| Objective:To optimize the conditions for phenotypic transformation of A549 cells induced by transforming growth factor-?(TGF-?)and to evaluate the main components of Polygonum cuspidatum(polydatin,resveratrol,and emodin)on TGF-?-indued phenotypic transformation of A549 cells.To optimize the conditions for phenotypic transformation of A549 cells induced by platelet-derived growth factor-BB(PDGF-BB)and to evaluate the main components of Polygonum cuspidatum(polydatin,resveratrol,and emodin)on PDGF-BB-indued phenotypic transformation of A549 cells.To optimize the conditions of phenotypic transformation of WI-38 cells induced by TGF-?,and to evaluate the effects of the main components of Polygonum cuspidatum on phenotypic transformation of WI-38 cells induced by TGF-?.By examining the correlation between drugs with anti-lung fibrosis activity and cell phenotype transformation at the cellular level,a reliable protein evaluation target was found,and a drug evaluation screening model based on the phenotypic characteristics of mesenchymal cells was established.To provide new ideas and basis for drug screening and treatment of pulmonary fibrosis diseases at the cellular level.Methods:1.Toxicity of main components of Polygonum cuspidatum on A549 cells:A549 cells in logarithmic growth phase were grown on 96-well plates and treated for 72 h with different concentrations of polydatin,resveratrol,emodin,and paeoniflorin.Cell viability was detected by MTT assay and the cytotoxicity of the main components of Polygonum cuspidatum on A549 cells was evaluated.2.Intervention effect of main components of Polygonum cuspidatum on phenotypic transformation of A549 cells induced by TGF-?:different concentrations of TGF-?acted on logarithmic growth phase A549 cells at different times,with SM a-actin and other mesenchymal cell proteins,to establish an effective mesenchymal transformation model.According to the established phenotypic transformation model of A549 cells,the main components of Polygonum cuspidatum and TGF-?are administered simultaneously,to evaluate intervention effect of the main components of Polygonum cuspidatum on TGF-?-induced phenotypic transformation of A549 cells.3.Intervention of PDGF-BB-induced phenotypic transformation of A549 cells with the main components of Polygonum cuspidatum:TGF-(3,Ang II and PDGF-BB act on logarithmic growth phase A549 cells for 24 h,with SM a-actin and other indicator protein of mesenchymal cells,to establish an effective mesenchymal transformation evaluation basis.According to the established phenotypic transformation model of A549 cells,the main components of Polygonum cuspidatum and PDGF-BB are administered simultaneously,to evaluate intervention effect of the main components of Polygonum cuspidatum on PDGF-BB-induced phenotypic transformation of A549 cells.4.Intervention effect of main components of Polygonum cuspidatum on phenotypic transformation of WI-38 cells induced by TGF-?:TGF-?,Ang ? and PDGF-BB act on logarithmic growth phase WI-38 cells for 24 h,with SM a-actin and other indicator protein of mesenchymal cells,to establish an effective mesenchymal transformation model;based on established mesenchymal transformation models.The main components of Polygonum cuspidatum and TGF-? were administered simultaneously,and the main components of Polygonum cuspidatum was evaluated for TGF-? induced WI-38 Intervention of cell phenotype transformation.Results:1.The cytotoxicity test results of the main components of Polygonum cuspidatum on A549 cells showed that polydatin was administered to A549 cells in a concentration range of 31.25-500 ?M,but it had a certain inhibitory effect but no cytotoxicity;resveratrol administered to A549 cells between the concentration of 4 and 80 ?M,there is a certain inhibition but no cytotoxicity;emodin in the range of 8.75?17.5?M administered to A549 cells,there is a certain inhibition but no cytotoxicity,greater than 17.5 ?M concentration,there are significant cell death and significant cytotoxicity;Paeoniflorin administered to A549 cells in the range of 50-200 ?M,has a certain pro-proliferative effect in a dose-dependent,and has no cytotoxicity but has a certain cell inhibition to A549 cells in the range of 500-1000?M.2.The results of the intervention of TGF-P on the phenotypic transformation of A549 cells induced by the main components of polydatin cuspidatum showed that the expression of E-cadherin,SM a-actin,vimentin and fibronectin was up-regulated after TGF-P induced the phenotypic transformation of A549 cells.According to immunofluorescence and western blot results,A549 cells were induced at 1 ng/mL TGF-? for 24 h,and the phenotype transformation was most obvious.With polydatin(10 ?g/mL?25 ?M),resveratrol(5?g/mL?22 ?M),emodin(1 ?g/mL?3.5 ?M),paeoniflorin(10 ?g/mL?20 ?M)and TGF-?(1 ng/mL)were administered to A549 cells.The results showed that polydatin,resveratrol and paeoniflorin inhibited TGF-?-induced phenotypic transformation of A549 cells.3.The results of the intervention of PDGF-BB-induced phenotype of A549 cells with the main components of polydatin cuspidatum showed that the phenotype transformation of A549 cells induced with TGF-?(1 ng/mL),Ang II(6 nmol/L),and PDGF-BB(60 ng/mL),the protein expression of E-cadherin,SM a-actin,vimentin and fibronectin was up-regulated.According immunofluorescence and western blot results,60 ng/mL PDGF-BB was induced A549 cells for 24 h with the most obvious phenotypic changes.Administration with polydatin(25 pM),resveratrol(20 ?M),emodin(3.5 ?M),paeoniflorin(20 ?M)and PDGF-BB(60 ng/mL)In A549 cells,the results showed that polydatin,resveratrol,emodin,and paeoniflorin inhibited PDGF-BB-induced phenotypic transformation of A549 cells,and emodin had the best inhibitory effect on PDGF-BB-induced phenotypic transformation of A549 cells and concentration-dependent.4.The results of interventions of the main components of polydatin cuspidatum on phenotypic transformation of WI-38 cells induced by TGF-? showed that phenotypic transformation of WI-38 cells induced by TGF-?(1 ng/mL),Ang II(6 nmol/L),and PDGF-BB(60 ng/mL),the expression of E-cadherin,SM a-actin,vimentin,and fibronectin protein was up-regulated.According to immunofluorescence and western blot results,WI-38 cells was induced by 1 ng/mL TGF-P for 24 h with the most obvious phenotypic changes.WI-38 cells was given with polydatin(25 ?M),resveratrol(20 ?M),emodin(3.5 ?M),paeoniflorin(20?M),and TGF-?(1 ng/mL),fluorescence results showed that polydatin,resveratrol and paeoniflorin inhibited the phenotypic transformation of WI-38 cells induced by TGF-?,while emodin hardly inhibited the phenotypic transformation of WI-38 cells induced by TGF-? Western blot results showed that emodin and paeoniflorin inhibited the phenotypic transformation of WI-38 cells induced by TGF-P,and resveratrol and polydatin showed little inhibition.In summary,the main components of polydatin cuspidatum has inhibitory effects on the phenotypic transformation of WI-38 cells induced by TGF-?.Conclusion:The main components of Polygonum cuspidatum can inhibit the phenotypic transformation of A549 cells induced by TGF-?.The main components of Polygonum cuspidatum inhibit the PDGF-BB-induced phenotypic transformation of A549 cells.The phenotype of the main components of Polygonum cuspidatum has not obvious effect intervention on WI-38 cells induced by TGF-?. |
PDF Full Text Request