乳腺癌4T1细胞中过表达鼠白介素-10对宿主抗肿瘤免疫应答的影响  

Effects of mouse interleukin-10 overexpression in breast cancer 4T1 cells on host anti-tumor immune response

作　　者：王晓倩 李丹丹[2] 王丹[2] 王孟影 贺龙梅 王晓琴[1] 马运峰 WANG Xiaoqian;LI Dandan;WANG Dan;WANG Mengying;HE Longmei;WANG Xiaoqin;MA Yunfeng(Clinical Laboratory of First Affiliated Hospital,Xi’an Jiaotong University,Xi’an 710061,China;Department of Pathogenic Microbiology and Immunology,School of Basic Medical Sciences,Xi’an Jiaotong University;Clinical Laboratory of Shanxi Traditional Chinese Medicine Hospital;Institute of Infection and Immunity,Xi’an Jiaotong University Translational Medicine Center)

机构地区：[1]西安交通大学第一附属医院检验科,西安710061 [2]西安交通大学基础医学院病原生物学与免疫学系 [3]陕西省中医医院检验科 [4]西安交通大学转化医学研究院感染免疫研究所

出　　处：《山西医科大学学报》2021年第5期559-564,共6页Journal of Shanxi Medical University

基　　金：国家自然科学基金面上项目(81872026,81472822);陕西省自然基金面上项目(2020JM-022)。

摘　　要：目的研究鼠白介素-10(mIL-10)对BALB/c小鼠三阴性乳腺癌4T1细胞生长以及荷瘤小鼠抗肿瘤免疫应答的影响。方法将重组质粒pLenti-CMV-mIL-10-GFP-puro以及pLenti-CMV-GFP-puro分别与慢病毒包装质粒共转染293T细胞,收集病毒上清感染4T1细胞,从而获得过表达mIL-10的细胞株即4T1/mIL-10(实验组)以及4T1/GFP(对照组);酶联免疫吸附试验(enzyme linked immunosorbent assay,ELISA)以及流式细胞术(flow cytometery,FCM)检测感染效率及细胞中mIL-10的表达水平;细胞计数检测细胞的增殖能力;应用FCM检测细胞表面CD86、主要组织相容性复合体Ⅱ类抗原(MHCⅡ)、程序性死亡配体1(programmed cell death ligand 1,PDL1)的表达。将4T1/GFP、4T1/mIL-10细胞分别皮下注射至BALB/c小鼠,建立小鼠乳腺癌皮下移植瘤模型即未治疗组和mIL-10治疗组,观察肿瘤生长情况,绘制生长曲线,并使用流式细胞仪检测肿瘤组织中T细胞的变化。结果成功构建过表达mIL-10的4T1细胞,ELISA以及FCM检测结果表明,与对照组比较,实验组的mIL-10表达水平显著增加(P<0.05)。与对照组比较,实验组细胞的增殖以及表面分子CD86、MHCⅡ分子、PDL1的表达无显著差异(P>0.05)。动物实验模型表明,与未治疗组比较,mIL-10治疗组肿瘤组织中CD8^(+)T/CD4^(+)T的比例以及CD8^(+)T细胞IFN-γ的表达水平明显增加,肿瘤生长受到抑制(P<0.05)。结论mIL-10能够促进宿主抗肿瘤免疫应答,从而抑制乳腺癌的生长。Objective To investigate the effect of mouse interleukin-10(mIL-10)on the growth of BALB/c mouse triple negative breast cancer 4T1 cells and host anti-tumor immune response.Methods The 293T cells were co-transfected with recombinant plasmids plenti-CMV-IL-10-GFP-Puro or plenti-CMV-GFP-Puro and lentiviral packing plasmids,,and then the viral supernatants were harvested.4T1 cells were infected by viral supernatants to obtain cell lines with overexpression of mIL-10,namely 4T1/mIL-10(experimental group)and 4T1/GFP(control group).The infection efficiency and the expression of mIL-10 in the cells were detected by enzyme linked immunosorbent assay(ELISA)and flow cytometry(FCM).The cell proliferation was detected by cell count.The expression levels of CD86,MHCⅡ,and programmed death ligand 1(PDL1)on cell surface were determined by FCM.4T1/GFP and 4T1/mIL-10 cells were subcutaneously injected into BALB/c mice to establish mouse breast cancer tumor models in untreated group and mIL-10 group,respectively.The tumor size was measured and the T cell infiltration in tumor tissues was detected by FCM.Results The 4T1 cells overexpressing mIL-10 were successfully constructed.ELISA and FCM assay results showed that the expression of mIL-10 in experimental group was significantly increased compared with control group(P<0.05).There was no significant difference in cell proliferation and the expression levels of CD86,MHCⅡand PDL1 between experimental group and control group(P>0.05).Compared with untreated group,the ratio of CD8^(+)T/CD4^(+)T and the expression of IFN-γin CD8^(+)T cells were significantly increased in mIL-10 group(P<0.05),while the tumor growth was inhibited(P<0.05).Conclusion The mIL-10 can promote the host anti-tumor immune response and inhibit the growth of breast cancer.

关 键 词：三阴性乳腺癌 mIL-10 4T1细胞 

分 类 号：R737.9[医药卫生—肿瘤]