射频消融亚致死性温度对肝癌干细胞的影响  

The effect of radiofrequency ablation with sublethal temperature on the stem cells of hepatocellular carcinoma

作　　者：郝美君[1,2] 刘凯[1] 郭向华[1] 欧阳雅博[1] 乔录新[1] 石英[1] 陈德喜[1] 郑加生[1,2] HAO Meijun LIU Kai GUO Xianghua OUYANG Yabo QIAO Luxin SHI Ying CHEN Dexi ZHENG Jiasheng(Affiliated Beijing You ＇an Hospital of Capital Medical University, Beijing 100069, China)

机构地区：[1]首都医科大学附属北京佑安医院 [2]北京市肝病研究所

出　　处：《介入放射学杂志》2017年第7期636-640,共5页Journal of Interventional Radiology

基　　金：国家自然科学基金(81402556;81272266);NSFC-NIH国际合作项目(81361120401);首都发展基金(首发2014-1-1151);北京市肝病研究所流式平台基金(BJIH-01602)

摘　　要：目的研究亚致死温度射频消融(RFA)对肝癌干细胞(LCSC)产生及其相关转录因子表达的影响。方法利用小鼠Hep1-6肝癌细胞株和肝细胞癌(HCC)患者临床样品,检测LCSC相关标志物和转录因子的表达。结果不同温度分别刺激Hep1-6细胞后发现,45℃是不能诱导细胞死亡的亚致死性温度。流式细胞仪(FCM)检测显示45℃处理可引起CD13^+、CD44^+、CD90^+、CD133^+Hep1-6细胞水平明显上调,提示45℃温度导致Hep1-6细胞中以上各型LCSC产生增加;实时定量聚合酶链反应(RT-q PCR)检测显示45℃温度导致CD13、CD90、CD133 m RNA水平明显上调。CD13 m RNA水平在5例HCC患者复发肝癌组织中均明显上调,CD133 m RNA在4例复发肝癌中上调,CD90 m RNA仅在1例复发肝癌中上调;FCM检测显示CD13^+LCSC水平在4例复发肝癌中明显上调,CD133^+LCSC水平仅在1例复发肝癌中上调,提示CD13^+LCSC水平上调与45℃温度关系更密切。RT-q PCR检测显示4例CD13^+LCSC上调的复发肝癌患者13个LCSC相关转录因子中Sox2、Stat1明显上调,FCM检测显示45℃处理Hep1-6细胞后Sox2、Stat1 m RNA明显上调。用Sox2、Stat1 si RNA分别沉默了Sox2、Stat1基因,表明Sox2、Stat1均参与了45℃温度诱导的CD13^+LCSC产生。结论 RFA治疗中45℃亚致死性温度所致CD13^+LCSC水平增高与Sox2、Stat1表达有关。该结果对肝癌复发研究有一定借鉴意义。Objective To study the effect of radiofrequency ablation （RFA） with sublethal temperature on the production of liver cancer stem cells （LCSCs） and on the expression of LCSCs-related transcriptional factors. Methods Mouse hepl-6 hepatoma cell line and clinical samples of patients with hepatocellular carcinoma （HCC） were used to test the expressions of LCSCs-related markers and transcriptional factors. Results Different temperatures were used to stimulate Hepl-6 cells, and it was proved that the temperature of 45℃ was a sublethal temperature that could not induce cell death. Flow cytometry testing showed that treatment with 45℃ could obviously increase CD13^＋, CD44^＋, CD90 and CD133^＋ Hepl-6 cells, suggesting that treatment with 45℃ could increase the production of above mentioned types of LCSCs in hep1-6 cells. Real-time quantitative polymerase chain reaction （RT-qPCR） assay indicated that the temperature of 45% could cause significant increase in CD13, CD90 and CD133 mRNA. In all 5 HCC patients, CD13 mRNA in the recurrent HCC lesions was remarkably increased, CD133 mRNA was increased in 4 patients with recurrent HCC, and CD90 mRNA was increased in only one patient with recurrent HCC. Flow cytometry testing revealed that CD13^＋ LCSCs were strikingly increased in 4 recurrent HCC patients, while CD133^＋ LCSC was increased in only one patient, suggesting that more close correlation existed between the increase of CD13^＋ LCSCs and the temperature of 45℃. RT-qPCR assay showed that in 4 recurrent HCC patients with increased CD13^＋ LCSC, the Sox2 and Statl among 13 LCSCs-related transcriptional factors were obviously increased. Flow cytometry testing showed that 45℃ treatment also increased the expression of Sox2 and Statl mRNA in Hep1-6 cells. Finally, Sox2 and Stat1 could be knockdown by siRNAs, indicating that both Sox2 and Statl transcriptional factors were involved in 45℃-induced production of CD13^＋ LCSCs in Hep1-6 ceils. Conclusion In RFA therapy, the use of sublethal te

关 键 词：肝细胞癌 射频消融 亚致死温度 肝癌干细胞 

分 类 号：R735.7[医药卫生—肿瘤]