结肠癌细胞株HT-29与Colo-205增殖的分子机制研究  

The proliferation mechanism of colon cancer cell lines HT-29 and Colo-205

作　　者：李小文 曹琼 徐凝馨 胡艳芬 朱剑军 刘铭[1] 李莉[1] LI Xiao-wen;CAO Qiong;XU Ning-xin;HU Yan-fen;ZHU Jian-jun;LIU Ming;LI Li(Department of Basic Medical Sciences Center,Key Laboratory of Cellular Physiology,Ministry of Education,Shanxi Medical University,Taiyuan Shanxi 030001,China)

机构地区：[1]山西医科大学基础医学研究中心细胞生理学教育部重点实验室,山西太原030001

出　　处：《毒理学杂志》2022年第2期130-136,共7页Journal of Toxicology

基　　金：国家自然科学青年基金(81902513);山西省回国留学人员科研资助项目(2020-076)。

摘　　要：目的通过蛋白激酶抑制分析探讨了结肠癌细胞株HT-29与Colo-205增殖的分子机制。方法选择多种蛋白激酶抑制剂(剂量从0.0006μmol/L开始,倍增到20μmol/L,共16个剂量组)作用于HT-29与Colo-205细胞,CCK8法检测细胞增殖,筛选有效抑制2种细胞增殖的药物;转染siRNA敲低HT-29细胞中酪氨酸蛋白激酶SRC的表达,并采用慢病毒表达质粒在Colo-205细胞中过表达SRC,检测蛋白激酶抑制剂对两种细胞增殖的影响;构建HT-29细胞裸鼠荷瘤模型,检测抑制剂对小鼠肿瘤生长的影响。结果达沙替尼(Dasatinib)或BMS-754807单独作用于HT-29细胞后,抑制效果不明显,而两者联合使用,可有效抑制HT-29细胞增殖(P=0.005)。AZD-6244单独作用于Colo-205细胞,可明显抑制细胞增殖(P=0.027)。BMS-754807单独作用于敲低SRC的HT-29细胞株,与联合使用BMS-754807和Dasatinib组相比,对细胞的抑制效果基本一致。Colo-205过表达SRC后,与对照组相比,AZD-6244抑制细胞增殖效果更明显(P=0.021)。HT-29裸鼠荷瘤实验结果表明,与对照组和BMS-754807组相比,联合用药组的小鼠肿瘤体积增长率明显降低(P=0.03)。结论HT-29细胞与Colo-205细胞的增殖受不同信号通路的调控,其中,HT-29细胞的增殖同时受胰岛素受体信号通路与SRC信号通路调控,且两条通路相互独立;Colo-205细胞增殖主要受MEK信号通路调控。Objective To explore the molecular mechanism of proliferation of colon cancer cell lines HT-29 and Colo-205 by protein kinase inhibition analysis.Methods HT-29 cells and Colo-205 cells were treated with a variety of protein kinase inhibitors(PKIs),CCK8 assay was used to detect cell proliferation and drugs that inhibit the proliferation of two kinds of cells were screened.SRC-siRNA was used to knock down the expression of SRC in HT-29 cells,SRC with the phosphorylation site of SRC-527 deleted was overexpressed using lentiviral vector transfection in Colo-205 cells,and the proliferation was detected using CCK8 assay.A nude mouse tumor model of HT-29 cells was constructed to detect the effect of inhibitors on tumor growth in vivo.Results When HT-29 cells were treated with dasatinib or BMS-754807 respectively,the inhibitory effect was not obvious,but the combination of both drugs potently inhibited the proliferation of HT-29 cells(P=0.005).The proliferation of Colo-205 cells was significantly inhibited by AZD-6244(P=0.027).The inhibitory effect of BMS-754807 on HT-29 cells after knocking down SRC was consistent with that of BMS-754807 combined with dasatinib.AZD-6244 inhibited the proliferation of Colo-205 cells more significantly than that of the control group after overexpression of SRC with SRC-527 phosphorylation site deletion in Colo-205 cells(P=0.021).The result of tumor-bearing experiment in HT-29 nude mice showed that the growth rate of tumor volume in the combination group was significantly lower than that of the control group and BMS-754807 group(P=0.03).Conclusion The proliferation of HT-29 cells and Colo-205 cells is regulated by different signal pathways.The proliferation of HT-29 cells is mostly regulated by both insulin receptor signal pathway and SRC signal pathway,and these two pathways are independent.The proliferation of Colo-205 cells is mainly regulated by MEK signal pathway.

关 键 词：结肠癌 蛋白激酶抑制剂 HT-29细胞 胰岛素受体信号通路 SRC 

分 类 号：R735.3[医药卫生—肿瘤]