Acute effects of Cold Atmospheric Plasma on bottlenose dolphin skin fibroblasts: Implications for wound healing in cetaceans

Skin lesions are common in cetaceans and are challenging to treat due to environmental exposure and difficulties in administering therapies. Cold Atmospheric Plasma (CAP) is a non-invasive approach that may promote wound healing and reduce microbial infections through the generation of reactive oxygen and nitrogen species. Dermal fibroblasts play a central role in tissue repair, particularly in extracellular matrix production and wound contraction, making them key targets to evaluate CAP's therapeutic potential. In this study, a dermal fibroblasts cell line from bottlenose dolphins was exposed to short exposure (1, 2, 5 min) and long exposure (10 min) and analyzed at 0, 4, and 8 h post-treatment. Cell viability and cell cycle dynamics were assessed using MTT assays and high-content imaging. High-content imaging was employed to quantitatively evaluate cell cycle distribution and nuclear morphology through automated image analysis. Short CAP exposure enhanced cell cycle progression, increasing the proportion of cells in S and G2/M phases, whereas prolonged exposure induced early cell cycle alterations and led to nuclear collapse. These results indicate that CAP effects on fibroblasts are time-dependent with short exposures promoting early pro-survival and cell cycle-related responses, and longer exposures inducing cytotoxicity. Further studies are required to determine whether these early responses translate into sustained proliferative or regenerative effects and to optimize CAP protocols for potential therapeutic applications in cetacean skin repair.