Figure 1.
PDGF-IRES-Cre retrovirus induced tumor formation in mice with floxed Pten and p53.
(A) Kaplan-Meier curves comparing the survival of mice from our 4 experimental groups: All PIC injected Ptenf/f mice and Ptenf/f; p53f/f mice developed brain tumors with median survivals of 85 dpi and 27 dpi, respectively. Three mice from the control groups died from unrelated causes during the study. The remaining control mice were sacrificed and analyzed at the end of the study, none showed evidence of tumor. The survival differences between all groups were highly significant. The p values for all comparisons are shown in the right panel. (B–E) Low and high powered micrographs showing H & E stains of end stage tumors in Ptenf/f mice (B and C) and Ptenf/f; p53f/f mice (D and E). Both tumors show histological features of GBM, including areas of pseudopalisading necrosis (N). (F–G) Bioluminescence imaging of tumor growth in Ptenf/f; p53f/f (F) and Ptenf/f (G) mice. (H) Plot showing the different growth rates of tumors in Ptenf/f (blue line) and Ptenf/f; p53f/f (red line) mice. Each line shows the changes in the mean value of the luciferase signal of 11 mice per group. Bars show the S.E.M at each time point.
Figure 2.
Immunophenotype of end stage tumors is consistent with OPC identity.
Each triptych shows the immunostaining (red), Hoechst nuclear stain (blue) and combined for low power (left) and high power (right) micrographs of ends-stage tumors in Ptenf/f; p53f/f mice. (A and B) Most cells in the tumor lacked Pten staining. (C–F)The vast majority of tumor cells express olig2 (C and D) and PDGFRα (E and F). (G and H) GFAP+ astrocytes were scattered throughout the tumor, but represented a minority of cells.
Figure 3.
Phenotypes of early tumor lesions in YFP; Ptenf/f mice indicated OPC identity.
(A–C) Schematic views show the distribution of YFP+ cells (green dots) at 3 dpi (A), 7 dpi (B) and 21 dpi (C). (D–F) Triple immunofluorescence shows YFP (green) ki67 (red) and Olig2 (blue) at 3 dpi (D), 7 dpi (E) and 21 dpi (F). The arrows in D and E mark triple positive cells. The tables below each panel show the counts of ki67+/YFP+ cells at each time point (total, Olig2+, and Olig2−). (G–I) Triple immunofluorescence shows the expression of different markers in tumor cells at 21 dpi: (G) shows ki67 (red), YFP+ (green) and PDGFRα+ (blue). (H) shows Olig2 (red), YFP (green) and GFAP (blue). (I) shows Olig2 (red), YFP (green) and NeuN (blue). (J–O) Low power montage of 21 dpi tumor shows Olig2 (J), YFP (K), PDGFRα (L), Hoechst (M), GFAP (N) and Pten (O).
Figure 4.
PDGF driven mouse tumors most resembled human Proneural GBM and express signatures of OPCs.
(A) Heat map of the Pearson correlation based classification of mouse and human GBM samples. Red to blue color scale shows the range from the highest positive to highest negative correlation. The correlation was computed for both mouse samples and TCGA human GBM. A sample is assigned the class with the highest correlation coefficients (see Methods for details). (B) Mouse tumors and human Proneural patients were both enriched with OPC genes. Sample wise: mouse tumors are labeled as black, Proneural as green, Neural as blue, Classical as red and Mesenchymal as yellow. Red to blue color scale shows the range from the highest to lowest enrichment score. These genes lists are provided in Table S1.
Figure 5.
DGE lists reveal heterogeneity in human Proneural GBM.
(A) GSEA of mouse tumors with DGE lists of Ptenf/f vs. Ptenf/f; p53f/f mouse tumors. Sample wise: Ptenf/f; p53f/f tumors were labeled as blue and Ptenf/f tumors as green. Red to blue color scale shows the range from the highest to lowest enrichment score. (B) GSEA ranking of TCGA Proneural patients with DGE lists of Ptenf/f vs. Ptenf/f; p53f/f mouse tumors. Horizontal red line on top of panel labels 1/3 of Proneural patients that most resemble Ptenf/f; p53f/f tumors, while blue line labels 1/3 of patients that most resemble Ptenf/f tumors. Similar patients from four datasets are pooled together to compare survival as in (C). Sample wise: Proliferative is labeled as blue, Mesenchymal as red and the rest of Proneural as green (See discussion). (C) Kaplan-Meier survival curve comparison of patients that resemble Ptenf/f; p53f/f mouse tumor vs. those that resemble Ptenf/f mouse tumor.
Figure 6.
Ontology analysis reveals many genes on DGE lists are related to p53 signaling.
(A) 104 members of DGE list were connected in Ingenuity pathway analysis. Out of these, 8 members of core p53 signaling pathway are highlighted as red. The shapes on the left indicate the functions and relationships of the genes. (B) The biological processes and disease pathways are significantly enriched with genes on the DGE list. The yellow line indicates the threshold to significance.