Fig 1.
Ex vivo non-contrast T2-weighted 2D MRI scans of ex vivo brains depicting the tumor progression.
Panels show MRI scans at day 0 (D0), D1, D3, D7, D14, and D21 post-inoculation of murine glioma Gl261 cells into the caudoputamen (CPu). Left (L) and right (R) of the brain are indicated. Images are representative of N = 3 mice per time point.
Fig 2.
Histopathology of Gl261 gliomas.
Four μm paraffin sections of murine Gl261 brain tumors stained with hematoxylin and eosin Y/eythrosin (A). High resolution of morphology of the Gl261 cells as the tumor grows and matures (B). Images are representative of N = 12 mice per time point.
Fig 3.
Ki67 proliferation index of Gl261 gliomas.
Ki67+ staining at the inoculation site of murine Gl261 gliomas (A). Scale bar, 200μm. Tumors at day 21 post-inoculation showed signs of hypoxia/necrosis (B). Scale bar, 500μm. Quantitation of five HPF for each murine brain (N = 12 per time point; C). Data are expressed as mean ± SEM per HPF. ap<0.05 vs D0, bp<0.05 vs D1, cp<0.05 vs D21 by Tukey’s multiple comparison test.
Fig 4.
Number of blood vessels in Gl261 gliomas.
CD31+ blood vessels at the inoculation site of murine Gl261 gliomas (A). Scale bar, 200μm. Quantitation of blood vessels (B) and lumen area (C) in five HPF (20x) for each murine brain (N = 12 brains per time point; B). Data are expressed as mean ± SEM per HPF. (B) ap<0.05 vs D0, bp<0.05 vs D1, cp<0.05 vs D3, dp<0.05 vs D14 by Tukey’s multiple comparison test. (C) ap<0.05 vs 21 by Tukey’s multiple comparison test.
Fig 5.
(A) Whole slide scans (4x) and MSI spectrally unmixed and pseudo-colored micrographs (20x) of the three immune cell panels for lymphocytes, macrophages and DC populations in the tumor infiltrate. (B-C) Quantitation of immune cell phenotypes in HPF for each murine brain (N = 6 brains per time point). Data are expressed as mean ± SEM per HPF (B) and as mean percentage normalized to day 0 (C). ap<0.05 vs D0, bp<0.05 vs D1, cp<0.05 vs D3, dp<0.05 vs D7, ep<0.05 vs all other time points by One-way ANOVA with Tukey’s multiple comparison test.
Fig 6.
Systemic immune cell populations.
Immune cell populations from spleen, bone marrow and peripheral blood from mice inoculated with glioma Gl261 cells. Cells were harvested at day 0 (sham) to day 21 time points and assessed by multi-color flow cytometry. Data are expressed as percentage of parent population (%). ap<0.05 vs D0, bp<0.05 vs D1, cp<0.05 vs D3, dp<0.05 vs D7; ep<0.05 vs D14; fp<0.05 vs D21 by Dunn’s Multiple Comparison Test.
Fig 7.
Plasma cytokine and chemokines levels from mice inoculated with glioma Gl261 cells.
Plasma was harvested at day 0 (sham) to day 21 time points and assessed by 23-plex immunoassay, except TGF-β1 (by chromogenic ELISA). Data are expressed as observed concentration in pg/ml. ap<0.05 vs D14; bp<0.05 vs D21; cp<0.05 vs D7; dp<0.05 vs D3; ep<0.05 vs D1 by Dunn’s Multiple Comparison Test.
Fig 8.
Color maps of the p-values for Pearson correlations between plasma cytokine levels and tumor and systemic immune cell populations at day-14 post-inoculation (A) and at all time points (B). Red, p>0.05, white p = 0.05, green p<0.05.