Single-Cell Atlas Reveals Complexity of the Immunosuppressive Microenvironment of Initial and Recurrent Glioblastoma

Abstract

The Glioblastoma (GBM) immune microenvironment plays a critical role in tumor development, progression, and prognosis. A comprehensive understanding of the intricate milieu and its interactions remains unclear, and single-cell analysis is crucially needed. Leveraging mass cytometry (CyTOF), we analyzed immunocytes from 13 initial and three recurrent GBM samples and their matched peripheral blood mononuclear cells (pPBMCs). Using a panel of 30 markers, we provide a high-dimensional view of the complex GBM immune microenvironment. Hematoxylin and eosin staining and polychromatic immunofluorescence were used for verification of the key findings. In the initial and recurrent GBMs, glioma-associated microglia/macrophages (GAMs) constituted 59.05 and 27.87% of the immunocytes, respectively; programmed cell death-ligand 1 (PD-L1), T cell immunoglobulin domain and mucin domain-3 (TIM-3), lymphocyte activation gene-3 (LAG-3), interleukin-10 (IL-10) and transforming growth factor-β (TGFβ) demonstrated different expression levels in the GAMs among the patients. GAMs could be subdivided into different subgroups with different phenotypes. Both the exhausted T cell and regulatory T (Treg) cell percentages were significantly higher in tumors than in pPBMCs. The natural killer (NK) cells that infiltrated into the tumor lesions expressed higher levels of CXC chemokine receptor 3 (CXCR3), as these cells expressed lower levels of interferon-γ (IFNγ). The immune microenvironment in the initial and recurrent GBMs displayed similar suppressive changes. Our study confirmed that GAMs, as the dominant infiltrating immunocytes, present great inter-and intratumoral heterogeneity and that GAMs, increased exhausted T cells, infiltrating Tregs, and nonfunctional NK cells contribute to local immune suppressive characteristics. Recurrent GBMs share similar immune signatures with the initial GBMs except the proportion of GAMs decreases.

PD-L1-Mediated Immunosuppression in Glioblastoma Is Associated With the Infiltration and M2-Polarization of Tumor-Associated Macrophages

“…Alternatively, stimuli such as IL-4, IL-14, IL-10 can induce macrophages towards an anti-inflammatory and cancer-promoting M2 phenotype (41,42). In GBM, TAMs were the predominant infiltrating immune cells and usually polarized to an immunosuppressive M2-like phenotype (43,44). This study shows that PD-L1 may be an important regulator of macrophages in GBM, supported by the positive correlation between them.…”

Sensei: How many samples to tell evolution in single-cell studies?

“…Although necessary approximations are made, the estimation is accurate as indicated by its consistency with the result of computer simulation and of patient data from real experiments. Results from previous single-cell studies are also consistent in the ballpark with Sensei's predictions[38]. Sensei runs in seconds on the front-end without requiring any connection to backend servers, providing versatile, secured utilities for researchers with limited resources.…”

Single-Cell Analysis Reveals Characterization of Infiltrating T Cells in Moderately Differentiated Colorectal Cancer

“…The strong effector functions are lost and various inhibitory receptors are expressed in exhausted T cells (T EX ) ( 33 ). Fu et al suggested that the tumor tissue showed high percentages of T EX and Treg cells compared to those in the peripheral blood ( 34 ), suggesting that tumor tissue showed relatively more immunosuppressive phenotypes. Consistently, CD8 + T EX cells were identified from the tumor tissue but not from the peripheral blood, in our study.…”