Single-Cell Transcriptome Analysis Reveals Neutrophil Populations in Gastric Cancer

The full neutrophil heterogeneity and function remains incompletely characterized in gastric cancer progression. Here, we profiled >21,854 mouse neutrophils with an average of 1,189 genes per cell using single-cell RNA sequencing to provide a comprehensive transcriptional landscape of neutrophil function and fate decision in tumor progression. By combining scRNA-Seq analysis with tumor model, flowcytometry, genetic mice and clinical sample, our results define four neutrophils subpopulations in gastric cancer. Importantly, we also identified the critical transcription factor for the generation of effective activated neutrophils targeted for tumor cells. We performed scRNA-seq analysis with bone marrow neutrophils, spleen neutrophils, blood neutrophils and tumor associated neutrophils from tumor bearing mice for 10 days, and tumor associated neutrophils from tumor bearing mice for 3 days. We also performed scRNA-seq analysis with tumor infiltrated CD45+ immune cells from WT and KO mice. Next, we detected the presence of neutrophils subpopulations in human gastric cancer by scRNA-seq of sorted human tumor infiltrated CD45+ immune cells.

胃癌进展过程中，中性粒细胞的完整异质性与功能尚未得到完全阐明。本研究通过单细胞RNA测序（single-cell RNA sequencing, scRNA-seq）对超过21854个小鼠中性粒细胞进行分析，每个细胞平均检测到1189个基因，以此构建肿瘤进展过程中中性粒细胞功能与命运决定的全面转录图谱。本研究结合单细胞RNA测序分析、肿瘤模型、流式细胞术（flow cytometry）、基因工程小鼠及临床样本，最终明确了胃癌中的4个中性粒细胞亚群。尤为重要的是，本研究还鉴定出可介导靶向肿瘤细胞的功能性活化中性粒细胞生成的关键转录因子。我们分别对荷瘤10天的小鼠的骨髓中性粒细胞、脾脏中性粒细胞、血液中性粒细胞及肿瘤相关中性粒细胞（tumor associated neutrophils, TAN），以及荷瘤3天的小鼠的肿瘤相关中性粒细胞进行了单细胞RNA测序分析。我们还对野生型（wild type, WT）和基因敲除型（knockout, KO）小鼠的肿瘤浸润CD45+免疫细胞进行了单细胞RNA测序分析。随后，我们通过对分选得到的人肿瘤浸润CD45+免疫细胞进行单细胞RNA测序，验证了人胃癌中中性粒细胞亚群的存在。