tudy on DEHP treated SHE cells in conditions similar to the present ones. Map Kinases such as Mapk3, Mapk4 and Mapk15 were tar geted by DEHP. Further investigations of Map Kinase pathways could be relevant due to their involvement in activities of transcription factors. The G protein coupled estrogen receptor was found to be over expressed in Differential Display. Gper can be activated by estrogen like compounds and its effect on cytoskeleton architecture has been reported. Because of its implication in the regulation of MAPK or TGF b pathways, it would be worth while to investigate gper further. Performances of DD The confirmation of differentially expressed genes by qPCR showed that the expression levels of more than 75% of genes identified by DD were confirmed by qPCR.

A comparative table of the sensitivity of DD versus qPCR is given in additional file 1. qPCR is more likely to quan tify subtle changes in the expression level of mRNAs at different concentrations while DD seems to be more sen sitive but is less discriminating. To summarize, 35% of the genes GSK-3 identified as differentially expressed in DD gave the same response at the same DEHP concentrations with qPCR while 40% were detected by DD at a lower DEHP concentration than with qPCR. Conclusion Transcriptional responses of SHE cells to DEHP were stu died in conditions inducing the cell neoplastic transforma tion, in order to identify gene expression changes in relation with effects of this non genotoxic carcinogen. Functions impacted by DEHP were found to be PPAR independent.

Effects on cytoskeleton related genes indi cated disturbances on actin polymerization and stabiliza tion, cell cell and cell matrix adhesion and protein trafficking. This is the first study that elucidates the genomic changes of DEHP on the organization of the cytoskele ton. Whether the expression changes of cytoskeleton related genes identified here such as coro1C, nrp2, kif23, are specific to DEHP or to cell transforming agents more generally would require further studies. To answer, the gene sets identified as significantly over or under expressed in this study must be explored on other non genotoxic carcinogens to identify biomarkers predictive of early events in the multistep carcinogenic process. Early disturbances in the expression of cytoskeleton related genes should be considered good candidates.

Methods Chemicals DEHP, purchased from Aldrich Chemicals was dissolved in the DMSO solvent. The latter was obtained from Sigma Aldrich and was used at a final concentration of 0. 1%. Nucleic acid stain Gelred purchased from Interchim was used at a final concentration of 1,10000. All chemicals used for this study were electrophoresis grade or molecular biology grade. Their origin is speci fied in the following sections. SHE cell culture and treatment SHE cells were isolated from Syrian hamster embryos at day 13 of gestation using the procedure described by Pienta et al. and in accordance with the modifica tions sugge