MiR-101负性调节人结肠癌细胞HCT116增殖机制的研究

Negative regulation of MiR-101 on the proliferation of colon cancer HCT116 cells and its mechanism

目的研究miR-101对结肠癌细胞增殖能力的影响及与分子学机制。方法人工干预miR-101在结肠癌细胞株HCT116中的表达,分别转染阴性对照物(对照组)、miR-101抑制物(实验Ⅰ组)、miR-101模拟物(实验Ⅱ组)48 h后,用细胞增殖法分析其对人结肠癌HCT116细胞增殖能力的影响。用免疫印迹法测定miR-101在调节Notch1表达中的作用。结果实验Ⅰ组、实验Ⅱ组及对照组的miR-101表达量分别为0.467,1.767,0.967。与对照组相比,mimic-miR-101使HCT116细胞中miR-101的表达水平明显提高,差异有统计学意义(P<0.05)。实验Ⅱ组细胞miR-101的表达水平较对照组显著降低,差异有统计学意义(P<0.05)。在72 h,实验Ⅰ组、实验Ⅱ组及对照组OD值分别为1.10,0.76,0.91。在96 h,实验Ⅰ组、实验Ⅱ组及对照组OD值分别为1.57,0.92,1.20。与对照组相比,实验Ⅱ组的细胞增殖能力明显被抑制,差异均有统计学意义(均P<0.05)。miR-101调节结肠癌细胞HCT116增殖的影响是通过靶向调节Notch1的表达来实现的。结论在结肠癌发生及进展过程中,miR-101发挥抑癌基因作用,可靶向干扰Notch1的表达,进而负性调节结肠癌细胞的增殖。

Objective To study the effects and the molecular mechanism of miR-101 on the proliferation of colon cancer HCT116 cells. Methods The expression of miR-101 was regulated and treated by negative control（ control group）, inhibitor-miR-101（ experimental Ⅰgroup）,mimic-miR-101（ experimental Ⅱ group） for 48 h,and the cell proli-feration was tested by [3-（ 4,5-dimethylthiazol-2-yl）-5-（ 3-carboxymethoxyphenyl）-2-（ 4-sulfophenyl）-2H-tetrazolium,MTS] assay. The Notch1 expression was also measured by Western blotting. Results The expression of miR-101 in experimental Ⅰgroup,experimental Ⅱ group and control group were 0. 467,1. 767,0. 967,respectively. The expression of miR-101 was significantly over-expressed by mimic-miR-101,or significantly inhibited by inhibitor-miR-101,compared with control group,the differences were statistica-lly significant（ all P〈0. 05）. After 72 h treatment,the OD values of experimentalⅠgroup,experimental Ⅱ group and control group were 1. 10,0. 76,0. 91. After 96 h treatment,the OD values of experimental Ⅰgroup,experimental Ⅱ group and control group were 1. 57,0. 92,1. 20. Compared with control group,the cell proliferation in experimental Ⅱ group were significantly inhibited, the differences were statistically significant（ all P〈0. 05）. The negatively regulated effect of miR-101 on the proliferation of HCT116 cell was taken by targeting Nocth1 expression. Conclusion As a cancer suppressor gene,miR-101 negatively regulated the proliferation of colon cancer HCT116 cells by targeting noth1 expression.