Fig. 2: Transcriptome analysis (scRNA-seq) demonstrates an immediate increase in neurodifferentiation upon EPO application.

a Experimental design of WT C57Bl6 mice (n = 3 per group), treated at P28 with a single i.p. injection of placebo or EPO (5000IU/kg), followed by isolation of the CA1 region after 6 h and processing for DropSeq analysis (processing of biological replicates performed separately, followed by pooling for graphical presentation and final analysis), as well as after 24 h for immunohistochemistry (figure created by Debia Wakhloo). b Visualization of hippocampal CA1 cell clusters using t-distributed stochastic neighbor embedding (tSNE). Each color represents a cluster of specific cells, characterized by a defined gene expression profile. c Individual cells derived from either EPO (red) or placebo (black)-treated mice are denoted. d Percentage of cells in the respective glutamatergic clusters per treatment condition (EPO: n = 583, placebo: n = 390); two-tailed Fisher’s exact test. e Representative images and quantification of Tbr1 and Tle4 staining (immature neuronal markers) in CA1 of mice at P29, i.e., 24 h after a single injection of EPO. Within bars, mouse n numbers indicated; mean ± SEM presented; two-tailed student’s t-test. f Trajectory analysis in Monocle2 of cells in the ‘Mature Glutamatergic1’ and the ‘Immature Glutamatergic’ cluster colored by pseudotime (the darker the more mature). g Trajectory colored by cell identity. Bar graph indicates average pseudotime of the respective clusters; mean ± SEM; two-tailed Mann–Whitney U test. h Trajectory colored and split by placebo (black; left) versus EPO treatment (red; right). Bar graph shows average pseudotime of cells in the respective treatment groups; two-tailed Mann–Whitney U test; mean ± SEM; see also Supplementary Figs. 1–3. Source data underlying graph e are provided as a Source Data File.