Figure 7

LincRNA-p21 enhances TGF-β/Smad signaling in hepatocyte by interacting with miR-30. (A) Effects of miR-30 on KLF11 and Smad7 expressions were determined by western blot and qRT-PCR. Left, AML12 were transfected with miR-30b mimics for 24 h. Right, primary hepatocytes were isolated from fibrotic liver injected with AdH-NC or AdH-miR-30. (B) miR-30 inhibits TGF-β/Smad signaling in hepatocyte. AML12 cells were transfected with miR-30b mimics, changed to serum-free DMEM for 24 h and then treated with TGF-β1 for 2 h. p-Smad2, p-Smad3 and total Smad2, Smad3 were detected by western blot. (C) Ectopic expression of lincRNA-p21 increases KLF11 expression in hepatocyte. AML12 cells were transfected with pCI-lincRNA-p21 for 24 h. KLF11 and Smad7 expressions were determined by western blot and qRT-PCR. (D) Ectopic expression of lincRNA-p21 enhances TGF-β/Smad signaling in hepatocyte. AML12 cells were transfected with pCI-lincRNA-p21 for 24 h and then treated with TGF-β1 for 2 h. (E) Knockdown of lincRNA-p21 reduces KLF11 expression in hepatocyte. AML12 cells were transfected with lincRNA-p21 siRNA for 24 h in the presence of TGF-β1. Top, lincRNA-p21 was determined by qRT-PCR. Bottom, KLF11 was detected by western blot. (F) Knockdown of lincRNA-p21 inhibits TGF-β/Smad signaling. AML12 cells were transfected with lincRNA-p21 siRNA for 24 h and then treated with TGF-β1 for 2 h. (G) Inhibition of miR-30 impairs the effects of lincRNA-p21 siRNA on TGF-β/Smad signaling. AML12 cells were co-transfected with lincRNA-p21 siRNA and miR-30b antagomir for 24 h and then treated with TGF-β1 for 2 h. Left, p-Smad2 and total Smad2 were detected by western blot; Right, miR-30b levels were determined by qRT-PCR. The results are shown as fold change compared with control group. Data are the mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01. The original immunoblot images are shown in Supplementary Figure S10.