Resveratrol Inhibits TWIST1 By Up-regulating BMP7 And Affects Epithelial-to-mesenchymal Transition In Pancreatic Cancer

Objective:To detect the expression of BMP7 between cancer and paracancerous tissues,and explore the relationship between resveratrol,BMP7 and TWIST1.Ultimately prove that resveratrol and BMP7 exhibited inhibitory effect on EMT process in Panc-1 cell.Methods:Immunohistochemistry were used to measure the expression differences of BMP7 in cancer tissue and adjacent normal tissue.Panc-1 cells were treated with Resveratrol(12.5 to 100 μM)at different times.(24h,48 h,72h).The proliferation of Panc-1 cells was determined by MTS test.Panc-1 cells were treated with Resveratrol(50μM)for 48 hours,the expression of BMP7 was measured using RT-PCR.Western blotting measured the expression level of BMP7,Twist1,Akt1 and EMT-related proteins.Lentivirus were used to overexpression and knockdown BMP7 in Panc-1 cell.Through transwell assays and wound healing assay,we observed the effect of EMT process in Panc-1 cells.Results:1.As shown by immunohistochemistry,the expression of BMP7 in in tumor-adjacent normal tissue was higher than that in pancreatic cancer tissue 2.With DMSO as the control group,resveratrol at doses from 12.5 to 100μM significantly decrease the viability of Panc-1 cells,which shows the dose and time dependent of resveratrol for Panc-1 cells.The inhibitory effect on Panc-1 cells was obvious at 50μM for 48 h.3.The results of RT-PCR showed that the expression of BMP7 at the concentration of 50 μ M was significantly higher than that of the control group.Using Western blotting,we found that the ratio of N-cadherin and Vimenten in experimental group were significantly decreased while the ratio of E-cadherin increased(P<0.05).We also found that the expression of Twist1 was suppressed.4.Western blot showed the overexpression of BMP7 significantly decreased the ratio of mmp-2 and mmp-9,compared with LV-control group.We also found that the ratios of Twist-1 was decreased in LV?BMP?7 group.5.In the group of sh-BMP7,the expression level of the N-cadherin and Vimenten and Twist1 were significantly increased with the decrease of E-cadherin compared with sh-control.Meanwhile,there was minor change in the protein expression between sh-BMP7 and sh-BMP7 with resveratrol(50μM).As for comparison,we recorded obvious inhibition of the ratio of Ncadherin and Vimenten and Twist1 between sh-BMP7 and sh-control with resveratrol(50μM).To investigate the relationship between BMP7 and Twist1,we established Co-transfection of BMP7 and Twist1 by overexpression Twist1 plasmid.Lv-BMP7 cells were transfected by Twist1 plasmid.Western blotting was used to confirm the transfection efficiency.We observed that the level of EMT marker protein in Lv-BMP7+ Lv-Twist1 were close to Panc-1 cell,significant differences compared to the Lv-BMP7 group.Conclusion: 1.Low expression of BMP7 in Pancreatic carcinoma 2.BMP7 inhibits the process of EMT in Panc-1 cells.3.Resveratrol could up-regulating BMP7,which inhibits Twist1 and suppress EMT progress in Pancreatic Cancer.