Fig. 3: Advanced maturation of neuron in enTsOrg revealed by spatial transcriptomic and in vivo electrophysiological analysis.
From: Engineered thoracic spinal cord organoids for transplantation after spinal cord injury
a, Schematic diagram of the experimental design. ST-seq at 7 weeks after transplantation and in vivo electrophysiological experiments at 14 weeks after transplantation. b, Representative immunofluorescence timeline of mature neuron marker (NeuN) and NPC marker (SOX2, Nestin) in the transplanted-organoids. c, Statistic analysis of NeuN-positive cells and SOX2 and Nestin-positive cells in sOrg and enTsOrg at different time points after transplantation. The folded line showing the trend of the population of cells. Two-way ANOVA followed by uncorrected Fisher’s LSD (n = 3 biological replicates per group). d, Spot plots of neuron and progenitor distributions in organoid transplant areas by ST-seq at 7 weeks after transplantation. e, Pie diagrams of the neuron and progenitor percentage in d. f, Violin plots of progenitor and neuron-related gene expression levels by ST-seq. g, Distribution of predicted developmental stages CS12, CS14, CS17 and CS19 cell types in the organoid-transplanted area of the spinal cord tissues at 7 weeks, by transcriptome analysis referring to human embryonic spinal cord dataset (GSE171892). h, Bar plots showing the spot numbers with CS12, CS14, CS17 and CS19 identification in transplanted area. i, Distribution of excitatory neurons and inhibitory neurons in the organoid-transplanted area of the spinal cord tissues. Bar plots showing the spot numbers with excitatory and inhibitory neuron identification, n = 2 biological replicates. j, Schematic diagram of in vivo electrophysiologic signal collection. k, Histograms showing distributions, mean firing rate (z-score) of all units in sOrg and enTsOrg at 14 weeks after transplantation; each point indicates a unit. Two-sided Kolmogorov–Smirnov test (record from 6 animals in sOrg and enTsOrg group). l, Neurons distribution based on waveform properties in each group; each symbol represents one unit. m, Representative waveforms of two discrete neuron populations: P-IN and P-PN, according to waveform properties analysis by k-means. n, Proportion of P-PN and P-IN in organoid grafts of sOrg and enTsOrg at 14 weeks (record from 6 animals in sOrg and enTsOrg group). o, Mean firing rate (z-score) quantification of the P-IN and P-PN in the organoid grafts; each point indicates a unit, and black lines indicate the mean value. Kolmogorov–Smirnov test, n > 3. Scale bars, 100 μm in b. Data represent the mean ± s.e.m. in c. Data in k are presented as box-and-whisker plots showing the median (centre line), the 25th–75th percentiles (box limits) and the minimum and maximum values (whiskers); all individual measurements are shown as points overlaid on the plot. CS, Carnegie stages.
