To explore the cellular heterogeneity and the differences in branched trajectory of pericytes between keloids and localized scleroderma, and to provide new clues for the pathogenesis and therapeutic targets of the two skin fibrotic diseases.

Single cell transcriptome sequencing (scRNA-seq) data of 3 cases of scleroderma, 4 cases of keloid and their corresponding 4 cases of adjacent normal skin samples were selected from GEO and GSA-Human databases, and the expression matrix of the data was drawn. Seurat 4.3.0 of R (4.2.2) was used to process the t-distributed stochastic neighbor embedding (t-SNE) visualization map. Monocle 2.24.0 was used to analyze the pseudo-temporal trajectory of pericytes.

The unsupervised clustering of keloid and scleroderma skin tissues revealed 19 different cell populations, among which C7 and C11 cells were pericytes, marked by high expression levels of PDGFRB and RGS5 genes, accounting for 7.53% of the total cells. Pericytes can be further divided into 8 subgroups. Pseudo-temporal analysis revealed a branched trajectory with two major branches, that is, cell fate 1 and cell fate 2, which could be further divided into 5 cellular states of pericytes (S1-S5). S4 constituted the most of the prebranch, which represented the cellular state of the initial pericyte phenotype. S5 constituted the most of the cell fate 1 branch, which represented the early differentiation state of the pericyte phenotype. S1, S2, S3 constituted the most of the cell fate 2 branch. S3 represented the intermediate differentiation state of the pericyte phenotype, while S1 and S2 represented the terminal differentiation states of the pericyte phenotype. Compared with the uniform distribution of various differentiation states of pericytes in normal skin, the keloid pericytes mainly distributed in the prebranch (S4), cell fate 1 (S5) and the first half of cell fate 2 (S3), representing cellular states of the initial, early and intermediate phases of the pericyte phenotype. Branched expression analysis modeling revealed the overexpression of SOX4, COL4A1, COL6A3, AHR, CXCL3 and IL1R1 genes, et cetera. On the other hand, the localized scleroderma pericytes mainly distributed in the bottom half of cell fate 2 (S1, S2), representing the final differentiated phase of pericyte phenotype, which overexpressed ACTA2 and MYH11 genes.

Pericytes in keloid and scleroderma are heterogenous and have different differentiation trajectories. Pericytes in keloid have stem-like characteristics, and play an important role in the pathologic characteristics of invasiveness and recurrence through high expression of genes related to cell stemness, epithelial-mesenchymal transition, invasiveness, and immune microenvironment regulation. However, pericytes in localized scleroderma may mainly transdifferentiate into myofibroblasts, leading to their fibrotic pathological phenotype.

Keloid; Localized scleroderma; Pericyte; Single-cell RNA-sequencing; Pseudo-time analysis