微波消融对肺癌肿瘤微环境的抗肿瘤作用及其机制
Anti-tumor effect and mechanism of microwave ablation on lung cancer tumor microenvironment
武少贤 1吴悦 1房章 1伍悠 1焦静 1陈陆俊 1郑晓 1蒋敬庭1
作者信息
- 1. 苏州大学附属第三医院肿瘤生物诊疗中心,江苏省肿瘤免疫治疗工程技术研究中心,苏州大学细胞治疗研究院,常州 213003
- 折叠
摘要
目的 探讨微波消融(MWA)对肺癌抗肿瘤的影响与作用机制.方法 将1×106肺癌细胞系3LL皮下注射到6～8周的雄性C57BL/6J小鼠背部两侧,当肿瘤最大径长到7 mm时,随机分为对照组和MWA组,对一侧肿瘤进行MWA处理,每隔1 d监测对侧肿瘤生长.在MWA后的第8天,运用流式细胞术对肿瘤微环境(TME)中免疫细胞群体比例及功能进行分析.在MWA后的1 d腹腔注射程序性死亡受体1(PD-1)或者细胞毒性T淋巴细胞相关蛋白4(CTLA-4)抑制剂(200µg/只),每隔1d监测对侧肿瘤生长.采用独立样本t检验比较两组间差异.使用双向方差分析来比较肿瘤生长曲线.结果 与对照组比较,MWA有效抑制小鼠对侧肿瘤生长,差异有统计学意义(780.70±239.90 比 1 794.00±195.20,t=3.275,P＜0.01).MWA 组小鼠对侧 TME 中 CD3+T 细胞(79.02±3.61 比 62.22±3.59,t=3.300,P＜0.05)、CD4+T 细胞(33.26±1.73 比 25.80± 1.68,t=2.686,P＜0.05)和 CD8+T 细胞(26.26±1.01 比 21.64±1.40,t=3.096,P＜0.05)比例显著高于对照组.MWA组小鼠肿瘤浸润Ly6G+MDSC细胞群体比例(33.00±3.25比44.86±1.46,t=3.327,P＜0.05)显著低于对照组.对CD4+T细胞的功能以及增殖分析表明,MWA组小鼠γ-干扰素(IFN-γ)+CD4+T 细胞(20.39±1.84 比 3.05±0.47,t=7.827,P＜0.01)、TNF-α+CD4+T 细胞(17.25±2.64 比 5.61±1.11,t=3.571,P＜0.05)和细胞核增殖抗原(Ki-67)+CD4+T 细胞(90.68±0.81比81.55±1.37,t=6.108,P＜0.01)比例显著高于对照组.此外,MWA组小鼠IFN-γ+CD8+T 细胞(37.46±5.09 比 6.84±1.52,t=4.973,P＜0.01)和 Ki-67+IFN-γ+CD8+T 细胞(35.75±4.62比6.55±1.48,t=5.204,P＜0.01)比例显著高于对照组.对T细胞表面免疫检查点分子表达分析表明,MWA组小鼠T细胞免疫球蛋白及黏蛋白结构域分子3(TIM-3)+CD8+T细胞(25.97±2.30 比 15.57±2.33,t=3.142,P＜0.05)和 PD-1+TIM-3+CD8+T 细胞(22.58±2.32 比12.77±2.81,t=2.617,P＜0.05)比例显著高于对照组.与MWA单独治疗组比较,将MWA与PD-1(514.50±106.40 比 883.60±144.80,t=1.911,P＜0.05)或 CTLA-4(534.40±82.06 比 961.90± 131.00,t=2.941,P＜0.05)抑制剂联用显著抑制小鼠对侧肿瘤生长.结论 MWA能够有效抑制肺癌肿瘤生长,将MWA联合PD-1抑制剂或者CTLA-4抑制剂产生更强抗肿瘤效果.
