In skin research, 2D monolayer in vitro models provide only limited fields of application as they do not sufficiently mimic physiological properties. Since in the spirit of 3R’s (Replacement, Reduction, and Refinement) animal models are to be avoided, new approaches like 3D skin equivalents (SE) are needed to close the in vitro/in vivo gap. Cell culture inserts to produce SE are commercially available, however, these inserts are expensive and limited regarding the experimental setup.
The aim of this study was to design and produce novel cell culture inserts fabricated on commercially available 3D-printers to generate full thickness human skin equivalents. A computer-aided design model was realized with extrusion-based 3D printing of polylactic acid filament. In consecutive steps the design of the inserts was improved and the feasibility confirmed in cell culture experiments. Cytotoxic effects of the final product were excluded by testing the inserts according to ISO-norm procedures. Finally, to verify usability, full thickness human skin equivalents were created on collagen/fibrin dermal scaffolds which were placed in the inserts. Histologic analysis proved the comparable quality of the constructs compared to commercially available products. In conclusion, here we demonstrate an efficient and cost-effective alternative to produce 3D-printed inserts for the generation of skin equivalents. The system is feasible with common 3D printers and allows high flexibility in the choice of printing materials, membrane materials as well as of shape and size. It is compatible with commonly used cell culture plates and easy to handle. Further studies will be performed with skin equvalents based on the presented technology.