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Laboratory Research
Ex vivo assays that utilize viable human skin are inherently physiological relevant models for studying wound healing. These models preserve many of the important cell types of human tissue, provide a full-thickness architecture, undergo native cellular interactions, and contain relevant extracellular matrix constituents. Moreover, they offer predictive insights into wound repair processes and provide a translational platform for evaluating therapeutics and wound care devices, potentially reducing reliance on animal studies or early-phase clinical trials. Importantly, ex vivo systems accommodate mechanistic investigations of epithelial regeneration, inflammation, and infection with relevant endpoint readouts such as histology, morphometric analyses, and omics-based bioanalytics. We present a case study illustrating the utility of this platform.
Methods:
An ex vivo viable human skin model of mature biofilm was utilized to study biodistribution and wound closure for cell-free amniotic fluid (cfAF) in viable tissue with morphometric, histological, and bioanalytical (LC-MS/MS) endpoints. A total of 24 skin samples were wounded and treated either by intradermal injection (Group 1) or by injection followed by daily topical applications (Group 2) over the course of 7 days. Skin was analyzed for wound closure (H&E), for cfAF localization (IHC), and LC-MS/MS was used to track a key peptide contained in cfAF.
Results:
The major finding from this pilot study show that cfAF treatments enhance re-epithelialization of wounded viable skin, with ~40% wound closure compared to controls (morphometric analysis), and results for cfAF injections showed clear dosing via immunohistochemistry. LC–MS/MS confirmed peptide recovery from tissue lysates and demonstrated suprabasal epidermal accumulation and moderate dermal penetration.
Discussion: This pilot study examined biodistribution of the serine protease, apha-1 antitrypsin (A1AT), known to contribute to the overall modulation of inflammation in wounds by neutralizing neutrophil elastase. These pilot results reveal that cfAF treatments decrease wound areas (morphometric analysis) and highlight differences in closure dynamics among conditions. IHC andLC–MS/MS highlightedd the specific biodistribution of the cfAF across the epidermis, and especially suprabasal and spinous layers, which indicates high A1AT accumulation. These results highlight the utility of the ex vivo human viable skin model to study wound healing, tissue responses and the ability to study cfAF delivery of bioactive factors to human skin layers to promote epithelial repair.