Abstract
Objective To explore the anti-tumor effect and mechanism of microwave ablation(MWA)on lung cancer.Methods Totally,1 × 106 3LL cells(lung cancer cell line)were subcutaneously injected into both sides of the back of 6-8 week old male C57BL/6J mice.When the maximum diameter of the tumor reached 7 mm,the animals were randomly divided into the control group and MWA group.The tumor on one side was treated with MWA,and the growth of the contralateral tumor was monitored every 1 day.On the 8th day after MWA,flow cytometry was used to analyze the proportion and function of immune cell populations in the tumor microenvironment(TME).After intraperitoneal injection of programmed cell death protein 1(PD-1)or cytotoxic T lymphocyte-associated antigen-4(CTLA-4)inhibitor(200 µg/mouse),the contralateral tumor growth was monitored every 1 day.The independent samples t-test was used to compare the differences between the two groups.Tumor growth curves were compared using two-way ANOVA.Results Compared with the control group,MWA effectively inhibited the growth of contralateral tumors in mice,and the difference was statistically significant(780.70±239.90 vs.1 794.00±195.20,t=3.275,P＜0.01).The proportion of CD3+T cells(79.02±3.61 vs.62.22±3.59,t=3.300,P＜0.05),CD4+T cells(33.26±1.73 vs.25.80±1.68,t=2.686,P＜0.05)and CD8+T cells(26.26± 1.01 vs.21.64±1.40,t=3.096,P＜0.05)in the contralateral TME of the MWA group mice was signif-icantly higher than that in the control group.The proportion of tumor-infiltrating Ly6G+MDSC cell popula-tion(33.00±3.25 vs.44.86±1.46,t=3.327,P＜0.05)in the MWA group mice was significantly lower than that in the control group.The functional and proliferative analysis of CD4+T cells in the MWA group mice revealed a significantly higher proportion of interferon-γ(IFN-γ)+CD4+T cells(20.39±1.84 vs.3.05±0.47,t=7.827,P＜0.01),tumor necrosis factor-α(TNF-α)+CD4+T cells(17.25±2.64 vs.5.61±1.11,t=3.571,P＜0.05)and proliferation cell nuclear antigen(Ki-67)+CD4+T cells(90.68±0.81 vs.81.55±1.37,t=6.108,P＜0.01)than the control group.In addition,the propor-tion of IFN-γ+CD8+T cells(37.46±5.09 vs.6.84±1.52,t=4.973,P＜0.01)and Ki-67+IFN-γ+CD8+T cells(35.75±4.62 vs.6.55±1.48,t=5.204,P＜0.01)in the MWA group mice was signifi-cantly higher than that in the control group.Analysis of immune checkpoint molecule expression on the sur-face of T cells showed that compared with the control group,the proportion of T cell immunoglobulin and mucin domain-containing protein 3(TIM-3)+CD8+T cells(25.97±2.31 vs.15.57±2.33,t=3.142,P＜0.05)and PD-1+TIM-3+CD8+T cells(22.58±2.32 vs.12.77±2.81,t=2.617,P＜0.05)in the MWA group mice was significantly increased in the control group.Compared with the simple MWA treat-ment group,the combination of MWA and PD-1 inhibitors(514.50±106.40 vs.883.60±144.80,t=1.911,P＜0.05)or CTLA-4 inhibitors(534.40±82.06 vs.961.90±131.00,t=2.941,P＜0.05)significantly inhibited the growth of contralateral tumors,and the difference was statistically significant.Conclusion MWA can effectively inhibit the growth of contralateral lung cancer tumors,and combined use of MWA with PD-1 or CTLA-4 inhibitors produces a more powerful anti-tumor effect.
关键词
微波消融/肺癌/肿瘤微环境/免疫细胞/程序性死亡受体1/细胞毒性T淋巴细胞相关蛋白4Key words
Microwave ablation/Lung cancer/Tumor microenvironment/Immune cells/Programmed cell death protein 1/Cytotoxic T lymphocyte-associated antigen-4引用本文复制引用
基金项目
国家自然科学基金(82303164)
国家自然科学基金(32270955)
江苏省卓越博士后计划(2023ZB512)
江苏省重点研发计划(BE2022721)
江苏省自然科学基金(BK20211065)
江苏省医学重点学科建设项目(YXZDXK202236)
常州市科技支撑计划社会发展项目(CE20235058)
出版年
2024
NETL
NSTL
万方数据