小鼠原位移植肝癌模型脏器内中性粒细胞异质性分析  

Analysis of neutrophil heterogeneity in orthotopic mouse models of hepatocellular carcinoma

作　　者：贺改霞 冯俊 周颖 李文娟[1] 梁映雪 尚琪 张栩锐 李宗芳[1,2] HE Gaixia;FENG Jun;ZHOU Ying;LI Wenjuan;LIANG Yingxue;SHANG Qi;ZHANG Xurui;LI Zongfang(National-Local Joint Engineering Research Center of Biodiagnostics and Biotherapy,Second Affiliated Hospital of Xi'an Jiaotong University,Shaanxi Xi'an 710004,China;Shaanxi Provincial Clinical Research Center for Hepatic&Splenic Diseases,Shaanxi Xi'an 710004,China)

机构地区：[1]西安交通大学第二附属医院生物诊断治疗国家地方联合工程研究中心,陕西西安710004 [2]陕西省肝脾疾病临床医学研究中心,陕西西安710004

出　　处：《现代肿瘤医学》2024年第23期4425-4431,共7页Journal of Modern Oncology

基　　金：国家自然科学基金(编号:81802455);陕西省自然科学基础研究计划项目(编号:2019JQ-962);西安交大基础临床融合项目(编号:YXJLRH2022098);西安交大二附院自由探索项目[编号:2020YJ(ZYTS)546-03]。

摘　　要：目的:建立小鼠原位移植肝癌模型,深入分析肝癌负荷下小鼠骨髓、脾脏和肝脏内中性粒细胞的异质性。方法:选取16只健康雌性BALB/c小鼠,6~8周龄,随机分为肝癌模型组和对照组。模型组通过肝脏注射H22细胞系建立肿瘤,对照组注射PBS。接种后18~20天,收集两组小鼠的骨髓、脾脏及肝脏组织,分离免疫细胞,流式细胞术检测中性粒细胞的比例,并探讨肝癌负荷下中性粒细胞的异质性。根据CXCR2和CD101分子的表达,我们分析了荷瘤小鼠脏器内四个不同成熟状态的中性粒细胞亚群。结果:与对照组相比,肝癌组小鼠的骨髓、外周血、脾脏和肝脏内中性粒细胞比例均显著升高;进一步分析发现荷瘤小鼠脏器内存在四个不同成熟阶段的中性粒细胞亚群,包括CXCR2^(-)CD101^(+)、CXCR2^(+)CD101^(+)、CXCR2^(+)CD101^(-)和CXCR2^(-)CD101^(-)亚群,这一发现支持了中性粒细胞的异质性。具体来说,在肝癌小鼠骨髓中CXCR2^(-)CD101^(+)亚群和CXCR2-CD101^(-)未成熟亚群比例显著增加,而CXCR2^(+)CD101^(+)成熟亚群和CXCR2^(+)CD101^(-)低成熟亚群比例则显著减少。脾脏和肝脏内的粒细胞亚群整体变化趋势类似,即CXCR2^(-)CD101^(+)亚群、CXCR2^(+)CD101^(-)低成熟亚群和CXCR2^(-)CD101^(-)未成熟亚群的比例均增加,而CXCR2^(+)CD101^(+)成熟亚群的比例显著减少,提示外周脏器内未成熟或低成熟中性粒细胞增加,而成熟中性粒细胞减少。结论:本研究揭示了肝癌进展中,小鼠骨髓、脾脏和肝脏内中性粒细胞数量的显著增加及其亚群比例的显著变化,为理解肝癌进展中中性粒细胞的表型异质性提供重要依据,并为进一步阐明中性粒细胞参与肝癌进展的机制提供了新的视角。Objective:To investigate the heterogeneity of neutrophils in bone marrow,peripheral blood,spleen,and liver of mouse model with orthotopic hepatocellular carcinoma(HCC).Methods:Sixteen healthy female BALB/c mice,aged at 6~8 weeks,were randomly divided into the orthotopic tumor-bearing group and control group.The tumor model was established by orthotopically injection of H22 cell line into the liver,while the control group received PBS injection as a sham surgery control.At 18~20 days post-inoculation,bone marrow,spleen,and liver tissues were collected from both groups of mice.Immune cells were isolated,and the proportion of neutrophils was detected by flow cytometry.The heterogeneity of neutrophilsubsets in HCC-bearing mice was examined based on the expression of CXCR2 and CD101 molecules.Results:Compared to the control group,the proportion of neutrophils significantly increased in the bone marrow(BM),peripheral blood,spleen,and liver of tumor-bearing mice.Further analysis revealed the coexistence of neutrophil subpopulations at different stages of maturation within these organs in HCC mice,including CXCR2^(-)CD101^(+),CXCR2^(+)CD101^(+),CXCR2^(+)CD101^(-),and CXCR2^(-)CD101^(-)subpopulations,supporting the heterogeneity of neutrophils.Specifically,in the BM of HCC mice,there was a marked elevation in the proportions of the CXCR2^(-)CD101^(+)subset and the CXCR2^(-)CD101^(-)immature subset,while the CXCR2^(+)CD101^(+)mature subset and the CXCR2^(+)CD101^(-)low maturity subset decrease significantly.A similar pattern was observed in the spleen and liver of tumor mice,with increased proportions of the CXCR2^(-)CD101^(+)subset,the CXCR2^(+)CD101^(-)low maturity subset,the CXCR2^(-)CD101^(-)immature subset,and decreased proportion of mature CXCR2^(+)CD101^(+)neutrophils,indicating an overall shift towards an increase in immature or less mature neutrophils and a decrease in mature neutrophils in peripheral organs.Conclusion:This study demonstrates a significant elevation in the number of neutrophils and notable alterat

关 键 词：中性粒细胞 肿瘤相关中性粒细胞 异质性 肝细胞癌 原位瘤模型 

分 类 号：R735.7[医药卫生—肿瘤]