上海交通大学学报（医学版）

上海交通大学学报（医学版）

• 论著（基础研究） • 上一篇    下一篇

姜黄素通过靶向PBK对结直肠癌HCT116细胞增殖的抑制作用

张生军1，刘敏丽2，常琦1，刘勇峰1   

  1. 1.延安大学附属医院普外科， 延安 716000; 2.延安大学医学院病理学教研室， 延安 716000
  • 出版日期:2016-07-28 发布日期:2016-08-31
  • 通讯作者: 刘敏丽, 电子信箱: zhangshengjun1973@126.com。
  • 作者简介:张生军(1973—)， 男， 副主任医师， 硕士; 电子信箱: zhangshengjun1973@126.com。
  • 基金资助:

    陕西省教育厅自然科学研究项目（09JK819）

Inhibiting effect of curcumin on the proliferation of colorectal cancer cell HCT116 via targeting PBK

ZHANG Sheng-jun1, LIU Min-li2, CHANG Qi1, LIU Yong-feng1   

  1. 1.Department of General Surgery, Yanan University Affiliated Hospital， Yanan 716000， China; 2.Department of Pathology, Medical College of Yanan University, Yanan 716000, China
  • Online:2016-07-28 Published:2016-08-31
  • Supported by:

    Natural Science Research Project of Education Department of Shanxi Provincial Government, 09JK819

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摘要：

目的 研究姜黄素抑制结直肠癌HCT116细胞增殖的分子机制。方法 体外培养HCT116细胞，四甲基偶氮唑蓝比色法（MTT）检测不同浓度姜黄素（0、10、20、50、100、200 μmol/L）对HCT116细胞增殖率的影响,实验分空白对照组、EGF组（单独加入20 ng/mL EGF）、EGF+姜黄素组；细胞锚定增殖实验检测药物对HCT116细胞增殖的抑制作用；体外结合及体外激酶实验检测姜黄素对磷酸化酶b激酶（PBK）活性的影响；Western blotting及免疫组织化学方法检测姜黄素对PBK下游信号通路的影响。结果 当姜黄素浓度<50 μmol/L时,对HCT116细胞影响较小；而当浓度达到100 μmol/L时，HCT116细胞凋亡明显增加。HCT116细胞在软琼脂内锚定增殖的结果显示：与EGF组比较，EGF+姜黄素组细胞克隆小且克隆数量少，具有药物浓度依赖性。体外结合及体外激酶实验结果显示姜黄素可以结合PBK，并抑制PBK的活性。Western blotting及免疫组织化学结果显示姜黄素明显下调PBK下游信号通路 ERK1/2及H3的磷酸化水平，具有时间和浓度依赖性。结论 在结直肠癌HCT116细胞中，姜黄素可能通过抑制PBK的活性，起到抗HCT116细胞增殖的作用。

关键词: 姜黄素, 结直肠癌, HCT116, 磷酸化酶b激酶, 抗肿瘤效应

Abstract:

Objective To explore the molecular mechanism of inhibiting the proliferation of colorectal cancer cell HCT116 by curcumin. Methods The HCT116 cells were cultured in vitro and assigned to the control group, EGF group (using 20 ng/mL EGF alone) and EGF+curcumin groups. The effects of different concentrations of curcumin (0, 10, 20, 50, 100, and 200 μmol/L) on the proliferation rate of HCT116 cells were detected with MTT assay. The inhibiting effect of curcumin on the proliferation of HCT116 cells was detected with cell anchoring proliferation assay. The effects of curcumin on the activity of phosphorylase b kinase (PBK) were detected with in vitro kinase assay and beads binding assay. The effects of curcumin on the downstream signal pathways of PBK were detected by Western blotting and immunohistochemistry. Results Curcumin at a concentration <50 μmol/L had small effect on HCT116 cells, while the apoptosis of HCT116 cells increased significantly when the concentration of curcumin reached 100 μmol/L. The results of cell anchoring proliferation assay for HCT116 cells in soft agar showed that the size and number of clones of HCT116 cells in the EGF+curcumin groups decreased in a concentration dependent manner as compared with the EGF group. The in vitro kinase assay and beads binding assay suggested that curcumin could bind with PBK and inhibit the activity of PBK. Results of Western blotting and immunohistochemistry indicated that curcumin significantly down-regulated the phosphorylation of PBK downstream signal molecules ERK1/2 and H3 in a concentration and time dependent manner. Conclusion Curcumin has anti-proliferation effect on colorectal cancer cell HCT116 via inhibiting the activity of PBK.

Key words: curcumin, colorectal cancer, HCT116, phosphorylase b kinase, antitumor effect