Re-Epithelialization

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.

Macrophages and inflammation play a beneficial role during wound repair with macrophages regulating a wide range of processes, such as removal of dead cells, debris and pathogens, through to extracellular matrix deposition re-vascularisation and wound re-epithelialisation. To perform this range of functions, these cells develop distinct phenotypes over the course of wound healing. They can present with a pro-inflammatory M1 phenotype, more often found in the early stages of repair, through to anti-inflammatory M2 phenotypes that are pro-repair in the latter stages of wound healing. There is a continuum of phenotypes between these ranges with some cells sharing phenotypes of both M1 and M2 macrophages. One of the less pleasant consequences of quick closure, namely the replacement with scar tissue, is also regulated by macrophages, through their promotion of fibroblast proliferation, myofibroblast differentiation and collagen deposition. Alterations in macrophage number and phenotype disrupt this process and can dictate the level of scar formation. It is also clear that dysregulated inflammation and altered macrophage phenotypes are responsible for hindering closure of chronic wounds. The review will discuss our current knowledge of macrophage phenotype on the repair process and how alterations in the phenotypes might alter wound closure and the final repair quality.

The skin, which serves as the first barrier of the human body, is particularly susceptible to exogenous injuries. Skin wounds, including acute burns and chronic non-healing ulcers, are commonly observed in clinics. Healing of skin wounds is a complex process, consisting of infiltration of inflammatory cells, cellular proliferation, and tissue remodeling phases, which restore the integrity and functions of the skin. Epithelialization is involved in wound healing through re-establishing an intact keratinocyte layer. Epidermal stem cells are indispensable for epithelialization, and they are regulated by multiple proinflammatory cytokines or growth factors. In this review, we summarize recent advances in the effect of these cytokines on migration, proliferation, and differentiation processes of epidermal stem cells. We also introduce promising therapeutic strategies targeting epidermal stem cells or related proinflammatory cytokines for patients with skin wounds.

Wound healing restores tissue integrity through tightly coordinated cellular and molecular programs, yet the biological processes that persist after apparent closure remain largely unexplored. Using a standardized mouse hard palate injury model combined with histological, lineage-tracing, molecular, and epigenetic analyses, we show that rapid re-epithelialization is followed by months of cellular remodeling. Although wounds re-epithelialized within two weeks, temporal analyses revealed that the regenerated epithelium maintained leading-edge-like features for months. Structural reorganization, including increased cell density and epithelial thickness, persisted well beyond closure, accompanied by sustained proliferation and elevated apoptosis mediated by the p53-p21 pathway, thereby removing cells with DNA damage during tissue remodeling. Lineage tracing revealed marked alterations in Wnt-responsive epithelial stem cells, which were transiently lost from the repaired epithelium and gradually repopulated the tissue over approximately four months. Repair-activated keratinocytes showed sustained expression of keratin 6 and keratin 17 and delayed recovery of differentiation markers keratin 10 and filaggrin, indicating prolonged epithelial activation despite morphological closure. Coordinated alterations in histone marks H2AK119ub, H3K27ac, and H3K27me3, and persistent immune infiltration, indicate lasting molecular and microenvironmental remodeling that maintains the tissue in a memory-like state. Together, these findings indicate that oral wound healing is an extended, multi-phase process in which post-healing remodeling restores epithelial homeostasis and ensures proper resolution of the repair response.