分化抑制因子1对结肠癌HT-29细胞增殖的影响

Effect of Id1 on proliferation of colon cancer HT-29 cells

目的检测分化抑制因子1(Id1)对结肠癌HT-29细胞VEGF表达的影响,探讨其在结肠癌增殖中的作用。方法 HT-29转染Id1表达质粒,RT-PCR方法和Western印迹法检测血管内皮生长因子(VEGF)mRNA和蛋白的表达,MTT法检测各组细胞增殖情况;合成Id-1干扰序列转染HT-29细胞后,RT-PCR及Western印迹检测HT-29细胞VEGF mRNA和蛋白的表达情况,采用MTT法检测各组细胞增殖情况。结果 Id1表达质粒空白组、对照组(空载体组)、过表达组VEGF/GAPDH mRNA分别为:0.002 99±0.000 687、0.003 49±0.001 34、0.031 3±0.006 05(P<0.01);VEGF/GAPDH蛋白分别为:0.132±0.035 1、0.131±0.053 0、0.245±0.032 6(P<0.01),MTT实验显示相对对照组,过表达组增殖明显快(P<0.01)。Id-1干扰空白组、对照组(空载体组)、抑表达组VEGF/GAPDH mRNA分别为:0.006 16±0.000 829、0.006 26±0.000 447、0.002 3±0.000 308(P<0.01);VEGF/GAPDH蛋白分别为:0.450±0.039 3、0.464±0.023 1、0.231±0.027 3(P<0.01)。MTT实验显示相对对照组,抑表达组增殖明显减慢(P<0.01)。Id1过表达能显著上调VEGF mRNA和蛋白表达,提高细胞的生长,Id1抑制后能显著下调VEGF mRNA和蛋白表达,减缓细胞的生长。结论在结肠癌HT-29细胞中Id1为VEGF的上游基因,Id1通过Id1↑→VEGF↑途径来影响结肠癌细胞增殖,在肿瘤血管生成中发挥重要作用。Id1有望成为结肠癌治疗的一个新靶点。

Objective To investigate the role of inhibitor of differentiation-I （ldl） on proliferation of colon cancer HT-29 cells by detecting the effect of Idl on the expression of vascular endothelial growth factor （VEGF） in hypoxia HT-29 cells. Methods The expression of VEGF mRNA and protein were detected by RT-PCR and Western blot assay in HT-29 cells after HT-29 cells transfeetion with Idl expressing plasmid. The proliferation of HT-29 cells was investigated by MTT methods after HT-29 cells transfection with Idl expressing plasmid. Results After transfection with ldl expressing plasmid, the VEGF/GAPDH mRNA expression of blank group, control group and over-ex- pression group detected by RT-PCR were 0. 002 99 ±0. 000 687, 0. 003 49±0. 001 34, 0. 031 3±0. 006 05（ P 〈0. 01 ） , and VEOF/GAPDH protein expression were 0. 132±0. 035 1 , 0. 131±0. 053 0, 0. 245±0. 032 6 respectively （ P 〈 0. 01 ）. MTT assay showed thai the cell proliferation of over-expression group was significantly faster than that of control group. After transfection with ldl-siRNA, the VEGF/GAPDH mRNA expression of blank group, control group and over-expression group were 0. 006 16±0. 000 829, 0. 006 26±0. 000 447,0. 002 3 ±0. 000 308 （ P 〈 0. 01 ） , and the VEGF/GAPDH protein were 0. 450± 0. 039 3, 0. 464±0. 023 1, 0. 231±0. 027 3 respectively （ P 〈 0. 01）. MTT assay showed that the cell proliferation of over-expression group was significantly slower than that of control group. The over-expresssion of Idl could promote the expressions of both the mRNA and protein of VEGF and ldl suppression could significantly reduce the expressions of both the mRNA and protein of VEGF. Conclusions Idl can down-regulate the expression of VEGF in HT-29 cells and play an important role in colon cancer proliferation by Id1↑-VEGF↑ way. Idl is expected to become a new target for colon cancer therapy.