Direct biological effects of fractional ultrapulsed CO₂ laser irradiation on keratinocytes and fibroblasts in human organotypic full-thickness 3D skin models

Molecular effects of various ablative and non-ablative laser treatments on human skin cells—especially primary effects on epidermal keratinocytes and dermal fibroblasts—are not yet fully understood. We present the first study addressing molecular effects of fractional non-sequential ultrapulsed CO₂ laser treatment using a 3D skin model that allows standardized investigations of time-dependent molecular changes ex vivo. While histological examination was performed to assess morphological changes, we utilized gene expression profiling using microarray and qRT-PCR analyses to identify molecular effects of laser treatment. Irradiated models exhibited dose-dependent morphological changes resulting in an almost complete recovery of the epidermis 5 days after irradiation. On day 5 after laser injury with a laser fluence of 100 mJ/cm², gene array analysis identified an upregulation of genes associated with tissue remodeling and wound healing (e.g., COL12A1 and FGF7), genes that are involved in the immune response (e.g., CXCL12 and CCL8) as well as members of the heat shock protein family (e.g., HSPB3). On the other hand, we detected a downregulation of matrix metalloproteinases (e.g., MMP3), differentiation markers (e.g., LOR and S100A7), and the pro-inflammatory cytokine IL1α.

Overall, our findings substantiate the understanding of time-dependent molecular changes after CO₂ laser treatment. The utilized 3D skin model system proved to be a reliable, accurate, and reproducible tool to explore the effects of various laser settings both on skin morphology and gene expression during wound healing.