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To protect the human skin from extensive solar radiation, melanocytes produce melanin and disperse it via melanosomes to keratinocytes in the basal and suprabasal layers of the human epidermis. Moreover, melanocytes are associated with pathological skin conditions such as vitiligo and psoriasis. Thus, an in vitro skin model that comprises a defined cutaneous pigmentation system is highly relevant in cosmetic, pharmaceutical and medical research. Here, we describe how the epidermal-melanin-unit can be established in vitro. Therefore, primary human melanocytes are implemented in an open source reconstructed epidermis. Following 14 days at the air liquid interface, a differentiated epidermis was formed and melanocytes were located in the basal layer. The functionality of the epidermal-melanin-unit could be shown by the transfer of melanin to the surrounding keratinocytes, and a significantly increased melanin content of models stimulated with either UV-radiation or the melanin precursor dihydroxyphenylalanine. Additionally, an UV50 assay was developed to test the protective effect of melanin. In analogy to the IC50 value in risk assessment, the UV50 value facilitates a quantitative investigation of harmful effects of natural UV-radiation to the skin in vitro. Employing this test, we could demonstrate that the melanin content correlates with the resilience against simulated sunlight, which comprises 2.5 % UVB and 97.5 % UVA. Besides demonstrating the protective effect of melanin in vitro, the assay was used to determine the protective effect of a consumer product in a highly standardized setup.

There is a great need for valuable ex vivo models that allow for assessment of cartilage repair strategies to reduce the high number of animal experiments. In this paper we present three studies with our novel ex vivo osteochondral culture platform. It consists of two separated media compartments for cartilage and bone which better represents the in vivo situation and enables supply of specific factors to the different needs of bone and cartilage. We investigated whether separation of the cartilage and bone compartments and/or culture media results in the maintenance of viability, structural and functional properties of cartilage tissue (study A). Next, we evaluated for how long we can preserve cartilage matrix stability of osteochondral explants during long-term culture of 84 days (study B). Finally, we investigated what the optimal defect size is without spontaneous self-healing occurring in this culture system (study C). It was demonstrated that separated compartments for cartilage and bone in combination with tissue-specific medium allows for long-term culture of osteochondral explants, while maintaining cartilage viability, matrixtissue content, structure and mechanical properties up to at least 56 days. Furthermore, it was shown that we can create critical size cartilage defects of different sizes in the model. The osteochondral model represents a valuable preclinical ex vivo tool for studying clinically relevant cartilage therapies, such as cartilage biomaterials for their regenerative potential, evaluation of drug and cell therapies or to study mechanisms of cartilage regeneration, which will undoubtedly reduce the number of animals needed for in vivo testing.

Surgical implantation of a biomaterial triggers foreign-body-induced fibrous encapsulation. Two major mechanisms of this complex physiological process are (I) chemotaxis of fibroblasts from surrounding tissue to the implant region, followed by (II) tissue remodeling. As an alternative to animal studies, we here propose a process-aligned \({in}\) \({vitro}\) test platform to investigate the material dependency of fibroblast chemotaxis and tissue remodeling mediated by material-resident macrophages. Embedded in a biomimetic three-dimensional collagen hydrogel, chemotaxis of fibroblasts in the direction of macrophage-material-conditioned cell culture supernatant was analyzed by live cell imaging. A combination of statistical analysis with a complementary parameterized random walk model allowed quantitative and qualitative characterization of the cellular walk process. We thereby identified an increasing macrophage-mediated chemotactic potential ranking of biomaterials from glass over polytetrafluorethylene to titanium. To address long-term effects of biomaterial-resident macrophages on fibroblasts in a three-dimensional microenvironment, we further studied tissue remodeling by applying macrophage-material-conditioned medium on fibrous \({in}\) \({vitro}\) tissue models. A high correlation of the \({in}\) \({vitro}\) tissue model to state of the art \({in}\) \({vivo}\) study data was found. Titanium exhibited a significantly lower tissue remodeling capacity compared to polytetrafluorethylene. With this approach, we identified a material dependency of both chemotaxis and tissue remodeling processes, strengthening knowledge on their specific contribution to the foreign body reaction.