Figure 4.

The impact of Icariin on Smad and MAPK signaling pathways in 16HBE cells. 16HBE cells were pretreated with or without Icariin (low, medium and high dose) for 6 h and then treated with or without 10 ng/ml TGF-β1 for 24 h. A. Western blot was used to detect the phosphorylation of Smad-2 and Smad-3 and total level of Smad-2/3 and Smad-4 proteins of Smad signaling pathway. B. The phosphorylation of Erk, Jnk and p38 and total level of Erk, Jnk and p38 proteins of MAPK signaling pathway were detected by western blot. C. The expressions of Smad-2, Smad-3 and Smad-4 mRNA were determined by quantitative real-time PCR. D. The expressions of Erk, Jnk and p38 mRNA were determined by quantitative real-time PCR. E. The inhibitory impact of Icariin on Smad2/3/4 complex formation in TGF-β1-induced 16HBE was detected by co-immunoprecipitation. F. The protein and mRNA of Snail were detected by western blot and quantitative real-time PCR. G. 16HBE were transfected with Pgl3-Basic-Snail-luc reporter plasmid, luminescence was measured by a luminometer. pRL-TK plasmids served as the correcting transfection efficiency. Results were expressed as the ratios between the activity of the reporter plasmid and Prl-TK. #P<0.05, ##P<0.01 versus control group, *P<0.05, **P<0.01 versus TGF-β1 group.