Fig. 3

Pathway and differential gene expression analysis confirm immune features across immune clusters. A Heatmap showing log-fold change in gene enrichment scores (derived from QuSAGE) in each immune cluster compared to all others in pedNST. Columns show MSigDB pathways with false discovery rate (FDR) < 0.1 in at least three immune clusters. Black box outlines pathways validated by protein in (B). B Ridge plots illustrating average z-scores of proteins [62] involved in immune pathways from A across immune clusters in 141 pedCNS samples (CBTN). Fraction indicates the number of protein readouts from the number of genes in each pathway. Pairwise two-sided Student’s t test with Bonferroni correction, *p < 0.05, **p < 0.01 and ***p < 0.001. Significance levels are shown for comparison to Paediatric Inflamed. C Volcano plots for differentially expressed genes (derived from DESeq2) in each immune cluster compared to other samples. Up- or downregulated genes with absolute log-fold change > 1.5 and FDR < 0.1 are shown in red or blue, respectively. Dashed line shows the log-fold change threshold. Top ten differentially expressed genes are annotated. D Heatmap showing median z-score expression of 59 genes with known immunoregulatory functions across immune clusters in pedNST