Effects Of CENP-EWT Gene Silencing On Human Colon Cancer Cellline HCT116

Objective: To explore the effect of down-regulating wild typecentromere protein E (CENP-EWT) expression by RNA interference onhuman colon cancer cell line HCT116.Methods: HCT116cells were divided into three groups: blank controlgroup, empty vector group(transfected with an empty vector), andexperiment group (transfected with CENP-EWT shRNA). After HCT116cells transfected with CENP-EWT shRNA,the mRNA expression ofCENP-EWT was examined by Nested PCR; cell proliferation was evaluatedby MTT assay; cell apoptosis was detected by Hoechst assay;cell migrationand invasion were detected by Transwell assay.Designed and synthesized for centromere protein E deletion(Varianttype Centromere protein E,CENP-E I) gene specific six pairs of siRNAinterference sequence,while previous experiments can effectively interferewith wild-type gene sequence of the interference of the centromere protein Esequence synthesis,using molecular cloning techniques were cloned intopSES-HUS adenovirus shuttle plasmid,the recombinant plasmid. Results: Compared with cells of the blank control and negative controlgroup,the expression of CENP-EWT mRNA (p<0.05) was down-regulatedin cells transfected with CENP-EWT shRNA group; the cell growthinhibition was significantly in the CENP-EWT shRNA group by MTTassay(p<0.05).the cell apoptotic rate was significantly increased in theCENP-EWT shRNAgroup by Hoechst assay(p<0.05). the cell migration andinvasion abilitywas evidently increased inthe CENP-EWTshRNAgroup byTranswell assay(p<0.05).Five pairs of siRNA for CENP-E I gene and one pair of siRNA forCENP-EWT,Successfully cloned into pSES-HUS adenovirus shuttleplasmid,and get a good transfection efficiency and interference effects.Conclusion: RNA interference targeting CENP-EWT genedown-regulates the CENP-EWT expression,inhibits cellproliferation,induces cell apoptosis,but increases cell migration andinvasion ability in human colon cancer cell line HCT116.