186 Chapter 6 Regenerative capacity of MatriDerm-based FSEs was similar to DED-based FSEs and ex vivo human skin at 2 weeks after burn injury To investigate the ability of the FSEs to function as burn injury models, we subjected the FSEs to a thermal contact injury of 80 °C for 20 s. In preliminary experiments we found that an injury with these settings caused sufficient damage to induce re-epithelization in similar models. Skin morphology and histology of the models were studied at T0, T + 1 week, and T + 2 weeks (Figure 3 and Supplementary Figure 2). At the macroscopic level, burn injuries presented in a rectangular shape which remained visible throughout the duration of the culture. In all models, destruction and release of the epidermis was clearly visible on the H&E-stained sections at T0. At T + 1 week and T + 2 weeks, re-epithelization of the epidermis was apparent in the ex vivo human skin model and the DED- and MatriDerm-based FSEs. In the Mucomaix-based model, re-epithelization did not take place. Although this model was clearly damaged, formation of a neo-epidermis was not detected. Figure 3. Effect of burn injury on cultured skin models. Images of ex vivo human skin (left) and full skin equivalents generated from de-epidermalized dermis, MatriDerm and Mucomaix (right). Models were produced from 3 different skin donors in duplicate. For the full skin equivalent models, T0 was after the initial 3 weeks of culture. Black scale bar = 100 µm. Next, we validated the regenerative capacity of the models. Therefore, we studied the presence of proliferating cells in the neo-epidermis and quantified the length of the re-epithelized epidermis (Figure 4). Ki67 staining revealed proliferating cells in and nearby the re-epithelized area in DED- and MatriDerm-based FSEs and the ex vivo human skin model (Figure 4A). In Mucomaix-based FSEs, there were hardly any positive cells present. BrdU was used to study 24 h proliferation in the models (Figure 4B). BrdUpositive keratinocytes were present in the newly formed basal layer of the ex vivo
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