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ShRNA-mediated gene silencing of β-catenin inhibits growth of human colon cancer cells

2007; Baishideng Publishing Group; Volume: 13; Issue: 48 Linguagem: Inglês

10.3748/wjg.v13.i48.6581

2219-2840

Wen‐Sheng Huang, Jianping Wang, Ting Wang, Jieyu Fang, Ping Lan, Jin-ping Ma,

RNA Research and Splicing

AIM:To observe the gene silencing mediated by the specific shRNA targeted against β-catenin and its effect on cell proliferation and cycle distribution in the human colon cancer cell line Colo205. METHODS:Two shRNA plasmid vectors against β-catenin were constructed and transfected into Colo205 cells with Lipofectamine TM 2000.The down-regulations of β-catenin, c-myc and cyclinD1 expressions were detected by RT-PCR and western blot analysis.The cell proliferation inhibitions were determined by MTT assay and soft agar colony formation assay.The effect of these two β-catenin shRNAs on cell cycle distribution and apoptosis was examined by flow cytometry. RESULTS:These two shRNA vectors targeted against β-catenin efficiently suppressed the expression of β-catenin and its down stream genes, c-myc and cyclinD1.The expression inhibition rates were around 40%-50% either at the mRNA or at the protein level.The shRNA-mediated gene silencing of β-catenin resulted in significant inhibition of cell growth both on the culture plates and in the soft agar.Moreover, the cancer cells showed significant G0/G1 arrest and increased apoptosis at 72 h post transfection due to gene silencing. CONCLUSION:These specific shRNAs targeted against β-catenin could have a gene silencing effect and block the WNT signaling pathway.They could inhibit cell growth, increase apoptosis, and induce cell cycle arrest in Colo205 cells.ShRNA interference against β-catenin is of potential value in gene therapy of colon cancer.