Fig. 5

Functional properties of GBM-educated Mg. A Signature score of genes associated with migration, sensome, phagocytosis, and antigen presentation per myeloid cluster: CL0-1 Ho-Mg, CL2-6: Mg-TAMs, CL7: Mo, CL8: BAMs. B Ex vivo assessment of migratory capacity in CD11b⁺ myeloid cells isolated from nude mouse normal brain (Nu-NB) and PDOXs P8, P13 (n = 3 mice/condition, mean ± SEM, **p < 0.01, ***p < 0.001, one-way ANOVA with Tukey’s HSD correction). C Ex vivo assessment of phagocytic capacity in CD11b⁺ myeloid cells isolated from Nu-NB and PDOXs P8, P13 (n = 3–4 mice/condition, mean ± SEM, *p < 0.05, one-way ANOVA with Tukey’s HSD correction). D Phagocytic uptake measured in CD45⁺CD11b⁺Ly6G⁻Ly6C⁻CD206⁻ Mg, CD45⁺CD11b⁺Ly6G⁻Ly6C⁺CD206⁻ Mo and CD45⁺CD11b⁺Ly6G⁻Ly6C⁻CD206⁺ BAMs. Representative example is shown for PDOX P13 tumor core (mean ± SEM, n = 3). E Phagocytic uptake measured in Mg in Nu-NB and PDOXs P8, P3, P13 (n = 3 mice/condition, ***p < 0.001, **p < 0.01, mean ± SEM, one-way ANOVA with Tukey’s HSD correction). An unstained control (CTR) represents Mg without E. coli particle incubation. For PDOXs P3 and P13 tumor core and distant normal brain area were collected. F Comparison of phagocytic uptake in CD45^high and CD45^low Mg in PDOX P8 (n = 3 mice, mean ± SEM, *p < 0.05, two-tailed Student’s t test). G Representative flow cytometry graphs and quantification of Mg positive for CD11c and CD86 in Nu-NB and PDOXs P3, P8, and P13 across different brain regions (n ≥ 3 mice/condition each, mean ± SEM, ****p < 0.0001, **p < 0.01, one-way ANOVA with Tukey’s HSD correction). An unstained control is shown for each population (CTR). H Representative immunofluorescence images depicting CD11c staining in Iba1⁺ cells in the Nu-NB and PDOX P8 tumor core. Scale bar: 20 µm. White arrows point CD11c⁺ positive signals. I Representative flow cytometry graphs showing activation of MHC-II (I-A/I-E epitope) in Mg, Mo, and BAMs in the tumor core and distant brain. Examples shown for PDOX P13 and P3 (mean ± SEM, n = 3 mice). J Representative immunofluorescence images depicting MHC-II and CD11c co-expression of Iba1⁺ cells in the P8 PDOX tumor core. Quantification represents mean values from 2 technical replicates. Scale bar: 20 µm. White arrows point MHC-II⁺CD11c⁺ signals in Iba1⁺ cells. MHC-II⁺Iba1.⁻ cells may represent tumor cells expressing rarely MHC-II [58]