中文版 | English
题名

CD11bint F4/80hi肿瘤相关巨噬细胞在小鼠肝转移瘤内分化路径及功能解析

其他题名
Differentiation and functional analysis of CD11bint F4/80hi tumor-associated macrophage in mouse liver metastasis
姓名
姓名拼音
QIAO Ting
学号
12032633
学位类型
硕士
学位专业
071009 细胞生物学
学科门类/专业学位类别
07 理学
导师
许扬,任欢
导师单位
南方科技大学医学院;南方科技大学医学院
论文答辩日期
2023-05-12
论文提交日期
2023-07-01
学位授予单位
南方科技大学
学位授予地点
深圳
摘要

研究背景及目的:肝脏是结直肠癌最常见的远处转移器官,由于转移瘤快速进展及耐药性等因素,目前的治疗手段获益有限,对于结直肠癌肝转移(colorectal liver metastasesCRLM)仍需找出更为有效的治疗策略。 近年来免疫疗法在多种癌症中取到了显著疗效,可能是 CRLM 治疗的新希望 。 CRLM 肿瘤微环境中 的肿瘤相关巨噬细胞( tumor associated macrophageTAM)具有高度异质性,在肿瘤进展中起重要作用。我们通过建立 CRLM 小鼠模型对 CRLM TAM 的分化途径及功能进行分析研究,探索其促进肿瘤进展新机制,以期发现靶向巨噬细胞新靶点和治疗方案。

实验方法:通过经脾注射 MC38 细胞构建免疫适能 C57BL/6 小鼠 CRLM 模型,模拟结直肠癌晚期患者癌细胞经门静脉进入肝脏并发生肝转移的过程。通过多次建模确定 CRLM 肝内生长模式及疾病分期。分别选取不同肿瘤微环境及免疫细胞或基质细胞等组织和细胞样本,使用批量转录组测序(bulk RNA sequencingbulk RNA-seq)对系列 CRLM 或细胞样本进行测序。基于获得的 bulk RNA-seq 数据,以生物信息数据分析软件 ImmuCCmMCP-counter 等对免疫细胞浸润、肿瘤纯度和基质细胞评分等进行分析计算;联合流式细胞术分析 CRLM 微环境中的免疫细胞的组成及占比等。进一步通过提取 CRLM 组织、肝脏或骨髓原代细胞包括骨髓来源单核细胞、枯否细胞、F4/80+ TAM(肿瘤相关巨噬细胞)等进行分离和体外培养;体外与肿瘤细胞或其上清共培养等。并通过实时荧光定量 PCR 等技术明确相关基因表达;以及肿瘤血管生成基因表达模式相关的生信分析等,以明确CRLM 内单核巨噬细胞细胞分化途径及分析 TAM 促进血管生成的功能等。

实验结果:TAM 是快速进展的小鼠 CRLM 免疫微环境中逐渐增多和最丰富的免疫细胞,并随 CRLM 进展表型发生变化。Ly6C+ 骨髓单核细胞是TAM 重要来源。我们发现 CD11bhi Ly6C+的单核细胞被募集至 CRLM 微环境后获得驻留巨噬细胞标记 F4/80 表达,在肿瘤微环境中转变为 CD11bint F4/80hi Ly6C- 的成熟 TAM 亚型。通过生物信息软件分析比较 CRLM 中异质性的 CD11b+ TAM F4/80+ TAM bulk RNA-seq 数据发现,该两群巨噬细胞在来源及功能上虽有重叠但也有显著差异,相对于 CD11b+ TAMF4/80+ TAM 在器官形成和促血管生成方面显著激活,而 CD11b+ TAM 则显示髓系细胞去分化和细胞增殖等显著特征。生物信息学分析显示,两类TAM 均显示独特于肝脏血管、肿瘤组织血管的血管生成相关的表达模式。 我们进一步在体外共培养体系中证实,F4/80+ TAM 可显著促进血管生成关键基因例如 CD34CD31VEGFTIE2 等的激活。

结论:TAM CRLM 免疫微环境中丰度最高的免疫细胞,在 CRLM 进展过程中起到重要作用且来源及功能存在异质性,F4/80+ TAM 显著促进肿瘤血管生成。

 

关键词
语种
中文
培养类别
独立培养
入学年份
2020
学位授予年份
2023-06
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生物学
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条目标识符http://sustech.caswiz.com/handle/2SGJ60CL/544727
专题南方科技大学医学院
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谯婷. CD11bint F4/80hi肿瘤相关巨噬细胞在小鼠肝转移瘤内分化路径及功能解析[D]. 深圳. 南方科技大学,2023.
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