Figure 4.

Stiff condition induced sprouting angiogenesis of endothelial cells and enhanced drug resistance of GBM cells. (A) Immunofluorescence staining of SOX2 (GBM cells) and CD31 (endothelial cells) in the stiff co-culture condition vs. the soft co-culture condition. White arrow: sprouting angiogenesis. Scale bars: 50 μm. (B) mRNA expressions of angiogenesis markers SPP1 and VEGFA in TS576 in (1) sphere culture, (2) tumor-only soft condition, (3) co-culture soft condition, (4) tumor-only stiff condition, and (5) co-culture stiff condition measured by real-time quantitative PCR (RT-qPCR). N=3 technical replicates for each condition. P-values were calculated using two-way ANOVA corrected for multiple comparisons using Dunnett test. **, p < 0.01; ****, p < 0.0001. (C) Invasion patterns of TS576 cells in 3D co-culture models. (D) TS576 viability after 6 days of Temozolomide treatment. n=3 technical replicates for each condition. P-values were calculated using ordinary one-way ANOVA corrected for multiple comparisons using Tukey test. *, p < 0.05; **, p < 0.01; ****, p < 0.0001. (E) mRNA expressions of drug resistance markers ABCG2, MGMT, and CXCL12 in TS576 measured by RT-qPCR. N=3 technical replicates for each condition. P-values were calculated using two-way ANOVA corrected for multiple comparisons using Turkey test. **, p < 0.01; ****, p < 0.0001.