oct4在肝癌干细胞中的表达及对其耐药性的影响 - 第三军医大学学报.doc

肝癌细胞Oct4、ABCG2对其耐药性影响的初步研究Toronto, Ontario, Canada , Toronto General Research Institute，University of Toronto2） [摘要] 目的 富集培养肝癌干细胞，初步探讨八聚体4（Oct4）和ATP结合盒转运蛋白ABCG2在肝癌细胞及肝癌干细胞中的表达及其对耐药性的影响。方法 使用流式细胞仪从肝癌细胞中分选肝癌干细胞，无血清培养肝癌干细胞球。实验分为肝癌细胞组及肝癌干细胞组（检测HepG2、Hep3B两种细胞系）。单细胞克隆形成实验分析检测细胞增殖能力。分别给予肝癌细胞及肝癌干细胞阿霉素处理，MTT法测阿霉素处理后各组细胞的存活率，PCR法及western blot法分别检测各组细胞CD90，CD133，Oct4及ABCG2基因mRNA及蛋白水平的表达。结果 肝癌干细胞单个细胞增殖能力强于肝癌细胞，克隆形成分析显示14 d克隆形成率Hep3B细胞组为8/27（30%）、Hep3B CSCs组为12/23（52%），p0.05；HepG2细胞组克隆形成率为7/38（18%）、HepG2 CSCs组为9/27（35%），p0.05。用阿霉素处理肝癌细胞及肝癌干细胞后，MTT法测定结果显示肝癌干细胞组细胞活性高于肝癌细胞组，处理48 h后各组细胞活性：HepG2细胞组细胞活性为（38.17 ± 6.92）%、HepG2 CSCs组为（69.88 ± 5.43）%，p0.05；Hep3B细胞组（50.16 ± 4.89）%，Hep3B CSCs组为（78.53 ± 7.86）%，p0.05。 RT-PCR及qPCR结果显示肝癌干细胞组中CD90、CD133、Oct4及ABCG2基因mRNA表达较亲代细胞组显著上调（p0.05）。Western blot结果显示肝癌干细胞组Oct4及ABCG2蛋白水平显著上调，与亲代细胞组间表达差异有显著性（p0.05）。 结论CD90，CD133，Oct4及ABCG2均高表达于肝癌干细胞，且Oct4及ABCG2与肝癌干细胞耐药性密切相关。 [关键词] 肝癌干细胞；CD90；CD133；Oct4；ABCG2；耐药性 [中图法分类号] R735.7 [文献标志码] A Analysis of Oct4 and ABCG2 expression and chemoresistance in liver cancer stem cells Jia Qian1, Gao Jian1, Zhang Xiaoli1，Xiang Ying1, Deng Tao2 （1Department of Gastroenterology, Second Affiliated Hospital, Chongqing Medical University, Chongqing, China, 400010, 2M5G 2M1 Toronto, Ontario, Canada , Toronto General Research Institute，University of Toronto） [Abstract] Objective To enrich liver cancer stem cells (CSCs) and, explore the differential expression and chemotherapy resistance of octamer 4 (Oct4) and the ATP-binding cassette transporter protein ABCG2 between liver cancer cells and liver CSCs. Methods Liver cancer stem cells (detect HepG2, Hep3B cell lines) were isolated by flow cytometry, cells cultured in serum-free medium to form spheres. Cells were divided into two groups, including liver cancer cells group and liver CSCs group. Cell proliferation capacity was detected by single-cell clonogenic assay. Cell activity was detected by MTT assay after doxorubicin was pretreated. The expressi