Effect of plumbagin on epithelial-mesenchymal transition and underlying mechanisms in human tongue squamous cell carcinoma cells
Author(s): Li Bin, Pan Shuting, Qiu Jiaxuan
Pages: 421-426
Year: 2017 Issue: 7
Journal: Chinese Journal of Stomatology
Keyword: Carcinoma; squamous cell; Tongue; Plumbagin; p38 mitogen-activated protein kinases; Reactive oxygen species;
Abstract: Objective To study the effect of plumbagin on epithelial-mesenchymal transition (EMT) and underlying mechanisms in human tongue squamous cell carcinoma (TSCC) cells.Methods Methyl thiazolyl tetrazolium assay was apllied to examine the proliferation inhibition effect and half maximal inhibitory concentration (IC50) of plurnbagin (0.1,1.0,5.0,10.0,20.0 μmol/L) in 12,24,48 h in TSCC cells.Transwell assay was used to count the number of transmembrane cells and scratch test was performed to examine cells mobility.The flow cytometry was applied to measure intracellular reactive oxygen species (ROS) level in control group,plumbagin group (1.0 μmol/L,24 h) and glutathione (GSH)+plumbagin group.The expression of E-cadherin,vimentin,Slug,p38 mitogen activated protein kinases (p38MAPK) and phospho-p38MAPK (p-p38MAPK) proteins were determined by Western blotting.The expression of E-cadherin,vimentin and Slug were detected by Western blotting in control group,plumbagin group,activator combined group (p38MAPK activator + plumbagin) and inhibitor combined group (p38MAPK inhibitor+plumbagin).Results After the treatment of plumbagin for 12,24,and 48 h,the IC50of TSCC cells were 10.3,3.1,1.5 μ mol/L.After treated by 1.0 μ mol/L plumbagin for 24 h,the number of transmembrane cells were significantly reduced ([50± 13],P＜0.05) in comparison to control group (204±6),as well as the cells mobility ([18.2± 2.3]％,P＜0.05) in comparison to control group ([49.3± 1.2]％).Compared to control group (2.32±0.52),the ROS level was increased in plumbagin group (902.20± 10.69),while compared to plumbagin group,the ROS level was reduced in GSH combined group (2.18±0.15).In comparison to control group,the expression of E-cadherin was up-regulated (P＜0.05),and vimentin,Slug,p-p38MAPK/p38MAPK were down-regulated in plumbagin group (P＜0.05).In comparison to plumbagin group,the expression of E-eadherin was down-regulated (P＜0.05),and vimentin,Slug,p-p38MAPK/p38MAPK were up-regulated in GSH combined group (P＜0.05).Treatment of cells with p38MAPK activator could decrease the expression of E-cadherin significantly (P＜0.05) and increase the expression of vimentin (P＜0.05) and Slug (P＜0.05) in comparison to plumbagin group.Treatment of cells with p38MAPK inhibitor could increase the expression of E-cadherin significantly (P＜0.05) and decrease the expression of vimentin (P＜0.05) and Slug (P＜0.05) in comparison to plumbagin group.Conclusions Plumbagin inhibits EMT via ROS/p38MAPK-mediated pathway in human TSCC cells.
Pages: 421-426
Year: 2017 Issue: 7
Journal: Chinese Journal of Stomatology
Keyword: Carcinoma; squamous cell; Tongue; Plumbagin; p38 mitogen-activated protein kinases; Reactive oxygen species;
Abstract: Objective To study the effect of plumbagin on epithelial-mesenchymal transition (EMT) and underlying mechanisms in human tongue squamous cell carcinoma (TSCC) cells.Methods Methyl thiazolyl tetrazolium assay was apllied to examine the proliferation inhibition effect and half maximal inhibitory concentration (IC50) of plurnbagin (0.1,1.0,5.0,10.0,20.0 μmol/L) in 12,24,48 h in TSCC cells.Transwell assay was used to count the number of transmembrane cells and scratch test was performed to examine cells mobility.The flow cytometry was applied to measure intracellular reactive oxygen species (ROS) level in control group,plumbagin group (1.0 μmol/L,24 h) and glutathione (GSH)+plumbagin group.The expression of E-cadherin,vimentin,Slug,p38 mitogen activated protein kinases (p38MAPK) and phospho-p38MAPK (p-p38MAPK) proteins were determined by Western blotting.The expression of E-cadherin,vimentin and Slug were detected by Western blotting in control group,plumbagin group,activator combined group (p38MAPK activator + plumbagin) and inhibitor combined group (p38MAPK inhibitor+plumbagin).Results After the treatment of plumbagin for 12,24,and 48 h,the IC50of TSCC cells were 10.3,3.1,1.5 μ mol/L.After treated by 1.0 μ mol/L plumbagin for 24 h,the number of transmembrane cells were significantly reduced ([50± 13],P＜0.05) in comparison to control group (204±6),as well as the cells mobility ([18.2± 2.3]％,P＜0.05) in comparison to control group ([49.3± 1.2]％).Compared to control group (2.32±0.52),the ROS level was increased in plumbagin group (902.20± 10.69),while compared to plumbagin group,the ROS level was reduced in GSH combined group (2.18±0.15).In comparison to control group,the expression of E-cadherin was up-regulated (P＜0.05),and vimentin,Slug,p-p38MAPK/p38MAPK were down-regulated in plumbagin group (P＜0.05).In comparison to plumbagin group,the expression of E-eadherin was down-regulated (P＜0.05),and vimentin,Slug,p-p38MAPK/p38MAPK were up-regulated in GSH combined group (P＜0.05).Treatment of cells with p38MAPK activator could decrease the expression of E-cadherin significantly (P＜0.05) and increase the expression of vimentin (P＜0.05) and Slug (P＜0.05) in comparison to plumbagin group.Treatment of cells with p38MAPK inhibitor could increase the expression of E-cadherin significantly (P＜0.05) and decrease the expression of vimentin (P＜0.05) and Slug (P＜0.05) in comparison to plumbagin group.Conclusions Plumbagin inhibits EMT via ROS/p38MAPK-mediated pathway in human TSCC cells.
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