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. 2016 Jun 28;5:e14845. doi: 10.7554/eLife.14845

Figure 4. ephrin-B2 is a glioblastoma oncogene controlling anchorage independent proliferation.

(a) Representative bioluminescent images of nude mice 20 days after intracranial injection of imNSC1, imNSC1-Efnb2, GSC1, GSC1Efnb2-/-. (b) Kaplan-Meier survival plots of the mice depicted in (a). Significance is given relative to imNSC1 for imNSC1-Efnb2 and to GSC1 for GSC1-Efnb2-/-. Log Rank Mantel Cox. (c, d) Left: Representative micrographs of GSC1 (c) and GSC2 (d) cells of indicated genotype grown in soft agar for 10d. Right: quantification of number of colonies formed in soft agar in all cultures, expressed as percentage over total number of seeded cells. n = 3, error bars depict s.e.m. One way ANOVA with Tukey correction. (e) Left: representative FACS profiles of cells grown in attachment or methylcellulose for 72 hr, showing DNA content by propidium iodide (PI) staining. Right: quantification of cell cycle phases from the FACS profiles. n = 3–5 as indicated by the dots. Error bars depict s.e.m., p values indicate significance of changes in G2/M phase. (f) Representative PI FACS profiles and quantifications of imNSC and GSC1 isolated from brain tissue 7 days after intracranial injection. n = 3. Error bars depict s.e.m. One way ANOVA with Tukey post hoc test shown for G2/M phase. See also Figure 4— figure supplement 1 and Figure 4— source data 1.

DOI: http://dx.doi.org/10.7554/eLife.14845.021

Figure 4—source data 1. Raw data for Kaplan Meier analysis, number of colonies formed in soft agar and cell-cycle analysis presented in Figure 4.
DOI: http://dx.doi.org/10.7554/eLife.14845.022
elife-14845-fig4-data1.xlsx (27.2KB, xlsx)
DOI: 10.7554/eLife.14845.022

Figure 4.

Figure 4—figure supplement 1. Anchorage-independent proliferation is independent of homotypic cell-cell contacts.

Figure 4—figure supplement 1.

(a) Left: representative phase contrast images of imNSC1-Efnb2 cells transduced with GFP- (Ad-GFP) or dominant negative N-Cadherin- adenoviruses (Ad-DN-Ncad) and cultured in soft agar for 10 days. Right: quantification of the percentage of cells forming colonies in soft agar. Error bars denote s.e.m. n = 3. (b) Western analysis of ephrin-B2 levels in GSC2 transfected with scrambled (scr) or Efnb2-specific (siEfnb2) siRNAs, confirming efficiency of the knock-down. n = 2 (c) Quantification of activated caspase3 positive cells in the indicated methylcellulose cultures. Values are expressed as percentage of total number of cells. See also Figure 4—figure supplement 1—source data 1.
DOI: http://dx.doi.org/10.7554/eLife.14845.023
Figure 4—figure supplement 1—source data 1. Raw data for all quantitative analyses shown in Figure 4—figure supplement 1.
DOI: http://dx.doi.org/10.7554/eLife.14845.024
elife-14845-fig4-figsupp1-data1.xlsx (27.8KB, xlsx)
DOI: 10.7554/eLife.14845.024