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ICE Krakow

ICE Krakow

ul. Marii Konopnickiej 17 30-302 Kraków


Cervantes, Diana (CELLINK)


As a result of animal welfare activism and public pressure, policy makers stablished a cosmetic ban for testing and marketing of finished cosmetic products and cosmetic ingredients tested on animals. This motivated the advancement of the scientific progress in this area and with it the creation of alternative models, specially of skin tissue since they are used in regulatory testing as well as in regenerative medicine applications. These skin models are a replacement to the use of animal models. Despite their great impact, there is an increasing need for models that allow the study of more complex physiological and pathological conditions. In this regard, 3D bioprinting offers the opportunity to create structures that resemble the histology of native tissues by allowing controlled spatial cell deposition, in combination of the use of relevant biomaterials, therefore, it is an important technique to achieve complexity in tissue models.
In general, some previously reported 3D bioprinted skin models and alternative skin models still contain “hidden” animal-derived components. In this study, we have focused on creating a 3D bioprinted skin model avoiding the use of animal-derived components. The skin model was created using a cellulose based bioink and the validation of the model was performed using animal-free reagents. The 3D bioprinted skin model was studied to determine its resemblance to native skin tissue.
Human dermal fibroblasts (HDFs) and normal human epidermal keratinocytes (NHEK) were expanded under 2D conditions prior to bioprinting. 3D bioprinting was performed using a BIO X bioprinter, HDFs formed the dermal layer and NHEKs the epidermal layer. The 3D bioprinted skin constructs were cultured for 14 days, of which the last 7 days, were in an air-liquid interface. The samples were analyzed on day 14. Cell health in the 3D construct was determined using live/dead assay. Analysis to assess the resemblance of the 3D model to native skin was performed using epidermal cell markers.
The 3D bioprinted constructs presented high cell viability, suggesting the bioink and bioprinting process did not have a negative effect on the cells. The histological data suggest different epidermal layers were formed including the spinous layer, indicating that the keratinocytes reached the final differentiation stage in the epidermis. The low expression of cytokeratin 10 and high expression of cytokeratin 14 suggests the keratinocytes had shifted from a proliferative to differentiated state.
Our findings suggest that the present 3D bioprinted skin model does resemble the native skin epidermis. Using non-animal-derived reagents is a feasible alternative in the creation of alternative models, it enables the development of models that can replace animal models while avoiding the use of animal derived components. Our 3D bioprinted model could be used as a foundation to generate skin models for specific diseases and as an alternative for the use of animal models for cosmetic and drug testing.

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