Study On The Relationship Between The Proliferation Inhibitory Effect Of Resveratrol In HCT116Cells And PI3K/Akt And Wnt/β-catenin

AimIn this investigation, we demonstrated the anti-proliferation andapoptosis-inducing activities of resveratrol in human colon cancer cells,and dissected the possible mechanisms underlying these effects.MethodsCrystal violet staining assay, HE staining, flowcytometry assay,immunofluorescence staining, RT-PCR, Immunohistochemistry, Westernblotting, and luciferase reporter assay were used in our investigation. Thedetail investigations were processed as following:1. Crystal violet staining, Western blot and immunofluorescence stainingwere employed to explore the effects of resveratrol on proliferation,apoptosis, and cell cycle arrest in HCT116colon cancer cells.2. A xenograft tumor model was used to confirm the anti-tumor effects ofresveratrol.3. We introduced RT-PCR, Western blot assay to test the effects of resveratrol on Akt1, Akt2, Akt1/2and p-Akt1/2protein levels.4. With PCR and Western blot assay, we detected the effect of resveratrolon expression of PTEN mRNA and protein level in HCT116coloncancer cells.5. Using recombinant adenovirus mediated PTEN over-expression orknockdown to determine the role of PTEN in resveratrol-inducedproliferation inhibitory effect in HCT116cells.6. We employed Western blot, luciferase reporter and RT-PCR assay todetect the effect of resveratrol on the protein level of GSK3β andp-GSK3β, Wnt/β-catenin signal activation and the mRNA level ofβ-catenin in HCT116cells.Results1. Resveratrol inhibits proliferation, promote apoptosis and arrest the cellcycle at G1phase in HCT116cells.2. Resveratrol inhibits the tumor growth in xenograft colon cancer model3. Resveratrol increases the mRNA and protein leve of PTEN in HCT116cells.4. Over-expression of PTEN strengthen the inhibitory effect of resveratrolin HCT116cells, however PTEN knockdown or PTEN specific inhibitorenhances the inhibitory effect of resveratrol.5. Resveratrol reduces p-Akt1/2protein levels, and over-expression ofPTEN potentiates this effect, whereas silence the expression of PTEN does no apparently change on p-Akt1/2protein levels.6. Resveratrol decreases the level of p-GSK3β, inhibits the transcriptionalactivity of β-catenin/TCF4, and down regulates the mRNA expressionof β-catenin.ConclusionResveratrol inhibits proliferation, promote apoptosis and cell cyclearrest in colon cancer cells. These effects may be mediated by inhibiting thesignaling transduction of PI3K/Akt and Wnt/β-catenin signal pathwaythrough up-regulating PTEN and down-regulating β-catenin, respectively.