Abstract
Bioprinting is an emerging technique for the fabrication of living tissues that allows cells to be arranged in predetermined three-dimensional (3D) architectures. However, to date, there are limited examples of bioprinted constructs containing multiple cell types patterned at high-resolution. Here we present a low-cost process that employs 3D printing of aqueous droplets containing mammalian cells to produce robust, patterned constructs in oil, which were reproducibly transferred to culture medium. Human embryonic kidney (HEK) cells and ovine mesenchymal stem cells (oMSCs) were printed at tissue-relevant densities (107 cells mL−1) and a high droplet resolution of 1 nL. High-resolution 3D geometries were printed with features of ≤200 μm; these included an arborised cell junction, a diagonal-plane junction and an osteochondral interface. The printed cells showed high viability (90% on average) and HEK cells within the printed structures were shown to proliferate under culture conditions. Significantly, a five-week tissue engineering study demonstrated that printed oMSCs could be differentiated down the chondrogenic lineage to generate cartilage-like structures containing type II collagen.
Highlights
While bioprinting has advanced significantly over the last 15 years, the pursuit of morphological complexity and biological functionality in fabricated cellular constructs remains challenging[9]
Ovine mesenchymal stem cells in printed constructs responded to transforming growth factor-β3 (TGF-β3) and underwent differentiation to form cartilage-like structures
Human embryonic kidney (HEK)-293T cells or ovine mesenchymal stem cells (oMSCs) were harvested and dispersed in a sterile bioink, which was kept at 37 °C prior to printing
Summary
While bioprinting has advanced significantly over the last 15 years, the pursuit of morphological complexity and biological functionality in fabricated cellular constructs remains challenging[9]. Techniques employing two dispensing methods in tandem have afforded cellularised structures with non-cellular elements such as structural frameworks[20, 25] or perfusable microchannels[45] Even with these advances in bioprinting technologies, there are still only limited examples of 3D printed constructs that contain two or more cell types[12, 13, 21, 25] and of these, only a few cases where the patterned cell features were less than 250 μm wide[34, 46]. Ovine mesenchymal stem cells (oMSCs) in printed constructs responded to transforming growth factor-β3 (TGF-β3) and underwent differentiation to form cartilage-like structures These data demonstrate that fundamental biological processes can remain intact after printing, which suggests that the approach presented here will be useful for complex tissue fabrication
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