Abstract
A large body of literature is available on wound healing in humans. Nonetheless, a standardized ex vivo wound model without disruption of the dermal compartment has not been put forward with compelling justification. Here, we present a novel wound model based on application of negative pressure and its effects for epidermal regeneration and immune cell behaviour. Importantly, the basement membrane remained intact after blister roof removal and keratinocytes were absent in the wounded area. Upon six days of culture, the wound was covered with one to three-cell thick K14+Ki67+ keratinocyte layers, indicating that proliferation and migration were involved in wound closure. After eight to twelve days, a multi-layered epidermis was formed expressing epidermal differentiation markers (K10, filaggrin, DSG-1, CDSN). Investigations about immune cell-specific manners revealed more T cells in the blister roof epidermis compared to normal epidermis. We identified several cell populations in blister roof epidermis and suction blister fluid that are absent in normal epidermis which correlated with their decrease in the dermis, indicating a dermal efflux upon negative pressure. Together, our model recapitulates the main features of epithelial wound regeneration, and can be applied for testing wound healing therapies and investigating underlying mechanisms.
Highlights
A large body of literature is available on wound healing in humans
We have efficaciously applied for the first time negative pressure to ex vivo human skin leading to the formation of typical blisters
Upon removal of the blister roof, we found neither damage of Col-IV nor epidermal components on the suction blister floor (=wound bed) which is in line with several studies showing by electron microscopic and histochemical analysis that the basement membrane stays intact after suction blister formation[14,20,22]
Summary
A large body of literature is available on wound healing in humans. a standardized ex vivo wound model without disruption of the dermal compartment has not been put forward with compelling justification. Incisional ex vivo human skin wounds created with a scalpel or partial-thickness wounds initiated with a small biopsy punch were shown to re-epithelialize[9,10] Unlike these models, where the basement membrane and partially the dermal structure are disrupted, several dermal-epidermal separation methods have been established providing a better basis to study re-epithelialization[11]. The blister roof consisted of a viable epidermis including the keratinocyte basal cell layer while leaving the basement membrane intact[15,16]. We have successfully utilized the device on human ex vivo skin and present that its application is comparable to in vivo experiments, and is a standardized, consistent and reproducible model, recapitulating the main features of epidermal wound regeneration
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