Fig. 3. Effect of miR-30c-1-3p on the expression of CYP3A4.

(A) Effect of miR-30c-1-3p on the expression of PXR and CYP3A4 Cells (LS180) were transfected by GenJet version II with the miR-30c-1-3p construct at 20, 200 and 800 ng. The vector plasmid was used to equalize the total amount of constructs. Cells were harvested 72 or 96 h post-transfection. Total RNA was isolated and the mRNA levels of PXR (Left) and CYP3A4 (Right) were determined by RT-qPCR. All experiments were performed three times in triplicate. Asterisk signs in the data indicate statistical significance from vector-transfected cells (P < 0.05). (B) Effect of miR-30c-1-3p on the induction of CYP3A4 Cells (LS180) were transfected as described above and treated with DMSO or 10 µM rifampicin (RIF) 72 h post-transfection. The treated cells were collected 24 h thereafter and the level of CYP3A4 mRNA was determined by RT-qPCR. All experiments were performed three times in triplicate. Asterisk signs in the data indicate statistical significance from vector-transfected cells or cells treated with DMSO (P < 0.05). (C) Effect of miR-30c-1-3p on the activation of CYP3A4-DP-Luc reporter Cells (LS180) were plated in 48 well-plates and transfected with a mixture containing 50 ng of the CYP3A4-DP-Luc reporter, 50 ng of miR-30c-1-3p or the vector along with 5 ng of the Renilla luciferase plasmid. The vector was used to equalize the total amount of constructs. After incubation at 37°C for 24 h, the transfected cells were treated with 10 µM rifampicin (RIF) or the same volume of DMSO for 48 h. Luciferase activities were determined with a Dual-Luciferase Reporter Assay System and the reporter activity was normalized based on the Renilla luminescence signal. All experiments were performed three times in triplicate. Asterisk signs in the data indicate statistical significance from vector-transfected and RIF-treated cells (P < 0.05).