Figure 4
Single-cell transcriptomics of midbrain periaqueductal gray (PAG) cells in CFA-induced chronic inflammatory pain model. (a) Overview of single-cell RNA sequencing (scRNA-seq) experimental design to define the mechanism of action (MOA) of ApAP and SRP-001. The periaqueductal gray (PAG) midbrain region was dissected to isolate cell nuclei for sequencing library generation using the 10 × Genomics Chromium platform. Single cell data was embedded after dimensionality reduction using uniform manifold approximation and projection (UMAP). Cell clusters were annotated, followed by differential gene expression analysis. (b) UMAP plots showing the distribution of annotated cell clusters color-coded by each cluster – astrocytes, excitatory neurons, inhibitory interneurons, oligodendrocytes, oligodendrocyte precursor cells (OPC), and microglia, from the aggregated clusters of all the 4 samples. (c) UMAP plots showing the distribution of annotated cell clusters generated by the workflow of Seurat across 4 different samples – Vehicle, CFA_Veh, CFA_ApAP, and CFA_SRP-001, respectively. (d–i) Marker gene feature plots showing selected distinctive marker genes used for cell annotation of single cell clusters into 6 different cellular subpopulations. (d,e) Oligodendrocytes (f), Astrocytes (g) OPC (h) Glutamatergic neurons i GABAergic neurons. (j–k) GO enrichment analysis results for top 50 differentially expressed genes between CFA_Vehicle and CFA_ApAP, and between CFA_Vehicle and CFA_SRP-001 – gene concept network showing the linkage between the DE genes and GO terms, and Barplot of the enriched GO terms from the selected genes.
