Background: While once believed to be a pro-apoptotic serine protease, in recent years, the role of Granzyme K (GzmK) has been redefined due to its role in augmenting inflammation. Found at low levels in the plasma of healthy individuals, extracellular GzmK levels are markedly elevated in response to certain infections and sepsis. As burn injuries invoke a large inflammatory response, we hypothesized GzmK is increased in burn injury and contributes to prolonged inflammation and impaired wound healing.
Methods: To evaluate GzmK expression, excised human burn biopsies were examined histologically. The role of GzmK was assessed in a murine model of thermal injury using wild-type or GzmK-/- mice. Wound morphometry, inflammation, Masson’s Trichrome, Collagen I/III ratio and skin tensiometry were assessed. To assess a direct role for GzmK on cutaneous inflammation, cultured keratinocytes were exposed to GzmK and cell viability, cytokine expression and in vitro wound healing was assessed.
Results: GzmK+ cells were significantly elevated in human burns compared to unwounded skin. Macrophages were the predominant cell type expressing GzmK, with classically activated M1 macrophages primarily responsible for both GzmK expression and secretion. A significant reduction in wound area, an increase in wound maturation and tensile strength was observed compared to equivalent burns in wild-type mice. Importantly, re-epithelialization showed the greatest degree of improvement of all parameters investigated, indicating keratinocytes may be especially susceptible to GzmK during healing. Cultured keratinocytes exposed to GzmK demonstrated impaired wound healing in vitro and a dose-dependent increase in pro-inflammatory cytokine release, including IL-6, which operated through a PAR-1 mediated pathway. GzmK-mediated cytotoxicity was not observed.
Conclusion: GzmK contributes to impaired wound healing, potentially by prolonging the pro-inflammatory stage of wound healing.