Single-cell RNA-sequencing of dermal fibroblasts demonstrates culture-induced changes and variable persistence of...

In vitro models of scarring and fibrosis are essential to improve our understanding of disease mechanisms and ultimately develop much-needed therapeutic strategies. The emerging appreciation of fibroblast heterogeneity leaves a knowledge gap about what is represented in typical fibroblast cultures: are the quantitative differences in fibroblast subtypes observed in pathological tissues represented, and are disease-associated molecular alterations of subtypes maintained? Single-cell (sc) RNA-seq on patient-matched keloid and adjacent non-lesional dermis was compared to sc- and bulk-RNA-seq of fibroblast cultures after 4+ passages. After culture, fibroblast subtypes assimilated, with clustering distinct from the original populations. Pseudo-bulk analysis of non-cultured mesenchymal fibroblasts showed cell-intrinsic keloid-versus-control transcriptional differences consistent with disease understanding; however, only a subset of these persisted in vitro. Cell-cell communication analysis provides insight into potential strategies to maintain cell populations and their in vivo phenotypes. This work provides a greater understanding of, and potential strategies to refine, essential human fibroblast culture models.