Monocyte - neutrophil interplay drives mesenchymal transition in GBM

Myeloid cells comprise the majority of immune cells in tumors, where their content and composition is not only driver mutation-specific, but also tumor type-dependent. While these cells are essential for shaping the tumor microenvironment, promoting tumor growth, and contributing to therapeutic resistance, targeting tumor-associated myeloid cells, including bone-marrow-derived monocytes and neutrophils, has not been successful. To eliminate monocyte recruitment, we employed CRISPR/Cas9-based methods to delete the region on murine chromosome 11 harboring the monocyte chemoattractant protein family (MCP) genes, Ccl2, Ccl7, Ccl8, Ccl12, and Ccl11, which we termed qMCP-KO. Using these qMCP-knockout mice in combination with genetically engineered mouse models (GEMM) of glioblastoma (GBM), we investigated myeloid infiltraion by sc-RNA seq. We found that when monocyte infiltraion was abolisehd, a compensatory influx of nuetrophil in these tumor occured. These neutrophil promote tumor growth by releasing TNF, contributing to tumor hypoxic responses and aggression. Overall design: We induced de novo brain tumor in WT, qMCP-KO mice of both genders. We isolated brain tumor cells at the humane endstage of these mice and ran sc-RNA seq on these cells to compare their functional differences.

髓系细胞（Myeloid cells）是肿瘤免疫细胞的主要组分，其丰度与组成不仅与驱动突变类型相关，还具有肿瘤类型依赖性。尽管这类细胞对塑造肿瘤微环境、促进肿瘤生长及参与治疗耐药过程至关重要，但靶向肿瘤相关髓系细胞——包括骨髓源性单核细胞与中性粒细胞——的治疗策略迄今尚未取得成功。为阻断单核细胞募集，我们采用基于CRISPR/Cas9的基因编辑技术，敲除了小鼠11号染色体上携带单核细胞趋化蛋白（monocyte chemoattractant protein family, MCP）家族基因（Ccl2、Ccl7、Ccl8、Ccl12及Ccl11）的基因组区域，将该模型命名为qMCP-KO。我们将qMCP敲除小鼠与胶质母细胞瘤（glioblastoma, GBM）基因工程小鼠模型（genetically engineered mouse models, GEMM）相结合，通过单细胞RNA测序（single-cell RNA sequencing, sc-RNA seq）分析肿瘤内髓系细胞的浸润特征。研究结果显示，当单核细胞浸润被完全阻断时，肿瘤内会出现中性粒细胞的代偿性浸润。这类中性粒细胞通过释放肿瘤坏死因子（tumor necrosis factor, TNF）促进肿瘤生长，参与肿瘤的缺氧应答与侵袭进展。实验整体设计：我们在野生型（wild type, WT）以及两种性别的qMCP-KO小鼠中诱导新发脑肿瘤，待小鼠达到人道终点时分离脑肿瘤细胞，通过单细胞RNA测序对其进行检测，以比较两组小鼠肿瘤细胞的功能差异。