单细胞 RNA 测序揭示胶质母细胞瘤的肿瘤内异质性和肿瘤相关巨噬细胞的促肿瘤亚群，其特征是 EZH2 过表达。

Single-cell RNA sequencing reveals intra-tumoral heterogeneity of glioblastoma and a pro-tumor subset of tumor-associated macrophages characterized by EZH2 overexpression.

机构信息

Department of Neurosurgery, Neurosurgery Research Institute, The First Affiliated Hospital, Fujian Medical University, Fuzhou, Fujian, China.

Department of Neurosurgery, the Second Affiliated Hospital of Fujian Medical University, Quanzhou, China.

出版信息

Biochim Biophys Acta Mol Basis Dis. 2022 Dec 1;1868(12):166534. doi: 10.1016/j.bbadis.2022.166534. Epub 2022 Aug 31.

DOI:10.1016/j.bbadis.2022.166534

PMID:36057370

Abstract

BACKGROUND

Glioblastoma (GBM) is a highly heterogeneous disease with poor clinical outcome.

AIM

To comprehensively dissect molecular landscape of GBM and heterogeneous distribution and potential role of Enhancer of zeste homolog 2 (EZH2) in tumor microenvironment (TME).

METHODS

Single-cell RNA sequencing (scRNA-seq) analysis was performed in GBM samples from 8 patients. Deconvolution analysis, immunofluorescence (IF) microscopy, reverse-transcription quantitative polymerase chain reaction (RT-qPCR), colony formation experiments, and Cell Counting Kit-8 (CCK-8) assays were performed to confirmed the potential role of EZH2 in TME cells.

RESULTS

Malignant cells exhibited remarkable heterogeneity in abnormal metabolic patterns. A mesenchymal-2-like (MES2-like) GBM subcluster with glial-immune dual feature was firstly discovered, which were associated with highly activated hallmark pathways, immune evasion associated transcription factor (IRF8), and poor survival. The oncogene, EZH2, was heterogeneously expressed in malignant cells and immune cells consistent with proliferative genes, cell-cycle transcription factors, and similar activated hallmark pathways. In a tumor-associated macrophages (TAMs) subset (macrophage.3), EZH2 was highly expressed with similar changes of transcriptomic dynamics with cell-cycle genes and macrophages M2-phetotype genes. In addition, the subset tightly interacted with malignant cells. Deconvolution analysis showed increased abundance of the subset in GBM compared to low-grade glioma (LGG) and significant association with worse prognosis. Functional verification experiments confirmed the pro-tumor role of TAMs with EZH2 overexpression in GBM.

CONCLUSIONS

Our study illustrated a MES2-like GBM subcluster characterized by glial-immune dual feature and highlighted the pro-tumor role of a TAMs subset characterized by EZH2 overexpression.

摘要

背景

胶质母细胞瘤（GBM）是一种高度异质性疾病，临床预后较差。

目的

全面剖析 GBM 的分子图谱以及增强子结合抑制因子 2（EZH2）在肿瘤微环境（TME）中的异质分布和潜在作用。

方法

对 8 例 GBM 样本进行单细胞 RNA 测序（scRNA-seq）分析。进行去卷积分析、免疫荧光（IF）显微镜、逆转录定量聚合酶链反应（RT-qPCR）、集落形成实验和细胞计数试剂盒-8（CCK-8）检测，以确认 EZH2 在 TME 细胞中的潜在作用。

结果

恶性细胞表现出异常代谢模式的显著异质性。首次发现了一种具有胶质-免疫双重特征的间充质 2 样（MES2 样）GBM 亚群，其与高度激活的标志性途径、免疫逃避相关转录因子（IRF8）和不良预后相关。致癌基因 EZH2 在恶性细胞和免疫细胞中呈异质表达，与增殖基因、细胞周期转录因子和相似的激活标志性途径一致。在一个肿瘤相关巨噬细胞（TAMs）亚群（macrophage.3）中，EZH2 高表达，与细胞周期基因和巨噬细胞 M2 表型基因具有相似的转录组动力学变化。此外，该亚群与恶性细胞紧密相互作用。去卷积分析显示，与低级别胶质瘤（LGG）相比，该亚群在 GBM 中的丰度增加，并与预后不良显著相关。功能验证实验证实了 TAMs 中 EZH2 过表达在 GBM 中的促肿瘤作用。