Fig 2: Dysregulated gene expression and defective neuronal migration in NANS-KO cerebral organoids.(A) Scheme depicting the experimental setup of neuronal differentiation and time-lapse imaging between WT and NANS-KO neurons. Scale bar, 10 μm. (B) Confocal imaging of WT and NANS-KO neurons after staining with antibodies against NeuN. Scale bar, 25 μm. (C) Quantification of different parameters of neuronal migration dynamics, tracked using live imaging. Resting TP, resting time points; n = 31 to 53 neurons from two independent experiments, **P < 0.01, ***P < 0.001, Student’s t test. (D) PCA of gene expression over time in WT and NANS-KO cerebral organoids. (E) Heatmap showing distinct developmental stages. DIV 0, stem cell stage; DIV 15, NPC stage; DIV 25, neurogenesis; DIV 35, the early developmental stage of neuronal maturation with concurrent gliogenesis. (F) Volcano plots of DEGs in WT and NANS-KO cerebral organoids at DIV 0, 15, 25, and 35. FC, fold change. (G) Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of DEGs in WT and NANS-KO cerebral organoids at DIV 35. (H) Heatmap showing DEGs associated with ECM organization and cell adhesion at DIV 35. AGE-RAGE, advanced glycation end product (AGE) and their receptor (RAGE); PI3K-Akt, phosphoinositide 3-kinase and Akt.