Cellular heterogeneity in hypertrophic burn scars in response to carbon dioxide laser therapy

Ablative fractional carbon dioxide (AFCO₂) laser therapy is used for treating pathological scarring. However, mechanisms underlying reduction in hypertrophic scarring are poorly understood.

We investigated cellular mechanisms of AFCO₂ laser therapy by performing single-cell RNA sequencing (scRNA-seq) on skin biopsies from burn survivors with hypertrophic scars before and after three sessions of AFCO₂ therapy. Scar reduction was assessed subjectively and objectively.

Those with a good response (GR) to laser therapy have scars less than 6 years from injury, whereas poor responders (PR) have scars over 6 years since injury. ScRNA-seq analysis of skin biopsies reveals that genes enriched in GR are associated with extracellular matrix and structure organisation (COL14A1, POSTN, SPARC); whereas genes enriched in PR are related to enhanced immune inflammatory responses (CXCL14, JUN, TNC). Notably, expression of the pro-fibrotic gene Engrailed-1 (EN1) remains elevated in PR scars compared to GR. The ECM-regulatory gene TIMP-1 (Tissue Inhibitor of Metalloproteinases 1) is also significantly upregulated in PR scars following treatment relative to GR scars, although at the protein level TIMP1 levels decrease in PR but increase in GR after therapy. The regenerative-associated gene, TRPS1 (Transcriptional Repressor GATA Binding 1) expression demonstrates opposing regulation post treatment, with upregulation observed in GR scars but downregulation in PR scars, underscoring distinct transcriptional trajectories associated with differential therapeutic outcomes. Finally, distinct intercellular communication networks and differentiation trajectories are observed after AFCO₂, with regenerative mesenchymal fibroblasts predominating in GR but inflammatory fibroblasts associated with PR.

We conclude AFCO₂ laser therapy is more effective if done early after injury and distinct fibroblast recruitment is associated with a good response, specifically regenerative fibroblasts.