Paired single-cell profiling reveals epithelial–stromal remodeling after ablative fractional carbon dioxide laser treatment in keloids

Background Keloids are fibroproliferative scars, but the cellular responses to ablative fractional carbon dioxide laser (AFCO2) treatment remain poorly defined. This study aimed to characterize treatment-associated epithelial, stromal and immune remodeling in patient-matched human keloid tissue. Methods We jointly reprocessed four previously reported untreated hypercellular-zone PRE specimens and four newly generated patient-matched POST specimens obtained 35 days after AFCO2 treatment from the same patients. All specimens were processed using the same Singleron Matrix/GEXSCOPE single-cell platform. Patient-aware pseudobulk differential expression, continuous transcriptional-program analysis, gene-set enrichment, trajectory and regulatory modeling, and cell–cell communication inference were integrated with independent reanalysis of GSE320017 and quantitative multiplex immunofluorescence and CD74 immunohistochemistry. The patient was the biological and inferential unit. Results After quality control, 74,937 cells were retained. A KRT6-dominant keratinocyte program increased in all four patients; KRT6B increased by 4.44 log2 units (false-discovery rate, 2.65 × 10 −5 ), with concordant increases in KRT6A and KRT16. Fibroblasts shifted toward an interferon/inflammatory program and away from a homeostatic extracellular-matrix program, whereas myeloid/APC cells showed lower HLA-DRA, C3 and MHC-II-associated antigen-presentation programs. Nine patient-supported mural-to-dendritic-cell or mural-autocrine communication axes weakened after treatment. Quantitative tissue imaging showed that CK6-family-positive area within CK14-defined epithelium increased from 2.70% to 65.84%, MMP10-positive area overlapping CK14-defined epithelium increased from 8.49% to 25.92%, and CD74-positive cell density decreased from 2,530.35 to 695.50 cells/mm². The independent hypertrophic-scar dataset showed context-dependent rather than uniform AFCO2 responses. Conclusions AFCO2 treatment was associated with coordinated epithelial, stromal and immune remodeling in human keloids, centered on a KRT6-dominant epithelial program, fibroblast-state redistribution and attenuation of an MHC-II-associated tissue context. These patient-consistent findings are exploratory and do not establish causal mechanisms.