肿瘤相关巨噬细胞促进索拉非尼耐药肝癌细胞的增殖及侵袭和迁移  

Tumor-associated macrophages promote the proliferation and migration as well as invasion of sorafenib-resistant hepatocellular carcinoma cells

作　　者：魏续福[1] 蒲俊良 郭振[1] 李婷婷[1] 朱迪[1] 吴忠均[1] WEI Xufu PU Junliang GUO Zhen LI Tingting ZHU Di WU Zhongjun(Department of Hepatobiliary Surgery, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, China)

机构地区：[1]重庆医科大学附属第一医院肝胆外科,重庆400016

出　　处：《细胞与分子免疫学杂志》2017年第5期617-622,共6页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(81672959)

摘　　要：目的探讨肿瘤相关巨噬细胞(TAM)与表皮生长因子受体/β联蛋白(EGFR/β-catenin)信号通路在肝癌索拉非尼耐药中的机制。方法采用Ficoll密度梯度分离法获取外周血单个核细胞,M-CSF培养诱导成TAM,并对其进行表型鉴定,ELISA检测TAM分泌的表皮生长因子(EGF)的水平,CCK-8法检测EGF对HepG2和SMMC7721肝癌细胞增殖的影响,TranswellTM实验检测EGF对HepG2和SMMC7721肝癌细胞侵袭和迁移的影响。将TAM与肝癌细胞共培养探讨下游信号通路,采用药物敏感实验培养R-HepG2和R-SMMC7721索拉非尼耐药细胞株,Western blot法和免疫组织化学染色检测β-catenin、EGFR蛋白水平和表达情况。结果初步分离并鉴定TAM,与HepG2、SMMC7721细胞相比,培养获得索拉非尼耐药R-HepG2和R-SMMC7721细胞的增殖、侵袭、迁移能力明显增加;索拉非尼耐药细胞中加入TAM后,其生长率明显高于对照组,索拉非尼耐药细胞和肝癌组织标本中的β-catenin、EGFR的蛋白表达均高于对照组。结论 TAM与EGFR/β-catenin信号通路促进索拉非尼耐药肝癌细胞的增殖、侵袭和迁移。Objective To investigate the roles of tumor-associated macrophages（TAMs） and epithelial growth factor receptor（EGFR）/β-catenin signaling pathway in sorafenib resistance of hepatocel ular carcinoma cel s.Methods Macrophages were isolated from peripheral blood mononuclear cells（PBMCs） and cultured in the presence of colony-stimulating factor（M-CSF） to obtain TAMs.Phenotypes of TAMs were identified.The level of epithelial growth factor（EGF） secreted by TAMs was detected by ELISA.CCK-8 assay was performed to verify the effects of EGF on HepG2 and SMMC7721 cel proliferation.TranswellTMassay was used to examine the effects of EGF on the invasion and migration ability of HepG2 and SMMC7721 cells.TAMs and hepatocellular carcinoma cells were co-cultured to study the downstream signaling pathways.Sorafenib-resistant HepG2 and SMMC7721 strains（R-HepG2 and R-SMMC7721 cells） were prepared and then subjected to Western blotting and immunohistochemistry to examine the expression levels of β-catenin and EGFR.Results TAMs we prepared were confirmed.Compared with HepG2 and SMMC7721 cells,R-HepG2 and R-SMMC7721 showed enhanced proliferation,invasion and migration abilities.The growth rates of sorafenib-resistant cell lines after co-cultured with TAMs were significantly higher than those of the controls.The protein expressions of β-catenin and EGFR in sorafenib-resistant cells and hepatocellular carcinoma tissues were higher than those in the controls.Conclusion TAMs and EGFR/β-catenin signaling pathway promote the proliferation,invasion and migration of sorafenib resistance of hepatocellular carcinoma cells.

关 键 词：索拉非尼耐药 肿瘤相关巨噬细胞 Β-CATENIN信号通路 

分 类 号：R735.7[医药卫生—肿瘤] R730.3[医药卫生—临床医学]