Effects of up-regulated gene-4 on the proliferation of colonic cancer cells / 中华消化外科杂志

ABSTRACT

Objective To study the effects of up-regulated gene-4 (URG-4) on colon cancer cellproliferation.MethodsColon cancer cell line with high expression of URG-4 was selected.The recombinant URG-4 siRNA retroviral vector was constructed and packaged by PT67 cell,then retroviral particles which can express URG-4 siRNA in mammal cell and its negative control were obtained.Expressions of URG-4 in MKN45,SW480,LoVo,HCT116,HT29 were detected by RT-PCR and Western blot,respectively.Recombinant virus (interference group),original virus (negative control group) and the same amount of PBS (blank group) were used to transfect LoVo cells respectively.Stably transfected cell lines were screened.The growth condition of cell lines in each group was assayed by MTT.All data were analyzed by the one-way analysis of variance and the t test.Results Sequencing results confirmed the successful construction of retroviral which expressed siRNA,the relative expression levels of URG-4 mRNA in MKN45,SW480,LoVo,HCT116,HT29 were 0.58 +0.02,0.63 ±0.03,0.81 ± 0.01,1.01 ± 0.02,0.91 ± 0.04 and the expression levels of URG-4 protein in the 5 cell lines were 0.73 ±0.02,0.85 ± 0.03,1.42 ± 0.01,0.80 ± 0.30,0.80 ± 0.04,respectively.High expression of URG-4 was observed in the LoVo cells.The expression of URG-4 mRNA in the LoVo cells in the interference group was 0.55 ±0.03,which was significantly lower than 1.15 ±0.02 of the negative control group and 1.15 ±0.01 of the blank group ( t =- 5.179,- 9.285,P ＜ 0.05 ).The inhibition rate of URG-4 mRNA in the interference group was 52.6％.The expression of URG-4 protein in the interference group was 0.82 ± 0.05,which was significantly lower than 1.46 ± 0.07 of the negative control group and 1.54 ± 0.04 of the blank group (t =-4.239,-3.704,P＜0.05).The inhibition rate of URG-4 protein in the interference group was 43.6％.The LoVo cells in each group grew exponentially.Compared with the negative control group,the cell growth of the interference group was inhibited during day 3 to day 6,which had statistical significant difference ( t =- 6.436,-6.045,-6.434,-4.285,P＜0.05).ConclusionInterference of the expression of URG-4 can inhibit the growth of LoVo cells.