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Figure 2: REM-induced JNK1/2 activation inhibits MDR1 expression. (a) Cells were treated with 100 μg/mL of either LEM or REM for 15 minutes. Tubulin was detected as an internal control. (b) Cells were treated with the indicatives for 24 hours, and then MDR1 mRNA and protein levels were examined. GAPDH and actin were used as internal controls. (c) To analyze the MDR1 promoter activities, MCF-7/Dox cells were transfected with MDR1-luc construct for 24 hours and then treated with the indicatives for another 6 hours. The luciferase assays were performed in triplicate, and value less than 0.05 (marked with an asterisk, *) was considered statistically significant. (d) MCF-7/Dox cells were treated with the indicatives for 30 hours and then incubated with rhodamine 123 for another 1 hour. Rhodamine 123 accumulation rate was analyzed by flow cytometry. (e) Cells were pretreated with SP600125 for 30 minutes and then treated with 100 μg/mL of REM. 24 hours after treatment, MDR1 mRNA and protein levels were examined. GAPDH and tubulin were used as internal controls. (f) To examine MDR1 promoter activities, MCF-7/Dox cells were transfected with MDR1-luc construct for 24 hours and then treated with REM for another 6 hours. SP600125 was treated 30 minutes before REM treatment. The luciferase assays were performed in triplicate and -value less than 0.05 (marked with an asterisk, *) was considered statistically significant.