Hydrogen Sulfide Suppresses Keratinocyte Migration in Scratch Assay
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Cell migration is crucial for maintaining physiological functions in the body and is driven by mechanical, electrical, or chemical stimuli. Hydrogen sulfide (H 2 S), a gaseous signaling molecule, has been shown to regulate cellular migration in a cell type and context-dependent manner. Keratinocytes are the predominant cell-type in the epidermis and drive re-epithelialization during wound healing. However, the effect of H 2 S on keratinocyte migration remains incompletely defined. Here, we demonstrate that H 2 S suppresses keratinocyte migration in a serum-restricted in vitro scratch assay. Intracellular H 2 S levels were modulated using fast-releasing donors H 2 S donors (NaHS or Na 2 S) to achieve supraphysiological levels or by siRNA knockdown of endogenous H 2 S-producing enzymes cystathionine-γ-lyase (CGL) and cystathionine-β-synthase (CBS) to obtain infraphysiological levels. Treatment with NaHS or Na 2 S (10 µM–5 mM, 24 h) reduced gap closure in a dose-dependent fashion, with negligible effects at ≤ 100 µM and progressive suppression at higher concentrations. Conversely, siRNA knockdown of CGL or CBS boosted keratinocyte migration by 30% versus scrambled siRNA control. Supplementation with a slow-release H₂S donor rescued the siCGL phenotype, restoring migration to control levels. Across experiments, intracellular H₂S level is negatively associated with keratinocyte migration. These findings suggest that H₂S can restrain keratinocyte motility under serum-restricted conditions, and that reports of accelerated wound closure with H₂S donors in vivo may reflect effects on other cell types and/or inflammatory pathways rather than direct enhancement of keratinocyte migration.
Significance Statement
Dysfunctional wound healing affects millions worldwide. While H₂S donors can accelerate wound closure in animal models, the cell-type–specific effects remain unclear. We show that, H₂S suppresses keratinocyte migration implicating non-keratinocyte mechanisms (e.g., endothelial, immune, fibroblast) in the in vivo benefit of H₂S donors and refining where in the healing cascade H₂S is most likely to help.
