Regeneration Of Merkel Cells In Engineered Skin Substitutes Grafted To Mice
Dorothy M. Supp1, Jennifer M. Hahn1, Kevin L. McFarland1, Kelly A. Combs1, Andrea L. Lalley1, Christopher M. Lloyd1, Steven T. Boyce2.
1Shriners Hospitals for Children - Cincinnati, Cincinnati, OH, USA, 2University of Cincinnati College of Medicine, Cincinnati, OH, USA.
Background: Engineered skin substitutes (ESS) containing primary human fibroblasts and keratinocytes were shown to provide long-term closure of excised full-thickness burn wounds, but relatively little is known about innervation of ESS. Merkel cells are specialized neuroendocrine cells of the epidermis that are required for light touch sensation. The goal of this study was to begin to characterize innervation of ESS and determine if Merkel cells are present after grafting to wounds. Methods: ESS were prepared with primary fibroblasts and keratinocytes, isolated from adult human skin samples obtained with IRB approval, and were grafted to full-thickness wounds in immunodeficient mice. Biopsies were collected at multiple time points for analysis by immunohistochemistry. Results: Cells positive for Merkel cell markers keratin 18 (KRT18) and keratin 20 (KRT20) were identified in the basal epidermis of ESS by 4 weeks after grafting, suggesting the presence of Merkel cells. These cells were positive for human leukocyte antigen (HLA-ABC), confirming their human origin. Fibers staining positive for the neuronal markers NCAM and PGP9.5 were found in apposition to KRT18/KRT20-positive epidermal cells by 16 weeks after grafting, suggesting association of Merkel cells with neurons. Conclusions: The results suggest that Merkel cells were regenerated in ESS following transplantation to mice. Although we hypothesize that Merkel cells were derived from precursors present in primary keratinocyte cultures, we are currently unable to rule out the presence of rare Merkel cells in keratinocyte cultures in vitro prior to preparation of ESS. The results suggest that fine touch perception may be regained in healed ESS, although this must be confirmed with additional studies analyzing nerve function.
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