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Ref. BSCHGR58/08(UCM), Ref. No. S2009/AGR-1469 (CAM) andConsolider-Ingenio 2010 No.CSD2007-063 (MEC), Spain.

doi:10.1016/j.toxlet.2012.03.506

138 Abstracts / Toxicology L

20-22se of in vitro assays for the prediction of the skin sensitizingotential of chemicals

ierre-Jacques Ferret, Marie-Pierre Gomez-Berrada, Carole Berge

Pierre Fabre Dermo Cosmétique, France

Purpose: Cutaneous sensitization results from the induction ofn immune response following the exposure of the skin to a sub-tance. Currently, the only validated alternative method remainshe Murine Local Lymph Node Assay (LLNA). Progress in the com-rehension of cutaneous sensitizing mechanisms and the need foronforming to the current legislation requirements supported theevelopment of new alternative methods to the animal experimen-ation. These methods will have to summarize the whole complexnteractions of the allergen with the immune system and need toe exploitable for any type of compound. Methods: The results pre-ented here were achieved with 40 substances of known sensitizingotential (25 cosmetic raw materials and 15 control substances)sing various methods: (1) the direct peptide reactive assay (DPRA),2) a dentritic cell activation assay (h-CLAT) and (3) the ARE-ependent gene expression in SENS-IS. The results are expressed inclasses (Slightly; Moderately; Strongly and Very strongly sensitiz-

ng) allowing to compare them. The correlation of individual testsas obtained by comparison to human Repeated Insult Patch Test

hRIPT) and LLNA data. Results and conclusion: We have observedgood correlation between the results obtained with DPRA and

n vitro models (75–90%) and a good correlation with those result-ng from the LLNA and hRIPT (75–85%). It appears obvious thatnly the combination of several alternative methods belonging tostrategy of evaluation could replace the LLNA. Moreover it woulde interesting to supplement these data by realization of studieselating to the Nrf2/Keap1 pathway.

oi:10.1016/j.toxlet.2012.03.503

20-23uman dendritic cell based model (DC-100) for

mmunotoxicity studies

ilvia Letasiova 1, Amy Hunter 2, Maureen Spratt 2, Alexrmento 2, Mitchell Klausner 2, Seyoum Ayehunie 2

MatTek IVLSL, Slovakia, 2 MatTek Corporation, Ashland, MA, Unitedtates

The immune system can be the target of a broad variety ofhemicals in the form of environmental contaminants, medica-ents, fabrics, pesticides, food additives and preservatives, and

ay-to-day household products with adverse effects to humanealth. Immunotoxicants can induce suppressive or stimulatoryesponses and DNA damage in components of the immune sys-em. In this study, dendritic cells (DC) were used to monitormmunotoxicant induced phenotypic changes (surface markerxpression), functional alteration (cytokine release), and DNA dam-ge (comet assay). We evaluated the effect of 5 immunotoxicompounds (ITC) and 2 reproductive hormones, estradiol and pro-esterone, on DC responses. Finally, we checked DNA damagef DC following treatment with the carcinogenic agent, methylethanesulfonate (MMS).DC were exposed to non-cytotoxic con-

entrations of the ITC or hormones for 24 h and then to LPS

or an additional 18 h. FACS analysis of ITC-exposed DC showedffects in the rank order of immunotoxicity (severe to low effect):ributyltin chloride > Cyclosporine A > Benzo(a)pyrene > Fursemide

211S (2012) S43–S216

and Urethane. Dose-dependent decreases in the secretion ofimmuno-stimulatory cytokines including interleukin (IL)-12, IL-6, and IL-1� were observed. Furthermore, treatment of DC withthe reproductive hormones, estradiol and progesterone, showeda decrease in lipopolysaccharide-induced CCR7 expression andIL-12 secretion. Exposure of DC to MMS also showed dose depen-dent DNA damage. Conclusion: The release of immunostimulatorycytokines and DNA damage may serve as endpoints to predictimmunotoxicity. Due to concerns with animal models in terms ofcost, ethical issues, and relevance to hazard assessment in humans,the human primary DC based assay is an attractive in vitro modelto predict the immunotoxicity of compounds and formulations.

doi:10.1016/j.toxlet.2012.03.504

P21: Metabolism and Kinetics

P21-01Inhibition of hepatic mixed function oxidase system bynatamycin

Arturo Anadon, Maria A. Martinez, Victor Castellano, MartaMartinez, alejandro Romero, Irma Ares, Eva Ramos, Maria R.Martinez-Larranaga

Universidad Complutense de Madrid, Spain

Natamycin (pimaricin), a polyene macrolide antibiotic, has beenused as a topical antimycotic agent in animals and humans for over40 years and it is also used as an antifungal agent in food processing.Natamycin appears to be an extremely effective inhibitor of mouldgrowth and mycotoxin production. The low solubility of natamycinmakes it suitable for use as a surface treatment on foods becausedoes not readily migrate into the interior and does not adverselyaffect flavour or appearance. For risk assessment, it is of primeinterest to obtain relevant information on the natamycin oxidativemetabolism.

The objective of this study is to establish if natamycin interactswith hepatic microsomal metabolising enzymes. Animals (maleWistar rats) were treated with natamycin (0.8, 2 and 8 mg/kg/dayi.p. 4 days). Natamycin-treated and control animals were sacri-ficed 24 h after the last administration and livers were removed.The livers were individually homogenized and microsomal pelletsprepared and stored at −80 ◦C for enzyme assays.

Natamycin decreased significantly the content of cytochromeP450, and the activities of aniline hydroxylase, erythromycin N-demethylase (CYP3A1), ethoxyresorufin O-deethylase (CYP1A1)and pentoxyresorufin O-depenthylase (CYP2B1). Natamycin alsoproduced a significant decrease in the hydroxylation of lauric acidassociated with the CYP4A subfamily. These data suggest reduc-tion in the formation of toxic products of oxidative metabolism,but natamycin as inhibitor of a number of cytochrome P450 medi-ated metabolic pathways, might cause clinically pharmacokineticdrug interactions.

Acknowledgements: This work was supported by projects