Effect of TKT on proliferation,migration and invasion of colorectal cancer cells
Objective To investigate the expression of transketolase(TKT)in colorectal cancer and its effect on the proliferation and migration and invasion ability of colorectal cancer cells.Methods The expression of TKT in colorectal cancer tissues and adjacent normal tissues was analyzed by quantitative PCR and an interactive analysis platform based on gene expression level values.Two plasmids(Ctrl plasmid and TKT overexpression plasmid)were used to transfect two colorectal cancer cell lines of SW480 and HCT116,respectively,in order to construct TKT-overexpressing stably-transfected cells(Ctrl group and TKT-Flag group).Meanwhile,two RNAs(TKT-CtrlRNA and TKT-shRNA)were used to transfect the above two colorectal cancer cells by recombinant lentiviral technique,in order to construct TKT knockdown stably-transfected cells(NC group and shTKT group).Then,the effect of TKT overexpression and TKT knockdown was detected by Western blot,and the effect of TKT on the proliferation of colorectal cancer cells was analyzed by CCK-8 assay,real-time label-free cellular analysis(RTCA),and cell clone formation assay.The HCT116 stably-transfected cells overexpressing TKT were subcutaneously injected into nude mice.After six weeks,the mice were sacrificed,and subcutaneous tumor tissues were taken.The effect of TKT on the tumor-forming ability of colorectal cancer cells was detected by measuring the tumor volume and mass.Transwell assay and wound healing assay were used to analyze the effect of TKT overexpression and TKT knockdown on the migratory and invasive abilities of the above two types of colorectal cancer cells.HCT116 stably-transfected cells overexpressing TKT and knockdown TKT were injected via the tail vein to establish a lung metastasis model.After eight weeks,the effect of TKT on the metastasis of colorectal cancer cells was further detected by in vivo imaging and H-E staining.The effect of TKT overexpression and TKT knockdown on epithelial-mesenchymal transition(EMT)associated proteins in colorectal cancer cells was detected by Western blot.Results Compared with adjacent normal tissues,TKT expression was significantly up-regulated in colorectal cancer tissues,and TKT gene expression also increased in colorectal cancer.Compared with the Ctrl group,TKT protein expression was up-regulated in the TKT-Flag group,the proliferation,migration,and invasion of colorectal cancer cells were elevated,the levels of the EMT-related proteins Vimentin,N-cadherin,and Snail increased,and the levels of E-cadherin protein decreased.Compared with the NC group,shTKT protein expression was down-regulated in the shTKT group,the proliferation,migration and invasion of colorectal cancer cells significantly decreased,the levels of EMT-related proteins Vimentin,N-cadherin and Snail decreased,and the levels of E-cadherin protein increased(P＜0.05).Conclusions TKT is highly expressed in colorectal cancer and enhances the proliferation,migration and invasion of colorectal cancer cells.
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