxicity of B[a]P were investigated in rat primary hepatocytes and in human bronchial Beas-2B cells. Several independent mechanisms were identified and their respective contribution to the overall protective effects was determined. A marked dose-dependent inhibition by C+K of B[a]P DNA-binding was found in cells of both origins. However, data showed that the significant induction by C+K of the detoxifying enzyme GST-Yp subunit is the key mechanism of protection against B[a]P DNA-binding in rat liver. In contrast, the phase I-mediated mechanism where C+K produce an inhibition of CYP 1A1 induction by B[a]P is of key significance for the C+K protection in human Beas-2B cells. Moreover, this effect suggests a novel mechanism of chemoprotection by the coffee diterpenes cafestol and kahweol.

Integrating bioassays and analytical chemistry as an improved approach to support safety assessment of food contact materials

ABSTRACT Food contact materials (FCM) contain chemicals which can migrate into food and result in human exposure. Although it is mandatory to ensure that migration does not endanger human health, there is still no consensus on how to pragmatically assess the safety of FCM since traditional approaches would require extensive toxicological and analytical testing which are expensive and time consuming. Recently, the combination of bioassays, analytical chemistry and risk assessment has been promoted as a new paradigm to identify toxicologically relevant molecules and address safety issues. However, there has been debate on the actual value of bioassays in that framework. In the present work, a FCM anticipated to release the endocrine active chemical 4-nonyphenol (4NP) was used as a model. In a migration study, the leaching of 4NP was confirmed by LC-MS/MS and GC-MS. This was correlated with an increase in both estrogenic and anti-androgenic activities as measured with bioassays. A standard risk assessment indicated that according to the food intake scenario applied, the level of 4NP measured was lower, close or slightly above the acceptable daily intake. Altogether these results show that bioassays could reveal the presence of an endocrine active chemical in a real-case FCM migration study. The levels reported were relevant for safety assessment. In addition, this work also highlighted that bioactivity measured in migrate does not necessarily represent a safety issue. In conclusion, together with analytics, bioassays contribute to identify toxicologically relevant molecules leaching from FCM and enable improved safety assessment.