Fetal Fibroblast Heterogeneity Defines Dermal Architecture during Human Embryonic Skin Development

To investigate the heterogeneity of fibroblasts in human fetal skin, we analyzed published single-cell RNA-sequencing data (7 and 16 weeks after conception) and performed single-molecule FISH to map their spatial distribution and predicted dynamic interactions. Clustering revealed 8 fibroblast populations with changes in developmental stage-specific abundance. Proliferative cells (MKI67+) were present at all stages. The appearance of dermal papilla (PRDM1+) and hair follicle (SLC26A7+) fibroblasts coincided with hair follicle maturation, whereas fibroblasts (apolipoprotein E positive) specifically associated with blood vessels increased in abundance as the vessels developed. HOXC5 was a marker of the most abundant fibroblasts 7-8 weeks after conception; this cluster was diminished 9-13 weeks after conception and undetectable subsequently. A second population (PLAT+) decreased in abundance with the same kinetics. Fibroblasts corresponding to papillary dermis (GRP+) were predominant 9-13 weeks after conception, whereas reticular dermal fibroblasts (ASPN+) were the major cluster 14-16 weeks after conception. Partition-based graph abstraction and pseudotime analysis indicated that the HOXC5+ fibroblasts were closely connected with the papillary and hair follicle fibroblasts, whereas the PLAT+ fibroblasts were connected with reticular and vascular fibroblasts. Dermal papilla fibroblasts were the most highly differentiated. Integration of fetal and adult datasets distinguished the adult and fetal papillary clusters from the reticular clusters.