Fig. 3 |. Rett syndrome fused and unfused organoids have similar cortical organization and cell type expression profiles.

a,b, Generation and immunohistochemical analyses of isogenic Cx and GE organoids from Rett syndrome patient hiPSC that either contain (iCtrl) or lack (Mut) MECP2 expression (see also³⁹). iCtrl and Mut Cx organoids exhibit comparable formation of neural progenitors (SOX2, TBR2), both deep and superficial layer neurons (CTIP2, BRN2), and inhibitory interneurons (GAD65, SST, and GABA) All images reflect representative images from at least 3 independently imaged sections, refer to supplementary table 4 for further details. c,d, D100 unfused iCtrl and Mut Cx organoids show minimal expression of GAD65, whereas ~20-25% of the cells in the Cx end of age matched Cx+GE organoids express GAD65, n = 3 independently generated organoids, 2631 cells, ns, not significant by a two-sided t-test. e,f, Immunohistochemical analysis of interneuron subtypes by the co-expression of GAD65 with SST, Calretinin, NPY or Calbindin in the Cx portion of day 100 iCtrl or Mut Cx+GE fusion organoids reveals the presence of multiple interneuron subtypes e. f, Cell counting reveals trends for all comparisons and a statistically significant difference between iCtrl and Mut samples with respect to the percentage of cells expressing calretinin, n = 3 fusion organoids per genotype, ≥ 980 cells for each sample counted, ns = not significant for all groups except calretinin. ANOVA P = 0.0003, F = 4.665, DF (between columns) = 7, followed by Sidak’s correction for multiple comparison between iCtrl and Mut for each marker; *P = 0.0391 for iCtrl vs Mut calretinin; ns for all other comparisons. Plots in d,f display the full distribution of individual data points with dotted lines indicating the median and quartile values.