The Relationship Between The Inhibitory Effect Of Berberine On Proliferation In Hct116Cells And Wnt/B-catenin Signal Pathway

AimThis study was aimed to investigate the relationship between theinhibitory effect of berberine on proliferation in HCT116colorectal cancercells and Wnt/β-catenin signal pathway, and unveil the possible mechanismof berberine for this function.MethodsCrystal violet staining assay, immunofluorescence staining,flowcytometry assay, RT-PCR, Western blotting, and luciferase reporterplasmid assay were employed in this study. The detailed methods were asfollowing:1. Crystal violet staining, immunofluorescence stain and flowcytometryassay were performed to assay the effects of berberine on proliferation,apoptosis, and cell cycle arrest in HCT116colorectal cancer cells,respectively.2. Luciferase reporter assay was used to evaluate the influence of berberineon Wnt/β-catenin signal transduction. 3. Western blot assay was employed to detect the effect of bernerine on theexpression of β-catenin in the nucleus, cytoplasm, and the whole cellprotein in HCT116cells.4. Exogenous over-expression and knockdown techniques were introducedto determine the role of β-catenin in berberine-induced proliferationinhibitory effect in HCT116cells.5. GSK3β inhibitor LiCl was used to determine the role of β-catenin inberberine-induced proliferation inhibitory effect in HCT116cells.6. RT-PCR assay was performed to explore the effect of berberine onβ-catenin mRNAexpression in HCT116cells.Results1. Berberine could inhibit proliferation, induce apoptosis and arrest the cellcycle at G1phase in HCT116cells.2. Transduction activity of Wnt/β-catenin singnal was reduced by berberinein HCT116cells.3. Berberine decreased the β-catenin protein level of HCT116cells incytoplasm, nucleus, and whole cells.4. Exogenous over-expression of β-catenin attenuated the inhibitory effectof berberine in HCT116cells, whereas knockdown of β-catenin promotesthe inhibitory effect.5. LiCl, an inhibitor of GSK3β, failed to reverse the inhibitory effect ofberberine on proliferation and the reduction of β-catenin protein level in cytoplasm, nucleus, and whole cell in HCT116cells.6. Berberine inhibited the mRNA expression of β-catenin in HCT116cells.ConclusionThe effects of berberine on proliferation inhibition, apoptosis inductionand cell cycle arrest in HCT116cells are relative to Wnt/β-catenin signalpathway. This may at least partly result from the inhibition of β-cateninmRNA expression by berberine in HCT116cells.