Figure 1.

Unbiased clustering of scRNA-seq data reveals the major cell types of the glomerulus. (A) T-distributed neighbor embedding (t-SNE) representation of 5287 glomerular cells from a healthy C57BL/6J mouse. Labels indicate clusters identified by unsupervised clustering analysis. (B) Violin plots of marker gene expression in each cluster shown in (A). (C) Violin plots showing expression of glomerular capillary endothelial cell (Ehd3) or arteriolar endothelial cell (Fbln2, Mgp, Trpv4, Bmx) marker genes in clusters 2 and 4 (D). PCA plot of normal glomerular cells. Cluster labels are identical to (A). Proximity of clusters 3 and 5 indicate high degree of similarity between the two clusters. (E) Violin plots showing expression levels of genes specific to mesangial cells or SMCs/JG cells. The numbers in parentheses indicate cluster number. (F) Immunofluorescence staining of kidney sections shows mesangial-specific expression of the identified marker genes. PDGFRβ, which stains mesangial cells in the glomerulus (dotted circle) and stromal cells outside the glomerulus, was used as reference. Scale bars, 20 μm. (G) Expression levels of the indicated genes are shown in a t-SNE plot of SMCs/JG cells. The two subclusters corresponding to AA SMCs and EA SMCs are outlined. (H) Immunofluorescence staining of kidney sections shows specific staining of calponin 1 in the vascular SMCs of the AA. Scale bar, 10 μm. Normalized expression levels are shown in the violin plots. DAPI, 4′,6-diamidino-2-phenylindole; α-SMA, α-smooth muscle actin.