Fig. 3.

C9-ALS iAstrocytes affect cell dynamics and paracellular permeability on BMEC-like cells A) BMEC passive barrier as shown by TEER following BMEC-like cells/iAstrocytes co-culture for C9-ALS BMEC-like cells. Error bars represent the standard deviation of triplicate Transwell™ filters. Statistical significance was determined using One-Way ANOVA (****p < 0.0001). N = 3. B) BMEC-like cells were either treated with control or C9-iAs-conditioned media. BMEC-like cells were fixed in 100% MeOH after 48 h of the experiment. Scale bar equals 50µM. Plotted results represent confocal image z-stacks analysed in 3D volume. Claudin-5 is shown in green and Glucose-1 transporter (Glut-1) in red; and DAPI in blue for the nuclei staining. Error bars represent the standard deviation of triplicate Transwell™ filters. Statistical significance was determined using One-Way ANOVA (****p < 0.0001). N = 3. At least a total of 5 images were acquired per each condition, with a minimum of 3 technical replicates for a total of 3 biological replicates (N = 3, total images per condition = 45). C) The LDH assay was performed after 2 days of iAstrocytes conditioned media treatment on BMEC-like cells. Untreated refers to BMEC-like cells on their indicated endothelial media and non-cond iA media are BMEC-like cells treated with plain iAstrocytes media to evaluate the effect of the media on the BMEC-like cells. Control (CTR) and C9-ALS media (ALS). Error bars represent the standard deviation of triplicate wells. Statistical significance was determined using One-Way ANOVA (****p < 0.0001). N = 3