Fig. 5: Adjustment of neuronal cell numbers during later differentiation.

A Representative immunocytochemistry images of hiPSCs-derived neurons treated with UNC0638 (250 nM) at days 30, 35 and 45 of differentiation stained with NeuN (green) and counterstained with DAPI (blue). Scale Bar, 50 µM. Graph shows quantitation of NeuN expression (mean average intensity of cell nuclei, all data shown) at day 35 time point. B Total number of NeuN-cells at days 25, 30, 35 and 45 of differentiation in the absence and presence of UNC0638, n ≥ 3 independent experiments. C Western blot analysis at days 25 and 30 to examine the expression of cleaved-caspase-3 in the presence and absence of UNC0638 (250 nM). Top panel shows the expression of uncleaved caspase-3 (inactive), middle panel shows the expression of cleaved-caspase-3 (activated) and bottom panel shoes expression of GAPDH. Quantification of Western blot analysis was performed by normalization to GAPDH, n ≥ 3. D qRT-PCR analysis to examine the change in the expression of caspase-3 in hiPSCs-derived neurons treated with UNC0638 (250 nM). The expression of caspase-3 increased in UNC0638-treated neurons and patient iPSC compared to untreated neurons. Fold change over untreated neurons (mean). n ≥ 3 independent experiments. Data were presented as Mean±SEM and analysed by student’s t-test or One-way ANOVA with post hoc comparisons using Dunnett’s multiple comparisons test comparing to control samples.