Fig 2: MGDep is associated with decreased BrMet and shifting of PD1− Tregs and γδ T cells under ABX conditions.(A and B) Dot plot of the relative gene expression level for (A) predicted receptors in the CD3+ T cell subset for interaction with top predicted ligands in microglia SC2 and (B) top predicted Tnf target genes in the CD3+ T cell subset.(C) Percentages of CD3+ SC7: CD4+ Tregs (left) and CD3+ SC11: CD3Low ILC3-like (right) of the CD3+ T cell subset at each time point and condition.(D) Schematic for MGDep and ABX treatment (top), stereoscope images of BrMet in control or MGDep mice on ABX treatment (left), and quantification of tumor number in each group (quantification based on pooled data from two independent experiments with ≥8 mice per group) (right). Scale bar: 5 mm.(E and F) Percentages (E) of the indicated CD4+FOXP3+ and (F) TCRgd+ T cell populations in control and MGDep BrMet-burdened brains.(G) Schematic of MGDep and vehicle/ABX treatment in tumor-naive mice.(H and I) Percentages (H) of the TCRgd+ and (I) CD8+ (left) and CD4+ (right) T cell populations in control and MGDep tumor-naive brains in vehicle- and ABX-treated mice.Plots in (A)–(C) were derived from pooling three biological replicates per time point and condition. Dot plots in (A) and (B) have relative gene expression level depicted by color scale and percentage by dot size. Data in (C) were analyzed with two-tailed Student’s t test, and error bars represent SD. Each dot represents 1 mouse in (D)–(F), (H), and (I), with data analyzed by two-tailed Student’s t test, error bars representing min and max, and center representing mean. Immune cells in (E), (F), (H), and (I) were enriched from brain tissue and measured by flow cytometry. See also Figures S6 and S7 and Tables S2 and S3.

Fig 3: NK T cell subsets are altered with MGDep and ABX treatment in BrMet-burdened brains.(A) Dot plot of the relative gene expression level of predicted MHCI receptors across CD3+ T cell SCs. Relative gene expression level is depicted by color scale and percentage by dot size.(B) Feature plots showing cells in the CD3+ T cell subset with co-expression of CD8 (top) and Klrd1 (bottom) with Klrc1.(C–E) Percentages of the indicated (C) NK T cell, (D) CD8 T cell, and (E) NK cell populations in control and MGDep BrMet-burdened brains.(F) Biaxial plot depicting the CD3+ SC2: NK T cell subset with and without Klrc1 and Lamp1 gene expression (left) and stacked bar charts of NK T cell subsets at each time point and condition (right).(G) Violin plot of DEGS on subset NK T cell populations.Plots in (A), (B), (F), and (G) were derived from pooling three biological replicates per time point and condition. Each dot represents 1 mouse in (C)–(E), with data analyzed by two-tailed Student’s t test, error bars representing min and max, and center representing mean. Immune cells in (C)–(E) were enriched from brain tissue and measured by flow cytometry. Data in (G) were analyzed by Wilcoxon rank-sum test. See also Figure S7 and Tables S2 and S6.