干涉NFATc1表达影响细胞周期运转抑制结肠癌细胞增殖的机制

Mechanism study of interfered NFATc1 expression suppressing cell cycle progression in colorectal cancer cell

目的根据活化T-细胞核因子1(NFATc1)在结肠癌中的表达及预后情况,利用shRNA转染细胞干涉NFATc1表达,检测NFATc1对结肠癌细胞增殖各表型的影响,并初步分析可能的机制。方法利用TCGA数据库分析NFATc1高表达对结肠癌预后的影响,取临床结肠癌患者术后样本进行免疫组化染色,比较NFATc1在结肠癌组织和癌旁组织中的表达差异。通过将shRNA转染结肠癌细胞干涉NFATc1后,利用CCK-8测定NFATc1干涉后的细胞增殖变化;利用克隆形成实验预测NFATc1干涉后结肠癌细胞成瘤潜力;利用碘化丙锭染色细胞,通过流式细胞仪分析细胞周期分布变化;利用qPCR检测NFATc1对结肠癌细胞周期相关因子转录活性的影响。结果生存曲线显示NFATc1高表达的结肠癌患者预后更差。临床结肠癌组织NFATc1的表达高于癌旁正常组织。干涉NFATc1导致结肠癌细胞的增殖速率明显降低,克隆形成能力显著减弱,细胞周期出现G0/G1期停滞。qPCR结果表明干涉NFATc1后,多个关键的细胞周期抑制因子转录活性明显上升。结论NFATc1通过抑制细胞周期调控因子转录活性促进细胞周期运转,从而促进结肠癌细胞增殖和成瘤能力。

Objective Based on the expression of NFATc1 and prognosis in colorectal cancer,we interfered NFATc1 via shRNA transfection,analyzed the effect of NFATc1 on colorectal cancer cell proliferation,and further explored potential mechanisms.Methods We explored the correlation between NFATc1 expression and the prognosis of colorectal cancer using the TCGA database.Thereafter,we compared the expression of NFATc1 in colorectal cancer tissues and adjacent normal tissues by immunohistochemical staining of postoperative samples from clinical colorectal cancer patients.We analyzed the effect of NFATc1 on colorectal cancer cell proliferation by measuring CCK-8 after NFATc1 interfered using shRNA transfection;Tumorigenic potential of colorectal cancer cells was measured with clone formation assay;Cell cycle distribution was measured by flow cytometry after propidium iodide stai-ning.The effect of NFATc1 on the transcriptional activity of cell-cycle-related factors was measured by qPCR.Results Based on the TCGA data,we found that high NFATc1 expression in colorectal cancer patients was associated with poor prognosis.The expression of NFATc1 in clinical colorectal cancer tissues was significantly higher than that in adjacent normal tissues.Additionally,interference with NFATc1 inhibited the proliferation rate of colorectal cancer cells in vitro,and the clonogenic capacity of cells was impaired.As expected,the cell cycle was arrested at the G0/G1 phase.The qPCR results indicated that the knockdown of NFATc1 increased the transcriptional activity of multiple key cell cycle inhibitors.Conclusion NFATc1 promotes cell cycle progression by inhibiting the transcriptional activity of cell cycle regulatory factors,thereby promoting the proliferation and tumorigenic ability of colorectal cancer cells.