Granzyme B Exacerbates Atopic Dermatitis Through Filaggrin, E-cadherin And Desmoglein Cleavage And Disruption Of Barrier Function
Christopher Turner1, Stephanie Santacruz1, Matthew Zeglinski1, Christine Wang1, Katlyn Richardson1, Sho Hiroyasu1, Hongyan Zhao1, Yue Shen1, Gail Gauvreau2, Hermenio Lima2, Roma Sehmi2, David Granville1.
1University of British Columbia, Vancouver, BC, Canada, 2McMaster University, Hamilton, ON, Canada.
Granzyme B (GzmB) is a serine protease minimally expressed in normal skin but drastically elevated in multiple chronic inflammatory skin diseases. Extracellular GzmB cleaves multiple substrates within the extracellular matrix and dermal-epidermal junction, contributing to impaired wound healing. We hypothesize GzmB contributes to development, severity and impaired healing in atopic dermatitis (AD). To evaluate GzmB expression, excised human AD skin was examined histologically. GzmB’s role was assessed in a murine model of AD, comparing GzmB-/- and wild-type (WT) mice. Macroscopic severity scores, wound morphometry, inflammation and barrier protein expression were assessed. To identify additional GzmB substrates, in vitro cleavage assays were performed. To assess mechanism, cultured keratinocytes were exposed to GzmB with barrier function, barrier protein expression and cell viability assessed. GzmB was significantly elevated in human and mouse AD compared to healthy skin. A significant reduction in inflammation, lesion size, epidermal thickness and AD severity scores were observed in GzmB-/- compared to WT mice. Topical administration of a small molecule GzmB inhibitor to WT mice also reduced AD severity compared to vehicle-treated controls. Cultured keratinocyte monolayers exposed to GzmB showed a dose-dependent increase in barrier function impairment. In vitro, GzmB effectively cleaved Filaggrin, E-cadherin, desmoglein-1 and -3. Keratinocyte monolayers exposed to GzmB showed disordered E-cadherin, desmoglein-1 and -3 expression. As expected, epidermal expression of Filaggrin (stratum corneum only), E-cadherin, desmoglein-1 and -3 were all decreased in WT AD-affected mice compared to healthy skin controls. However, GzmB-/- AD-affected mice displayed a significantly greater expression of each of these proteins than WT AD-affected ears. GzmB is elevated in AD and contributes to disease severity. Mechanistically, GzmB disrupts barrier function, cleaving key proteins linked to the pathogenesis of AD. GzmB inhibition provides a potential therapeutic option.
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