目的 探讨化疗耐药与肿瘤干细胞的关系以及逆转化疗耐药的途径,指导抗肿瘤合理用药。方法 分别培养普通型和耐药型肝癌细胞系HepG-2,建立细胞增殖模型观察化疗药物氟尿嘧啶和干细胞诱导分化剂全反式维甲酸对肝癌细胞的增殖抑制作用,通过流式分析术比较普通型和耐药型HepG-2细胞中肿瘤干细胞的含量差异,运用Western Blot和Realtime-PCR技术检测普通型和耐药型肝癌细胞中干性基因的表达水平。结果 氟尿嘧啶单用时,其对耐药型HepG-2细胞的增殖抑制作用明显低于普通型细胞;加入全反式维甲酸后,耐药型细胞的增殖速率显著降低。与普通型HepG-2细胞相比,耐药型细胞中CD44和CD326阳性细胞的比例明显增高,ABCG2、BMI、OCT4的mRNA和蛋白表达量均显著升高;而加入维甲酸后,ABCG2、BMI、OCT4蛋白表达水平又有不同程度的下降。结论 化疗耐药与肿瘤干细胞的富集相关,维甲酸可以下调肿瘤干细胞相关基因的表达、逆转肿瘤细胞的耐药性。这为临床上合理应用抗肿瘤药物提供了新的思路。
Abstract
OBJECTIVE To investigate the relationship between chemotherapy resistance and cancer stem cells, probe the way to reverse drug resistance, and guide the rational medical treatment. METHODS Hepatocellular carcinoma cell lines, parent and chemoresistant HepG-2, were cultured with fluorouracil (5-FU). CCK-8 cell proliferation assay was used to determine the inhibition of 5-Fu and all-trans retinoic acid (ATRA), a cell differentiation inducer. The content difference of cancer stem cells between parent and chemoresistant HepG-2 cell lines was examined by flow cytometry. Western blot and realtime PCR were preformed to probe the difference of related gene expression level between parent and chemoresistant HepG-2 cell lines. RESULTS Our data supported that the inhibitory effect of 5-Fu alone on chemoresistant HepG-2 cells proliferation was lower than on parent HepG-2 cells. After the addition of ATRA, chemoresistant HepG-2 cells proliferation was reduced obviously. Flow cytometry RESULTS revealed that compared with parent HepG-2 cells, the ratio of CD44 and CD326 positive cells in chemoresistant HepG-2 cells was much more than that in parent HepG-2 cells. Furthermore, realtime PCR and Western blot suggested that ABCG2, BMI and OCT4 expressions were increased in chemoresistant HepG-2 cells than those in parent HepG-2 cells, while ATRA could attenuate these differences on protein expressions. CONCLUSION Cancer stem cells enrichment relates to chemotherapy resistance, which can be reversed by ATRA through down-regulation cancer stem cells associated gene expression. This can provide new ideas for clinical rational use of anti-tumor drugs.
关键词
肝癌 /
耐药性 /
肿瘤干细胞 /
氟尿嘧啶 /
全反式维甲酸
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Key words
hepatocellular carcinoma /
drug resistance /
cancer stem cell /
fluorouracil /
all-trans retinoic acid
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中图分类号:
R965
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参考文献
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脚注
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