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安罗替尼在肝癌中耐药的差异mRNA筛选

谷俊谋 王立博 曾德俊 陆勤伟 董凯 梁若鹏 王维杰 朱荣涛 孙玉岭

引用本文:
Citation:

安罗替尼在肝癌中耐药的差异mRNA筛选

DOI: 10.3969/j.issn.1001-5256.2021.02.022
基金项目: 

国家自然科学基金 (81870457)

国家自然科学基金 (81900558)

利益冲突声明:本研究不存在研究者、伦理委员会成员、受试者监护人以及与公开研究成果有关的利益冲突,特此声明。
作者贡献声明:谷俊谋负责课题设计,资料分析,撰写论文;王立博、曾德俊、陆勤伟、董凯、梁若鹏、王维杰、朱荣涛参与收集数据,修改论文;孙玉岭负责拟定写作思路,指导撰写文章并最后定稿。
详细信息
    作者简介:

    谷俊谋(1993—),男,主要从事肝脏相关疾病的研究

    通信作者:

    孙玉岭,doctorssh@126.com

  • 中图分类号: R735.7

Differentially expressed mRNA involved in the resistance of liver cancer to anlotinib

  • 摘要:   目的  运用ceRNA芯片筛选可能参与肝癌细胞对安罗替尼耐药过程的mRNA。  方法  利用大剂量冲击联合低剂量诱导的方法建立对安罗替尼耐药的肝癌细胞,用CCK8实验进行验证耐药细胞在安罗替尼作用下的细胞增殖差异;运用ceRNA芯片检测耐药肝癌细胞与正常肝癌细胞的基因表达差异;运用实时荧光定量PCR(real-time PCR)对部分芯片测出的部分基因差异进行验证。计量资料两组间比较采用独立样本t检验,Kaplan-Meier法对肝癌样本的总生存期进行生存分析,log-rank检验比较生存率差异。芯片筛选结果使用Fisher精确检验。  结果  耐药肝癌细胞与正常肝癌细胞基因表达差异较大,通过缩减范围筛选出差异最大的10个基因进行分析。与耐药和肿瘤生长相关的基因有4个,分别为BIRC2、BIRC7、ABCC2、MAPK8。其中BIRC2、ABCC2、MAPK8表达水平下降(P值分别为0.001 4、0.001 2、0.011 8),BIRC7的表达水平增多(P<0.001)。real-time PCR的验证结论与芯片一致(t值分别为10.74、32.65、18.34、2.80,P值分别为0.000 4、0.000 1、0.000 1、0.044 8)。BIRC7的高表达与MAPK8的低表达对应显著减少的生存期(P值分别0.022 0、0.005 6)。  结论  BIRC2、BIRC7、ABCC2、MAPK8在对安罗替尼耐药的肝癌细胞中差异表达,可能参与了肝癌细胞对安罗替尼耐药的过程。

     

  • 图  1  CCK8法检测安罗替尼对Huh7和Huh7/R的抑制作用

    图  2  差异mRNA热图见二维码

    图  3  正常肝癌细胞与耐药肝癌细胞差异基因的GO富集分析图

    图  4  正常肝癌细胞与耐药肝癌细胞差异基因的KEGG pathway分析图

    图  5  4个基因的mRNA在正常肝癌细胞和耐药肝癌细胞中的表达水平比较

    注:a, BIRC2;b, ABCC2;c, MAPK8;d.BIRC7。

    图  6  BIRC2、ABCC2、MAPK8、BIRC7的生存分析

    注:a, BIRC2; b,ABCC2; c, MAPK8;d, BIRC7。

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  • 收稿日期:  2020-09-29
  • 录用日期:  2020-11-27
  • 出版日期:  2021-02-20
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