Canine skin organ culture under serum free conditions: morpho-physiology and mast cell behaviour in response to degranulating and modulatory stimuli

ntroduction: Although canine skin models are alreadyavailable as either monocellular or organotypic cultures,they only partly recapitulate normal skin organization andfunction. Skin full thickness cultures may offer uniqueopportunities to study skin morpho-physiology in a bio-logically relevant 3D environment, complying with theprinciples of the 3Rs in animal research. Moreover, theymay provide insight in the effects of various agents on theskin. Mast cells (MCs) are tissue-homing leukocytes, den-sely packed with metachromatic granules, containinghighly active biological compounds (e.g., cytokines, growthfactors) that can be rapidly released upon stimulation. Oneof the most recently discovered regulatory mechanism ofMC degranulation relies on palmitoylethanolamide (PEA),an endogenous lipid compound and an endocannabinoid-like molecule. Recently, a clinical study has shown the effi-cacy and safety of PEA in reducing pruritus and skinlesions in dogs with atopic dermatitis.Materials and Methods: Skin biopsies sampled from cli-ent-owned dogs presented for surgery were cultured intriplicate, under submerged conditions, in Williams’ Emedium supplemented with penicillin/streptomycin,insulin, hydrocortisone and glutamine. Experiments wereperformed to evaluate the effect of ultramicronized PEA(30 lM) on skin viability and mast cell degranulationinduced by compound 48/80 (10 lg/mL). One-wayANOVA in conjunction with Bonferroni’s Multiple Com-parison Test was used for statistical comparisons.Results: Compared to vehicle, PEA treatment did not induce any epidermal nor dermal morphological changes suggestive of damage. Compound 48/80 induced MC degranulation and PEA treatment resulted in a significant decrease in the percentage of degranulating MCs (P < 0.05) without effect on MC density (number per mm2).Conclusion: The study corroborated the use of canine skin organ cultures as useful models for studying morpho-physiology features of the skin and its responses to various agents. It also confirmed previous data on the lack of adverse effects of PEA and its ability to down-modulate skin MC degranulation.