Comparison of Ah receptor-mediated luciferase and ethoxyresorufin O-deethylase expression in H411E cells: Their use as bioassay tools
Conference
·
OSTI ID:203627
- Michigan State Univ., East Lansing, MI (United States)
- Univ. of Wageningen (Netherlands)
The response of a recombinant rat hepatoma cell line (H411E-luc), containing a luciferase expression vector under control of dioxin-responsive enhancers, was characterized to several polychlorinated aromatic hydrocarbons (PHAHs). The new cell system was compared with the widely used wild-type cell line (H411E-wt), which expresses Ah receptor-mediated cytochrome P-450 1A1 induction. The authors also report an improved method for the H411E-wt bioassay which allows for the rapid screening of environmental samples for Ah-active PHAHs. This method makes use of 96-well plates, a plate-reading spectrofluorometer and a fluorescence-based protein assay that enables the simultaneous measurement of resorufin and protein. Both cell lines demonstrated a dose-dependent increase in Ah receptor-mediated response upon exposure to several known Ah agonists. H411E-luc cells were 3-fold more sensitive than H411E-wt cells to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The detection limit and ED{sub 50} of TCDD for EROD induction in H411E-wt cells were 0.6 and 4.9 fmole/well, respectively; for luciferase induction in H411E-luc cells 0.2 and 1.4 fmole/well. The detection limit in H411E-wt cells was a 50-fold improvement over that reported previously and comparable to that of a chicken embryo hepatocyte bioassay developed by Kennedy et al. The tested PHAHs exhibited a similar structure-activity relationship in H411E-luc as in H411E-wt cells. Binary mixtures of TCDD, PCB-126 and PCB-77, dosed at various ratios, showed no departure from additivity, while PCB-153 at a concentration of 14 nmole/well reduced the potency, without affecting the efficacy, of TCDD in both cell lines. This study supports the use of such luciferase-transfected cell lines as bioanalytical tools for screening Ah-active components.
- OSTI ID:
- 203627
- Report Number(s):
- CONF-9511137--; ISBN 1-880611-03-1
- Country of Publication:
- United States
- Language:
- English
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