title: Construction of interference vector targeting Ep-CAM gene and its effects on colorectal cancer cell proliferation.
abstract: BACKGROUND
  Prior study indicates that abnormal protein expression and functional changes in the development and progression of colorectal cancer is related to gene expression. The aim of this study was to construct an interference plasmid targeting the Ep-CAM gene and to investigate its effects on the proliferation of colorectal cancer cells.
  
  
  METHODS
  In this study, HT-29 and HCT-116 colorectal cancer cell lines were selected as cell models. The double-stranded micro (mi)RNA oligo was inserted into the pcDNATM6.2-GW/EmGFPmiR vector, which is an expression of miRNA. Lipofectamine™ 2000 was used to transfer plasmid into the empty plasmid group (transfected pcDNATM6.2-GW/EmGFPmiR-neg) and the interference group (transfected pcDNATM6.2-GW/EmGFPmiR-Ep-CAM-1), respectively. Meanwhile, the nontransferred HT-29 and HCT-116 acts as the blank control group. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect the transfection efficiency. Western blot was used to detect Ep-CAM protein expression. The cell proliferation in each group was detected by using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay.
  
  
  RESULTS
  The results indicated that the Ep-CAM messenger (m)RNA expression in the interference group was lower significantly compared with that of the empty plasmid group and control group (P<0.01). Western blot analysis results showed that Ep-CAM protein expression was significantly lower in interference group compared with that of the empty plasmid group and the control group (P<0.01). MTT assay results demonstrated that the proliferation ability of cells in the interference group was significantly inhibited compared with the two other groups (P<0.05).
  
  
  CONCLUSIONS
  Silencing of Ep-CAM can significantly inhibit the proliferation of colorectal cancer cells.
authors.affiliation: Department of Gastroenterology, Binzhou People's Hospital, Binzhou, Shandong, People's Republic of China.
authors.forename: Yanmei
authors.initials: Y
authors.lastname: Qi
authors.affiliation: Department of General Surgery, Binzhou People's Hospital, Binzhou, Shandong, People's Republic of China.
authors.forename: Fengqiang
authors.initials: F
authors.lastname: Zhou
authors.affiliation: Department of General Surgery, Binzhou People's Hospital, Binzhou, Shandong, People's Republic of China.
authors.forename: Lu
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Department of General Surgery, Binzhou People's Hospital, Binzhou, Shandong, People's Republic of China.
authors.forename: Lei
authors.initials: L
authors.lastname: Liu
authors.affiliation: Department of General Surgery, Binzhou People's Hospital, Binzhou, Shandong, People's Republic of China.
authors.forename: Hong
authors.initials: H
authors.lastname: Xu
authors.affiliation: Department of General Surgery, Binzhou People's Hospital, Binzhou, Shandong, People's Republic of China.
authors.forename: Huiguang
authors.initials: H
authors.lastname: Guo
mesh_terms: D000951:Antigens, Neoplasm / Q000235:genetics*; D015815:Cell Adhesion Molecules / Q000235:genetics*; D049109:Cell Proliferation / Q000235:genetics*; D015179:Colorectal Neoplasms / Q000235:genetics*; D000071858:Epithelial Cell Adhesion Molecule; D020868:Gene Silencing; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D035683:MicroRNAs / Q000235:genetics; D010957:Plasmids; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D014162:Transfection
chemical_list: D000951:Antigens, Neoplasm; D015815:Cell Adhesion Molecules; C499379:EPCAM protein, human; D000071858:Epithelial Cell Adhesion Molecule; D035683:MicroRNAs; D012333:RNA, Messenger
keywords: Ep-CAM; colorectal cancer; epidermal growth factor; vector construction