Figure 4
Treatment of NRK-49F cells with trichostatin A or SP600125 (a JNK inhibitor) alleviates the TGF-β1-mediated myofibroblastic induction through modulating the Notch-2 signaling pathway. (a) Effect of TSA on the TGF-β1-mediated activation of Notch-2, α-SMA and fibronection. Western blot analysis was performed using cell lysates of NRK-49F cells that were treated with 5 ng/ml TGF-β1 in combination with TSA at 200 or 500 nM for 48 hr. (b) Effects of MAPK (JNK, p38 or ERK) inhibitors on TGF-β1-mediated myofibroblastic activation. NRK-49F cells were concurrently treated with TGF-β1 and 10 μM SP600125, 10 μM SB203580 or 20 μM PD98059 for 48 hr. Protein expression of phospho-JNK, phospho-p38, phospho-ERK, N2-ICD, α-SMA and fibronectin was determined by Western blot analysis. Results obtained from densitometric analysis are presented as means ± SEM from at least three experiments. #P < 0.05 vs. control group; *P < 0.05 vs. TGF-β1 group. (c) Representative immunofluorescence images of α-SMA (green color) staining in NRK-49F cells. The nuclei were counterstained with DAPI (blue color). Treatment of TGF-β1-treated cells with SP600125 or TSA dose-dependently reduced the formation of α-SMA positive cells. Proportions of α-SMA positive cells are shown in the bottom panel (n = 6). #P < 0.05 vs. control group; *P < 0.05 vs. TGF-β1 group and @P < 0.05 vs. the group treated with TGF-β1 plus SP600125 (5 μM) or TGF-β1 plus TSA (200 nM), respectively.
