Single-cell analysis identifies a VIM+ glioma stem-like vascular-immune state linked to myeloid OSM signaling and...

Gliomas show intertumoral and intratumoral heterogeneity, and isocitrate dehydrogenase (IDH) status is linked to biological behavior and clinical aggressiveness. Evidence suggests glioma progression depends on reciprocal crosstalk between glioma stem-like tumor cells (GSTCs) and the tumor microenvironment, where immune and vascular remodeling promote plasticity, invasion, and immune evasion. However, GSTC state architecture across IDH backgrounds and its links to vascular-immune cues remain unclear. We aimed to define IDH-associated GSTC states and their functional and regulatory features.

Single-cell RNA-sequencing data from 33 glioma patients (GSE278456) were analyzed using Seurat integration, clustering, differential expression analysis, and inferCNV-assisted malignant cell identification. Reclustered GSTCs were assessed by Gene Ontology (GO)/Hallmark enrichment, Area Under the Curve-based Cell-wise Gene Set Enrichment Analysis (AUCell), CytoTRACE, Monocle, Slingshot, CellChat, and pySCENIC to characterize states, developmental trajectories, cell-cell communication, and regulatory networks. After CEBPD silencing in U251 and LN229 cells, quantitative real-time PCR (qRT-PCR), Cell Counting Kit-8 (CCK-8), colony formation, Transwell migration, and apoptosis assays were performed.

Six cell populations and five GSTC subgroups were identified. A VIM+ GSTC state (C0), enriched in IDH-wild-type tumors, highly expressed CHI3L1, VIM, LGALS3, VEGFA, and SEC61G. Compared with other states, C0 showed stronger angiogenesis, hypoxia, IL6/JAK/STAT3 signaling, and epithelial-mesenchymal transition-related activities, with enrichment of oxidative phosphorylation, focal adhesion, cell-substrate junction, and actin/cadherin-binding programs. Trajectory analyses placed C0 at an early-to-intermediate node linked to divergent later states, indicating plasticity distinct from a purely proliferative state. CellChat suggested prominent myeloid-to-C0 communication, with OSM as a candidate input and LIFR/IL6ST as the receptor basis. pySCENIC identified a C0-preferential regulon landscape centered on CEBPD, RUNX2, POU2F3, TEAD3, and TEAD4, plus an M2 regulon module associated with stress adaptation and intracellular homeostasis. CEBPD knockdown reduced proliferation, colony formation, and migration while increasing apoptosis.

We identify an IDH-wild-type-enriched VIM+ glioma stem-like state with angiogenic, hypoxic, mesenchymal-like, and inflammatory features, and nominate myeloid-derived OSM signaling and CEBPD as candidate mediators of this vascular-immune-linked malignant phenotype. These findings provide a hypothesis-generating, state-based perspective on glioma progression across IDH backgrounds and highlight candidate biomarkers and mechanistic entry points for validation and therapeutic stratification.