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Minh SD, Below S, Muller C, Hildebrandt JP: Novel mammalian cell lines expressing reporter genes for the detection of environmental chemicals activating endogenous aryl hydrocarbon receptors (ArhR) or estrogen receptors (ER). Toxicol In Vitro. 2008 Dec;22(8):1935-47. Epub 2008 Sep 15.
We have constructed two vector systems (pDMS5, pSAB2) containing the promoter regions of the human CYP1A1 gene including xenobiotic response elements or the promoter region of the Xenopus laevis vitellogenin A2 gene including estrogen response elements, respectively, and the genes for green fluorescent protein and firefly luciferase. These vectors were transfected into CHO-K1 cells. Transiently transfected cells consistently responded to 1 nmol/l TCDD (dioxin) or 10 nmol/l 17ss-estradiol, respectively, with a 3-5 fold increase in luciferase activity. Permanent cell lines were selected by culturing transiently transfected cells under continued presence of antibiotics and dilution cloning. Cells which had stably integrated the vector-DNA into the genomic DNA were selected. SiF6 cells responded to treatment with TCDD, PCB126, benzo (a) pyrene or indirubin-3'-monoxime in the concentration range between 0 and 1 micromol/l. SiG12 cells responded to treatment with bisphenol A, 4-MBC and DDT in the concentration range between 0 and 10 micromol/l. Compared with the controls, luciferase mRNA-abundance (semi-quantitative RT-PCR) and luciferase activity (luminescence assay) were elevated up to 3-fold. Resveratrol or tamoxifen, respectively, worked as full antagonists. Luciferase expression was increased upon treatment of cells with extracts of spiked soil samples indicating that our systems are suited for screening of environmental samples. |
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