Abstract: 【Abstract】 Objective To identify and differentiate cell subsets in the epidermis and dermis of vitiligo skin lesions using single-cell RNA sequencing technology, and to study the relationship between them. Methods Skin samples were collected from 2 healthy people without immune or systemic diseases and 2 patients with stable non-segmental vitiligo in Department of Dermatology in the Third People′s Hospital of Hangzhou in September 2019. Single-cell transcriptome sequencing was performed on 11 000 cells in all the skin samples by using 10 × Genomics single-cell RNA-Seq technology. Cell subsets were analyzed, screened and counted by using Seurat software. Results Cluster analysis of gene expression in the 2 normal skin tissues revealed several cell subsets, including keratinocytes, fibroblasts, nerve cells and melanocytes, endothelial cells, tissue stem cells, and immune cells mainly consisting of dendritic cells and T cells. In the 2 vitiligo lesions, abnormal differentiation and quantity were observed in fibroblasts and 4 keratinocyte subpopulations. The proportion of fibroblasts was significantly lower in vitiligo lesions than in normal skin tissues （0 vs. 0.4%）, while the proportions of keratinocyte subpopulations 5, 6, 10 and 12 （8.03%, 7.36%, 3.52%, 0.91%, respectively） in vitiligo lesions were significantly higher than those in the normal skin tissues （4.47%, 3.53%, 2.69%, 0.28%, respectively, all P < 0.01）. Moreover, the above keratinocyte subpopulations were at the end of cell differentiation, and expressed very significant and specific marker genes, which were mainly closely related to cell-cell interactions and cell homeostasis. GO and KEGG analysis showed that keratinocyte subpopulations 5 and 6 were mainly related to intercellular connection, cell adhesion and cytoskeleton function, while the keratinocyte subpopulation 10 was closely related to cell homeostasis. Conclusion The single-cell sequencing technology was firstly used to study the transcriptional expression profile of vitiligo lesions in China, and preliminary analysis revealed 4 groups of keratinocytes with different quantity and functions, suggesting that abnormal differentiation and dysfunction of keratinocyte subpopulations may affect the occurrence and development of vitiligo.

Key words: Vitiligo, Single-cell analysis, Transcriptome, Keratinocytes, Fibroblasts, Non-segmental vitiligo