Figure 3
An activated EMT pathway is required for chemoresistance to MTA. (a) TGF-β induces a morphological switch from epithelial-to-mesenchymal phenotype and kaemperol reverses the process. H358, untreated H358 cells; H358_EMT, H358 cells treated with TGF-β (5 ng/ml) for 14 days; H358_EMT+Kae, H358_EMT cells treated with kaempferol (20 μM) for 5 days. Scale bar, 100 μm. (b) TGF-β treatment activates the EMT pathway whereas kaempferol inhibits the process. The mRNA level of EMT-TFs and N-cadherin in H358, H358_EMT and kaempferol-treated H358_EMT cells (H358_EMT+kaempferol; as above) were analyzed by qPCR. Data are shown as mean±S.D. of three independent experiments (n=3). (c) Chemoresistance to MTA is promoted by an activated EMT pathway and regressed by kaempferol. H358, H358_EMT and kaempferol-treated H358_EMT cells (H358_EMT+kaempferol) were treated with MTA for 5 days and analyzed by XTT assay. Data are shown as mean±S.D. of three independent experiments (n=3). (d) Micrographic images of chemoresistant H358_R cells treated with vehicle (H358_R) or with 20 μM kaempferol for 3 days (H358_R+kaempferol). Scale bar, 50 μm (e and f) Kaempferol treatment inhibits the expression of EMT-TFs and stem cell genes. The mRNA level of EMT-TFs and N-cadherin (e) and of stem cell genes (f) in vehicle- and kaempferol-treated H358_R cells (as above) was analyzed by qPCR. Data are shown as mean±S.D. of three independent experiments (n=3). (g) Kaempferol and SB431542 abrogate chemoresistance to MTA. H358_R cells treated with vehicle, kaempferol (20 μM) and SB431542 (40 μM) for 3 days were then exposed to MTA for another 5 days. Drug response was determined by XTT assay and data are shown as mean±S.D. of three independent experiments (n=3). *P<0.05; **P<0.01; ****P<0.001
