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

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(Ecadherin)、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...

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

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 ability of MCF-7 cells by inducing the occurrence of epithelial-mesenchymal transition(EMT).