Figure 1.

Hypoxia Pretreatment Enhances Tropism of NSC toward Human GBM Cells

(A and B) GFP⁺ NSCs treated with hypoxia (^HNSCs; 1% O₂; 24 hr) or control NSCs (^NNSCs) were compared for ability to migrate toward tumorspheres. ^HNSCs effectively cross the culture insert membrane and accumulate within tumorspheres of patient-derived GBM43 glioma cells visualized with DAPI nuclear stain (blue) (A) and U87MG mCherry cells (B). Right panels represent quantification of the relative presence of GFP⁺ NSCs, indicating significantly higher numbers of GFP^+HNSCs than ^NNSCs inside the tumorspheres by 24 hr of co-culture (n = 5, Student's t test; ^∗∗∗p < 0.001 for GBM43, ^∗p < 0.05 U87MG). Scale bars, 100 μm.

(C) Collagen hydrogel migration assay further demonstrates the enhanced tumor tropism by NSCs after hypoxia pretreatment; Confocal z-stack 3D reconstruction shows GFP⁺ NSCs (green) seeded on top of the collagen hydrogel layer migrating to the bottom of the culture dishes in both GBM43blue (blue cells, top panels) and U87MG mCherry adherent cultures (red cells, bottom panel) by 12 hr. Quantification of migrated GFP⁺ NSCs by microscopy visual field (right) confirms the increased “homing” by ^HNSCs in both the GBM43blue and U87MG mCherry cells after hypoxia pretreatment (n = 7; ^∗∗p < 0.01, Student's t test, error bars with SEM).