肿瘤相关成纤维细胞对肝癌干细胞的影响  

Effect of Cancer-associated Fibroblasts on Cancer Stem Cells in Hepatocellular Carcinoma

作　　者：冯仁鑫 陈升[1] 刘娇[1] 宁北芳[1] 谢渭芬[1] FENG Renxin CHEN Sheng LIU Jiao NING Beifang XIE Weifen.(Department of Gastroenterology, Changzheng Hospital, the Second Military Medical University, Shanghai （200003)

机构地区：[1]第二军医大学附属长征医院消化内科,200003

出　　处：《胃肠病学》2016年第9期517-522,共6页Chinese Journal of Gastroenterology

基　　金：国家自然科学基金(81572412)

摘　　要：背景:肿瘤相关成纤维细胞(CAF)是肿瘤微环境中一类特殊的间质细胞,研究显示CAF在一些恶性肿瘤中可调节肿瘤干细胞(CSC)的干细胞特性和致瘤性。然而在肝细胞癌(HCC)中,CAF对CSC的调控作用尚不清楚。目的:探讨CAF对肝癌干细胞的影响。方法:分别采用免疫组化和免疫荧光技术标记CD44阳性CSC和α-SMA阳性CAF,观察两者在人HCC标本中的位置关系。从HCC手术标本中分离、纯化CAF和癌旁成纤维细胞(PTF),收集相应条件培养基和干细胞条件培养基。以real-time PCR检测CAF或PTF条件培养基培养的人肝癌细胞株HuH-7中的肝癌干细胞相关基因表达。采用流式细胞术分选肝癌干细胞表面标记物EpC AM阳性HuH-7细胞,以CAF或PTF干细胞条件培养基培养细胞,行干细胞成球实验。结果:在HCC标本中,CAF与CSC位置紧邻。与PTF组和对照组HuH-7细胞相比,CAF条件培养基处理组HuH-7细胞肝癌干细胞表面标记物基因和自我更新相关基因表达显著上调(P均<0.05)。CAF干细胞条件培养基处理组未经分选的HuH-7细胞和EpC AM阳性HuH-7细胞,成球率分别显著高于相应PTF组和对照组细胞(19.15%、32.13%对10.89%、19.57%和12.57%、19.77%,P均<0.001)。结论:CAF在肝癌干细胞自我更新能力的维持和促进中起重要作用。Cancer-associated fibroblasts （CAF） are a class of specific mesenchymal cells in tumor microcnvironment. It has been demonstrated that CAF might regulate the sternness and tumorigenicity of cancer stem cells （CSC） in some malignancies, however, the regulatory effect of CAF on CSC in hepatocellular carcinoma （HCC） is not clear yet. Aims ： To explore the effect of CAF on CSC in HCC. Methods： CD44-positive CSC and ct-SMA-positive CAF were labeled by immunohistochemistry and immunofluorescence, respectively, for localization of CSC and CAF in human HCC tissue. CAF and peritumoral fibroblasts （PTF） were isolated and purified from HCC surgical specimens, and the respective conditioned medium （CM） and stemness CM were collected. Human HCC cell line HUH-7 was cuhured with CAF- or PTF-CM, and the expression of hepatic CSC-related genes in HUH-7 cells was evaluated by real-time PCR. HUH-7 cells positive for EpCAM, a cell surface hepatic CSC marker, were obtained by fluorescence-activated cell sorting, and cultured with CAF- or PTF-stemness CM for spheroid formation. Results： CAF was adjacent to CSC in HCC tissue. When compared with PTF- CM and controlled CM, CAF-CM up-regulated the expression of hepatic CSC surface marker genes and self-renewal-related genes in HUH-7 cells （ P all 〈 0.05 ）. The spheroid formation rates of unsorted HUH-7 cells and EpCAM-positive HUH-7 cells in CAF-stemness CM were significantly higher than those in PTF-stemness CM and controlled sternness CM （19.15%, 32.13% vs. 10.89%, 19.57% and 12.57%, 19.77%, P all 〈0.001）. Conclusions： CAF play important roles in the maintenance and enhancement of self-renewal capability of hepatic CSC.

关 键 词：癌 肝细胞 肿瘤相关成纤维细胞 肿瘤干细胞 自我更新 球形体 细胞 

分 类 号：R735.7[医药卫生—肿瘤]