基于单细胞转录组构建结直肠癌肿瘤相关巨噬细胞图谱

A SINGLE-CELL ATLAS OF TUMOR -ASSOCIATED MACROPHAGE IN COLORECTAL CANCER

颜培东

11849362

硕士

生物学

靳文菲

2020-06-05

2020-06-05

哈尔滨工业大学

深圳

肿瘤相关巨噬细胞是浸润于肿瘤组织中的一类巨噬细胞，也是肿瘤微环境中最多的免疫细胞。在结直肠癌的环境中，巨噬细胞不仅浸润于肿瘤微环境中，也暴露于肠道的复杂环境中，表现出极大的异质性。我们通过单细胞转录组测序技术，解析结直肠癌微环境中肿瘤相关巨噬细胞的图谱，以研究巨噬细胞的异质性。我们通过AOM/DSS诱导ApcMin杂合子小鼠，成功构建了小鼠炎症相关的结直肠癌肿瘤模型。我们通过酶消化法，制备不同组织的单细胞悬液。我们首先对小鼠肿瘤和癌旁组织进行单细胞测序，后利用流式细胞术分选出肿瘤中CD11b+髓系细胞，再进行单细胞转录组测序，共得到六组数据。通过对六组数据的分别分析以及合并分析，研究其基因表达特征，通过可视化方法分析，展示了细胞簇之间的异质性，并发现巨噬细胞簇主要来源于单核细胞。我们还表征了两个CX3CR1+的巨噬细胞细胞亚群，其表达特征与单核细胞来源的结肠巨噬细胞的极化特征相似。分析表明许多巨噬细胞簇同时表达M1型和M2型巨噬细胞的基因标记，处于M1/M2细胞型的中间状态。按照Vegfa的表达水平可以将巨噬细胞簇为高、中、低三类。Vegfa表达高的细胞簇除了表达巨噬细胞簇的特征分子也表达 TLRs、NLRs等炎症相关模式识别受体和NF-κB信号通路相关基因。一些细胞簇同时表达炎症小体和NF-κB负调节基因，共表达T细胞共刺激因子和T细胞免疫抑制因子。综上，通过单细胞转录组测序技术，我们构建了结直肠癌的肿瘤相关巨噬细胞图谱，为理解巨噬细胞在肿瘤微环境中的分布以及创建相关免疫疗法提供了参考。

Tumor-associated macrophages are a type of macrophages that infiltrate tumor tissues and are the most immune cells in the tumor microenvironment. In the environment of colorectal cancer, macrophages not only infiltrated the tumor microenvironment, but also exposed to the complex environment of the intestine, showing great heterogeneity. We induced ApcMin heterozygous mice through AOM/DSS and successfully established a mouse-related connective cancer tumor model. After obtaining the tumor, we prepared a single cell suspension by enzyme digestion. First, single-cell lesions were performed on mouse tumors and tissues adjacent to cancer, and then flow cytometry was used to sort out CD11b + cells in the tumors, and then single-cell transcriptome selection was performed. We got a total of six sets of data. Through separate analysis and combined analysis of the six sets of data, the gene expression characteristics are analyzed, and through a series of visual analysis, the heterogeneity between different cell clusters is analyzed. We found that these cell clusters basically arranged monocytes, and we also characterized two CX3CR1 + cell groups, whose expression characteristics are similar to the mutation characteristics of monocyte-derived colonic macrophages. We also found that many macrophage clusters express the characteristics of M1 and M2 macrophages at the same time, which is in the intermediate state of M2/M2 cell type. Based on the expression of Vegfa, we clustered macrophages into high, medium and low categories. Vegfa-expressed cell clusters are highly expressed while macrophage clusters have phenotypic characteristics. TLR, NLR and other metabolic related genes recognize receptors and NF-κB signaling pathway related genes through pattern recognition. Some cell clusters also express metabolic bodies and NF-κB negative regulatory genes, and some macrophage co-express T cell costimulatory factors and T cell immunosuppressive factors. Through single cell transcriptome targeting technology, we synthesized the tumor-associated macrophage map of colorectal cancer, which provides a reference for understanding the distribution of macrophages in the tumor microenvironment and related immunotherapy.

肿瘤相关巨噬细胞 结直肠癌 单细胞转录组测序 巨噬细胞异质性

tumor-associated macrophages colorectal cancer single-cell transcriptome sequencing macrophage heterogeneity

中文

联合培养

南方科技大学

颜培东. 基于单细胞转录组构建结直肠癌肿瘤相关巨噬细胞图谱[D]. 深圳. 哈尔滨工业大学,2020.