Thank you for your question! Yes, we performed pseudo-temporal ordering of all fibroblasts in our data set and overlaid RNA velocity vectors of those cells to better inform their trajectories. We found three distinct trajectories, all pointing toward regions high in Acta2/Tagln, which are markers of myofibroblasts (i.e. contractile cells). These results suggest that the distinct fibroblast subtypes all have the potential to acquire a fibrotic state. We also identified genes and transcription factors that are pseudo-temporally dependent, and found also TGF-beta genes to be changing. Currently, we are developing a pipeline for inferring cell-cell interactions, network structure and function of cells based on transcript abundance in single cells and one of the goals is to see what pathways change in these and other processes in latent space. If you have any further questions you can contact me at cfguerre@uci.edu.

Thank you for your question! Keloid and hypertrophic scars are fibrotic in nature, they have high amounts of collagen-producing fibroblasts (i.e. myofibroblasts) and, hence, have high amounts of collagen bundles. This could inherently affect the micro- and macro-environment of the wound and diminish skin integrity. Currently, we are co-opting a mouse model of keloid scars to investigate other aspects that may be limiting the natural function of non-pathological fibroblasts, such as metabolism.

Thanks! Yes, we used day 12 wounds for a couple of reasons. First, this time point is prior hair follicle and fat regeneration, which occurs 15 and 21 days after pounding, respectively. Second, this time point is prior complete wound closure - which corresponds with active cellular migration into the wound bed. We therefore reasoned that during this time point there would be increased cellularity and we might see a large pool of heterogeneous cell populations.

Thank you for your question! This is an active area of research that I am currently pursuing.

Thank you for your question. Yes, this is a recurring and important question. Mouse and human hair follicles (and also skin!) are very different not only in the density of follicles, but also in their structure and cycle dynamics. In our study, we use the wound induced hair neogenesis system as a model to study cell reprogramming during wound healing, and use hair follicle and fat regeneration as hallmarks of successful skin regeneration. Despite these differences, there have been reports of humans regenerating facial hairs after dermabrasion, and we ourselves have also shown that what we learned from mouse studies can be translated into humans by successfully reprogramming fat cells in vitro using scalp hair follicles and other molecules.

Excellent question! We would like to incorporate distinct fibroblast subsets and immune cells into the model (i.e. based on gene expression differences from single cell RNA-seq, proliferation dynamics, etc) and determine if heterogeneity can better inform, predict and resolve wound healing outcomes.