Specialized functions and sexual dimorphism explain the functional diversity of the myeloid populations during glioma progression.

Malignant gliomas are aggressive, hard-to-treat brain tumors. Their tumor microenvironment is massively infiltrated by myeloid cells, mostly brain-resident microglia, bone marrow (BM)-derived monocytes/macrophages, and dendritic cells that support tumor progression. Single-cell omics studies significantly dissected immune cell heterogeneity, but dynamics and specific functions of individual subpopulations were poorly recognized. We use Cellular Indexing of Transcriptomes and Epitopes by sequencing (CITE-seq) to precisely dissect myeloid cell identities and functionalities in murine GL261 gliomas. We demonstrate that the diversity of myeloid cells infiltrating gliomas is dictated by cell type and cell state. Glioma-activated microglia are the major source of cytokines attracting other immune cells, whereas BM-derived cells show the monocyte-to-macrophage transition in the glioma microenvironment. This transition is coupled with a phenotypic switch from the IFN-related to antigen-presentation and tumor-supportive gene expression. Moreover, we found sex-dependent differences in transcriptional programs and composition of myeloid cells in murine and human glioblastomas.

Single-Cell Omics in Dissecting Immune Microenvironment of Malignant Gliomas-Challenges and Perspectives.

Single-cell technologies allow precise identification of tumor composition at the single-cell level, providing high-resolution insights into the intratumoral heterogeneity and transcriptional activity of cells in the tumor microenvironment (TME) that previous approaches failed to capture. Malignant gliomas, the most common primary brain tumors in adults, are genetically heterogeneous and their TME consists of various stromal and immune cells playing an important role in tumor progression and responses to therapies. Previous gene expression or immunocytochemical studies of immune cells infiltrating TME of malignant gliomas failed to dissect their functional phenotypes. Single-cell RNA sequencing (scRNA-seq) and cytometry by time-of-flight (CyTOF) are powerful techniques allowing quantification of whole transcriptomes or >30 protein targets in individual cells. Both methods provide unprecedented resolution of TME. We summarize the findings from these studies and the current state of knowledge of a functional diversity of immune infiltrates in malignant gliomas with different genetic alterations. A precise definition of functional phenotypes of myeloid and lymphoid cells might be essential for designing effective immunotherapies. Single-cell omics studies have identified crucial cell subpopulations and signaling pathways that promote tumor progression, influence patient survival or make tumors vulnerable to immunotherapy. We anticipate that the widespread usage of single-cell omics would allow rational design of oncoimmunotherapeutics.

Single-cell RNA sequencing reveals functional heterogeneity of glioma-associated brain macrophages.

Microglia are resident myeloid cells in the central nervous system (CNS) that control homeostasis and protect CNS from damage and infections. Microglia and peripheral myeloid cells accumulate and adapt tumor supporting roles in human glioblastomas that show prevalence in men. Cell heterogeneity and functional phenotypes of myeloid subpopulations in gliomas remain elusive. Here we show single-cell RNA sequencing (scRNA-seq) of CD11b+ myeloid cells in naïve and GL261 glioma-bearing mice that reveal distinct profiles of microglia, infiltrating monocytes/macrophages and CNS border-associated macrophages. We demonstrate an unforeseen molecular heterogeneity among myeloid cells in naïve and glioma-bearing brains, validate selected marker proteins and show distinct spatial distribution of identified subsets in experimental gliomas. We find higher expression of MHCII encoding genes in glioma-activated male microglia, which was corroborated in bulk and scRNA-seq data from human diffuse gliomas. Our data suggest that sex-specific gene expression in glioma-activated microglia may be relevant to the incidence and outcomes of glioma patients.