肿瘤相关巨噬细胞对乳腺癌MCF-7细胞侵袭迁移能力的影响  

Influence of tumor-associated macrophages on invasion and migration of MCF-7 cells

作　　者：李旭 黄新宇 赵琳[3] 吴芳[4] 王玉珍 廖子君 安改丽[5] LI Xu;HUANG Xinyu;ZHAO Lin;WU Fang;WANG Yuzhen;LIAO Zijun;AN Gaili(Department of Oncology, Shanxi Provincial Cancer Hospital, Xi'an 710061, China)

机构地区：[1]陕西省肿瘤医院肿瘤内科,西安710061 [2]陕西富平县医院内五科,711700 [3]西安交通大学第一附属医院肿瘤内科,710061 [4]西安交通大学附属第一医院儿科,710061 [5]陕西省人民医院肿瘤内科,710068

出　　处：《临床肿瘤学杂志》2022年第1期14-20,共7页Chinese Clinical Oncology

基　　金：西安市科技计划项目[20YXYJ0005(4)];陕西省自然科学基础研究计划项目(2020JM-674);陕西省肿瘤精准医学重点实验室开放课题(KLTPM-SX2017-C3);陕西省人民医院科技人才支持计划项目(2021JY-11)。

摘　　要：目的观察肿瘤相关巨噬细胞(tumor-associated macrophages,TAMs)对乳腺癌MCF-7细胞侵袭迁移能力的影响,并初步探索其机制。方法选取人单核细胞系THP-1,体外经佛波酯(PMA)、人白细胞介素-4(IL-4)诱导获得TAMs细胞模型;通过流式细胞术(FCM)检测TAMs表面标记分子CD206表达水平;MCF-7细胞与TAMs共培养后,光学倒置显微镜观察细胞形态;利用Transwell小室分别检测MCF-7细胞的侵袭和迁移能力;蛋白印记法(Western blotting)检测E-钙黏蛋白(E-cadherin)、N-钙黏蛋白(N-cadherin)、闭合蛋白(Occludin)及波形纤维蛋白(Vimentin)的表达;采用酶联免疫吸附法(ELISA)测定细胞培养上清中转化生长因子-β(TGF-β)、肿瘤坏死因子-α(TNF-α)和血管内皮生长因子(VEGF)的浓度。结果与TAMs共培养后的MCF-7细胞伪足增多,细胞排列更松散。通过FCM检测到TAMs表面标记物CD206显著表达。Transwell实验结果显示,与TAMs共培养后的MCF-7细胞迁移能力增强,对照组穿过Transwell小室细胞数为(54±2)个,实验组为(110±6)个,差异有统计学意义(P<0.001);其侵袭能力显著增强,对照组穿透基质膜的细胞数为(41±1)个,实验组为(77±4)个,差异亦有统计学意义(P<0.001)。Western blotting检测结果显示,MCF-7细胞与TAMs共培养后,E-cadherin、Occludin蛋白表达较对照组显著降低,而N-cadherin和Vimentin蛋白表达量显著升高(P均<0.05)。ELISA法检测TAMs与MCF-7细胞共培养上清中的TGF-β、TNF-α、VEGF浓度均较对照组升高(P均<0.05)。结论TAMs与乳腺癌MCF-7细胞的浸润转移过程密切相关,TAMs可通过促进MCF-7细胞发生上皮间质转化(EMT)进而促进乳腺癌的浸润转移。Objective To investigate the influence of tumor-associated macrophages(TAMs)on invasion and migration of MCF-7 cells,and explore its preliminary mechanism.Methods Human mononuclear cell line THP-1 was induced by phorbol ester(PMA)and interleukin-4(IL-4)to obtain TAMs cell model in vitro.The TAMs surface marker molecule CD206 was detected by flow cytometry(FCM).After co-cultured MCF-7 cells with TAMs,the cell morphology was observed with an optical inverted microscope.The invasion and migration abilities of MCF-7 cells were assessed by transwell assay.Western blotting was performed to detect the expression levels of E-cadherin,N-cadherin,Occludin and Vimentin.Enzyme-linked immunosorbent assay(ELISA)was used to determine the concentrations of TGF-β,TNF-αand VEGF in cell culture supernatant.Results After co-cultured with TAMs,MCF-7 cells had more pseudopods and more looser cell arrangement.Significant expression of the TAMs surface marker CD206 was detected by FCM.Transwell assay showed that the migration ability of MCF-7 cells was enhanced after co-cultured with TAMs.The number of cells passing through the transwell chamber was(54±2)cells in the control group,and(110±6)cells in the experimental group,and the difference between the two groups was statistical significance(P<0.001).The invasive ability of MCF-7 cells was significantly enhanced meanwhile,the number of cells penetrating the stromal membrane in the control group was(41±1)cells,and in the experimental group was(77±4)cells,and the difference was statistically significant(P<0.001).Western blotting assay showed that after co-cultured MCF-7 cells with TAMs,the expression levels of E-cadherin and Occludin were down-regulated compared with the control group,while the expression levels of N-cadherin and Vimentin were up-regulated significantly(P<0.05).ELISA results showed that the TGF-β,TNF-αand VEGF levels in the co-culture group were significantly higher than those in the control group(P<0.05).Conclusion TAMs can promote the invasion and migration abil

关 键 词：乳腺癌 肿瘤相关巨噬细胞 侵袭迁移 上皮-间质转化 

分 类 号：R737.9[医药卫生—肿瘤]