Fig. 2

miR-9-5p over-expression abrogates TGF-β1-induced differentiation of human dermal fibroblasts into myofibroblasts. a–c qRT-PCR analysis for mRNA expression of α-SMA (a), Col1α1 (b), and FN (c) in HDFs transfected with 40-nM pre-miR-9-5p or pre-miR-NC and treated with TGF-β1 (5 ng/ml) for the indicated times. d Western blot analysis (top) and quantification (bottom) of α-SMA levels in HDFs transfected with 40-nM pre-miR-9-5p or pre-miR-NC and treated with TGF-β1 (5 ng/ml) for the indicated times (a.u., arbitrary units). The bar graph shows values after correction by GAPDH expression and normalized to control conditions. e Western blot analysis (top) and quantification (bottom) of FN levels in HDFs transfected with 40-nM pre-miR-9-5p or pre-miR-NC and treated with TGF-β1 (5 ng/ml) for the indicated times (a.u., arbitrary units). The bar graph shows values after correction by GAPDH expression and normalized to control conditions. f Sircol assay of secreted collagen proteins I-V was done in HDFs transfected with 40-nM pre-miR-9-5p or pre-miR-NC and treated with TGF-β1 (5 ng/ml) for the indicated times (a.u., arbitrary units). g Immunofluorescence staining of α-SMA (green) in HDFs treated with TGF-β1 (5 ng/ml) for 48 h after transfection with 40-nM pre-miR-9-5p or pre-miR-NC. Nuclei were stained with DAPI (blue). Scale bars: 100 μm. The bar graphs show mean ± SEM of three independent experiments, *P < 0.05, **P < 0.01, and ***P < 0.001 compared to control cells and ^# P < 0.05 and ^## P < 0.01 compared to its corresponding negative control time point