**Background:** The skin is the largest organ and serves as a protective barrier. Disruption of this barrier can lead to infection, making rapid wound healing critical. Very-long-chain fatty acids (VLC-FA, ≥24 carbons) are synthesized endogenously by the ELOVL4 enzyme, and mutations in ELOVL4 cause severe skin disorders and defective permeability barriers, indicating their essential role. While n-3 long-chain PUFAs (e.g., EPA, DHA) are known to improve skin barrier function and wound healing, the specific functions of n-3 very-long-chain PUFAs (VLC-PUFAs) in skin remain unclear. This study aimed to investigate whether dietary supplementation with n-3 VLC-PUFAs affects skin fatty acid composition and development in Atlantic salmon, and to elucidate their effects on cell migration in vitro using primary salmon keratocytes and human dermal fibroblasts.
**Methods:** A 4-week feeding trial was conducted with Atlantic salmon parr (initial weight ~6 g, final ~11 g) fed five isoenergetic diets containing increasing levels (0%, 2.5%, 5%, 7.5%, 10%) of a VLC-PUFA concentrate (VLC-Conc1), while EPA and DHA levels were kept constant across diets. Each diet was fed to three tanks with 100 fish per tank. Skin samples were collected at days 0, 18, and 28 for fatty acid composition analysis (phospholipid and triacylglycerol fractions) and histology (H&E and Von Kossa staining). For in vitro studies, primary keratocytes from Atlantic salmon scales were cultured with 10 µM or 20 µM of the VLC-PUFA 26:6 n-3, and cell migration from scales was assessed at 24, 30, and 57 hours. Human dermal fibroblasts (ATCC PCS-201-012) were treated with 3 µM VLC-Conc2 or 3 µM DHA, and a scratch assay was performed to evaluate cell migration over 14 hours. Gene expression (ELOVL4, CERS2, CPT1A, FGF2, TGFA, VEGFA1, IL8) was analyzed by qPCR at 0 hours and 1 day post-scratch.
**Key Results:** In vivo, dietary VLC-PUFA supplementation led to a linear increase in n-3 VLC-PUFA levels in skin phospholipids, with 28:8 n-3 increasing from 14.18 µg/g tissue (0% group) to 395.50 µg/g tissue (10% group) (P < 0.001). Epidermis thickness at day 18 was significantly greater in the 10% group (42.08 ± 1.84 µm) compared to the 2.5% group (28.96 ± 1.30 µm) (P < 0.0001), and at day 28, the 10% group (44.79 ± 2.09 µm) was thicker than the 2.5% group (34.30 ± 1.59 µm) (P = 0.0073). Mucus cell count at day 18 was higher in the 10% group (4.6 per 100 µm) versus the 2.5% group (2.4 per 100 µm) (P = 0.0002). Von Kossa staining revealed darker (more mineralized) scales in the 10% group compared to the 0% group. In vitro, salmon keratocyte migration from scales was significantly higher in both VLC-PUFA groups (10 µM and 20 µM) and the FGF group compared to control at 24 h (P = 0.01), 30 h (P = 0.03), and 57 h (P = 0.02). In human dermal fibroblasts, the VLC-Conc2 group showed a trend toward faster scratch closure (approximately 10% smaller scratch size at 14 h post-scratch) compared to control and DHA groups, though not significant (P = 0.2597). Gene expression at 0 h post-scratch showed significantly higher CPT1A (P < 0.0001) and VEGFA1 (P = 0.0044) in the VLC-Conc2 group, and significantly lower ELOVL4 in the DHA group (P = 0.0110). At 1 day post-scratch, CERS2 was significantly lower in control and VLC-Conc2 groups versus DHA (P = 0.0086), FGF2 was significantly lower in VLC-Conc2 (P = 0.0008), TGFA was significantly higher in VLC-Conc2 (P = 0.0028), and IL8 was significantly lower in VLC-Conc2 (P = 0.0206).
**Clinical Implications:** This study provides the first evidence that dietary n-3 VLC-PUFAs are deposited in skin and promote skin development (increased epidermis thickness, mucus cells, and scale mineralization) in Atlantic salmon. The in vitro findings suggest that VLC-PUFAs enhance cell migration in both fish keratocytes and human dermal fibroblasts, potentially accelerating wound healing. The gene expression changes (e.g., increased CPT1A and VEGFA1, modulated IL8) indicate that VLC-PUFAs may influence fatty acid oxidation, angiogenesis, and inflammatory responses during wound repair. These results have implications for aquaculture, where robust skin is critical during smoltification, and for human dermatology, where VLC-PUFA supplementation could be explored as a therapeutic strategy for impaired wound healing or skin barrier disorders. However, further in vivo studies are needed to confirm clinical relevance.