Supplementary Figure 7: ROS levels in CAFs, migration of CM-treated cancer cells, and GLUD1 expression.

(a) Fluorescence intensity of the ROS indicator CM-H2DCFDA in CAFs treated as indicated in glutamine/glutamate-free medium containing 5 mM NH₄Cl (n = 3 independent experiments). H₂O₂ was used as a positive control to induce ROS. (b) MDA-MB-231 cells were cultured in NH₄⁺-containing medium (3 g/L glucose; glutamine/glutamate-free; 5 mM NH₄Cl) that had been conditioned by CAFs treated as indicated. After 24 h, MDA-MB-231 cells were subjected to transwell migration assay and cells that had migrated within 16 h were quantified (n = 3 independent experiments). (c) Left: Relative RNA levels of GLUD1 in EV-treated CAFs grown in regular medium (3 g/L glucose; 4 mM glutamine) (n = 3 independent experiments). Right: Western blots showing protein levels of GLUD1 in EV-treated CAFs grown in regular medium. For the entire figure, data are shown as mean ± SD; statistical significance was assessed using one-way ANOVA. **P < 0.01, ***P < 0.001. Unprocessed original scans of blots are shown in Supplementary Figure 9. Source data are shown in Supplementary Table 5.