Figure 1
Design and validation of a high-throughput screening system based on the transcriptional activities of TCF/LEF, NF-κB and NRF2. (A) Workflow for the establishment of each cell lines for the screening system. The respective inducers or inhibitors were added for 24 hours to validate the sensitivity of the luciferase assay in each reporter cell lines of HCT116 (B) and DLD-1 (C). Luciferase activity was measured after stimulation with 0, 1.25, 2.5, 5, 10, 20 and 40 ng/ml TNF-α in NF-κB-reporter cell lines. Luciferase activity was measured after stimulation with 0, 1.25, 2.5, 5, 10, 20 and 40 μM tBHQ in NRF2-reporter cell lines. TCF/LEF-reporter cell lines were stimulated by Wnt3a CM and 5 and 10 μM iCRT14 in HCT116 cells or 20 and 40 μM iCRT14 in DLD-1 cells. Wnt3a CM was added at 10% of total volume. The data are shown as the relative value, with the value of the vehicle-treated cells set as 1 in each cell line. The data are the mean ± SD, n = 3. *p < 0.03, **p < 0.003 vs. control (vehicle-treated cells). CM, conditioned medium.
