Objective To investigate the effects of Tspan8 gene knockout combined with anlotinib on the proliferation, migration, invasion, and apoptosis of colon cancer SW480 cells.
Methods The plasmid was constructed by CRISPR/Cas9 technique, and Tspan8 gene was knocked out in SW480 cells. The knockout effect was detected by Western blot. The IC50 of anlotinib was calculated by MTT assay. The experiment was divided into control group, anlotinib group, Tspan8 knockout group, and combined group. Cell proliferation, migration, invasion, and apoptosis were detected by cell proliferation assay, clonal formation assay, scratch assay, Transwell chamber assay, and flow cytometry. Western blot was used to detect the effect of anlotinib on Tspan8 expression in SW480 cells.
Results SW480-KO-Ⅲ cells had the highest knockout efficiency in the Tspan8 knockout group. They were used in subsequent experiments. Different concentrations of anlotinib could inhibit the proliferation of SW480 cells at different times (P < 0.01) in a concentration-dependent and time-dependent manner (P < 0.01). According to IC50, 14 μmol/L was selected as the subsequent experimental concentration. Compared with those in the control group, the cell proliferation, migration, and invasion abilities in the anlotinib group, Tspan8 knockout group, and combined group were significantly decreased, and the cell apoptosis level was significantly increased (P < 0.05). The above changes in the combined group were more significant than those in the anlotinib group or Tspan8 knockout group (P < 0.05). Compared with that in the control group, the expression level of Tspan8 in SW480 cells in the anlotinib group was significantly decreased (P < 0.01).
Conclusion Tspan8 gene knockout combined with anlotinib can synergistically inhibit the proliferation, migration and invasion of SW480 cells. This combination can also promote the apoptosis of these cells.
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