【摘要】 目的观察过表达CDX2基因对人结肠癌细胞株增殖影响,并初步探讨其作用机制。方法通过基因转染技术将CDX2基因转染入结肠癌HT-29细胞株,经G418筛选后获得稳定转染细胞株,RT-PCR和Western blot检测CDX2基因表达。采用四甲基偶氮唑盐(MTT)法、流式细胞仪分析转染细胞的生物学行为。Western blot检测CDX2过表达对HT-29细胞内Cyclin D1和NF-κB磷酸化水平的影响。结果经脂质体转染和筛选,建立了稳定过表达CDX2人结肠癌HT-29细胞株。转染CDX2组与未转染组和空白质粒组相比,细胞的生长速度减慢(P <0. 05),细胞周期中G1/G0期比例增加(P <0. 05),而S期比例则减少(P <0. 05);与未转染组和空白质粒组比较,转染CDX2基因组HT-29细胞的Cyclin D1和p-NF-κB活性明显降低。结论 CDX2基因可能通过抑制NF-κB通路抑制Cyclin D1活性,从而抑制人结肠癌HT-29细胞增殖。更多还原

【Abstract】 Objective The aim of this study was to observe the effects of overexpressing CDX2 gene on proliferation of human colon cancer HT-29 cell line and to explore its mechanism. Methods CDX2 was transfected into HT-29 cells by gene transfection technology,and stable transfected cell clones with stably expressing CDX2 gene were obtained by G418 selection. The expression of CDX2 was detected by RT-PCR and Western blotting. The biological behavior of transfected cells were analyzed by methythiazoletertraolium( MTT) assay and flow cytometry. Western blot assay was used to detect the expression of cyclin D1 and NF-κB phosphorylation level in HT-29 cells with overexpressing CDX2 gene. Results A system for stably overexpressing CDX2 human colon cancer HT-29 cell line was established. Compared with the nontransfected and empty vector transfected HT-29 cells,the growth rate of the cells in the transfected CDX2 group was significantly slower( P < 0. 05). The proportion of G1/G0 phase of cell cycle was significantly increased( P < 0. 05),and the proportion of S phase was significantly decreased( P < 0. 05). Compared with the non-transfected and empty vector transfected cells,the expression of cyclin D1 and p-NF-κB activity were significant low in overexpressing CDX2 cells. Conclusion CDX2 gene may inhibit the proliferation of HT-29 cells through blocking the activity of NF-κB pathway and down-regulation of cyclin D1 protein expression.更多还原