Figure 1
Anti-fibrotic effects of HSM ethanol extract on the production of myofibroblast marker proteins α-SMA and fibronectin through suppression of Smad2/3 and Akt signaling pathways in MRC-5 human lung fibroblasts.
Cells were treated with various concentrations (1, 2 and 5%) of HSM extract for 24 h (a) or 48 h (b) and cell viability was measured using the MTT assay. (c,d) Cells were serum-starved overnight, before treatment with varying concentrations (1 and 2%) of HSM extract for 24 h and incubation with TGF-β1 (5 ng/ml) for 24 h. Expression of the myofibroblast markers α-SMA and fibronectin were evaluated in whole cell lysates by Western blotting. β-actin was used as a loading control. Relative protein levels were quantified by scanning densitometry and were normalized to β-actin. (e,f) Cells were pretreated with HSM extract (1 and 2%), followed by treatment with TGF-β1 (5 ng/ml) for another 6 h. Phosphorylated and total Smad2/3 and Akt were measured by Western blotting against phospho-Smad2/3, Smad2/3, phospho-Akt and Akt. Blots were analyzed by densitometry and the results were expressed as relative units. Data shown represent means ± SEM of three experiments performed in duplicate. #P < 0.05 versus untreated or ethanol–treated cells. *P < 0.05 versus control (TGF-β1–treated) cells.
