Differentiation Between Vascularized Composite Allograft Acute Skin Rejection and Delayed Type Hypersensitivity Reactions Based on Cytokine Analysis

Background: Skin rejection in vascularized composite allotransplantation is a major hurdle for wider clinical adoption in VCA. Histopathological alterations during acute skin rejection mimic inflammatory skin processes and are difficult to differentiate. The aim of this study was to identify protein expression patterns characteristic for acute skin rejection in reference to inflammatory skin processes, such as delayed type hypersensitivity reactions. Methods: Limb transplantations were performed using a Brown-Norway to Lewis rat allotransplant model. For the induction of skin inflammation a contact hypersensitivity (CHS) and delayed type hypersensitivity (DTH) model in Lewis rats were established. CHS and DTH result in skin irritation and swelling comparable to grade I-II rejection in our rat allotransplant model. While the DTH reaction is limited to the plantar skin, the CHS model allows for comparison with rejection also in hairy skin. Skin biopsies were taken at defined time points, where a histologically identical inflammatory response in all study groups was present. Protein levels of 14 inflammatory cytokines were assessed by Luminex™. Results: Based on the multivariate linear discriminant analysis, IL-12p70 and TNF-α were the major discriminators between skin rejection and inflammatory skin reactions. Principal component analysis identified IL-1α, GM-CSF, IL-6 and IL-18 as key drivers of rejection at this time point. Conclusion: The novel diagnostic platform enables for early and specific diagnosis of rejection and differentiation toward inflammatory skin diseases in this transplant model.