Assessment of morphological and immunohistological alterations in long-term keloid skin explants

Abstract

One of the major impediments in keloid research is the lack of a keloid animal model that can mimic human keloid. This imposes investigative constraints on studying cellular interactions and biochemical processes that normally occur in vivo. Our main objective is to establish an in vitro model for maintaining long-term viable keloid dermal explants as a tool for investigating the pathogenesis of keloid scar formation. Explants of adult keloid scars were cultured in vitro by embedding them in enriched collagen gel matrix and maintaining them for up to 6 weeks, whereupon changes in tissue morphology and cellular differentiation were examined. The effects of medium enrichment, air versus liquid submersion, and different substrates on the explants were examined. Our results indicated that keloid explants embedded in a collagen gel matrix were morphologically better preserved than explants placed on a plastic substrate. Explants with epidermis at the air-liquid interface had better morphology than collagen-submerged explants, and there were no differences between serum-free and serum-supplemented explant cultures. Immunohistochemical and apoptotic analyses were performed to assess cellular viability and differentiation. In situ hybridization confirmed that keloid fibroblasts had sustained collagen type I gene expression throughout the 6 weeks in culture, thus validating the integrity of a long-term keloid culture system. In conclusion, the collagen-embedded skin explant system demonstrates that keloid tissues could be maintained for up to 6 weeks for long-term in vitro studies.