Enhanced 4-Hydroxynonenal Resistance in KEAP1 Silenced Human Colon Cancer Cells
Figure 4
Effect of KEAP1 knockdown on NRF2 activity in HCT116 cells. (a) The mRNA level for KEAP1 was determined in HCT116 cells expressing scRNA (sc) and KEAP1-specific shRNA (shKEAP1). (b) ARE-driven luciferase activity was monitored in HCT116, sc, and shKEAP1 HCT116 cells. (c) The nuclear level for NRF2 protein was determined in HCT116, sc, and shKEAP1 HCT116 cells. Lamin B levels were used as a loading control. (d) Cellular total GSH contents were measured in HCT116, sc, and shKEAP1 HCT116 cells. (e) The basal mRNA levels for AKRs (AKR1C1/2, 1C2, 1C3, 1B1, and 1B10) were determined in HCT116, sc, and shKEAP1 HCT116 cells. Values are relative levels with respect to sc group and are the means ± SD of 3-4 experiments. (f) Cell viabilities were measured using MTT analysis following the incubation with H2O2 (350–650 μM) for 24 h. (g) Cell viabilities were measured following the incubation of cells with menadione (5–15 μM) for 24 h. The values are means ± SD from 8 wells. compared with the sc control.
