Figure 8. Inhibition of Smad4 by siRNA or hsa-miR-4535 mimic reverses the galangin-mediated enhancement of collagen synthesis in dermal fibroblasts. (A–C) HS68 cells were transfected with hsa-miR-4535 mimic (10 nM) or mimic NC (10 nM) for 1 h followed by cotreatment with galangin (30 μM) and H2O2 (200 μM) for 23 h. Hsa-miR-4535 (A) and Smad4 (B) levels were detected by qPCR to ensure successful transfection. (D–E) HS68 cells were transfected with siRNA Smad4 (20 nM) or siRNA NC (20 nM) for 1 h followed by cotreatment with galangin (30 μM) and H2O2 (200 μM) for 23 h. Smad4 levels were detected by qPCR to ensure successful transfection (D). Protein levels of Smad4, COL1A1 and COL3A1 were analyzed by western blotting (C, E). GAPDH was used as a loading control. Values shown are means ± SE. Quantification of the results is shown (n = 3) *P < 0.05, **P < 0.01, ***P < 0.001 vs. untreated control cells; #P < 0.05, ##P < 0.01, vs. H2O2-treated cells; $$P < 0.01, $$$P < 0.001 vs. H2O2 plus galangin-treated cells.