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3D bioprinting is the additive manufacture of biological materials, such as cells, and has been implemented over the last two decades to print 3D constructs which can mature into functional tissues. Although there are many bioprinting techniques, there are few that are capable of simultaneously printing two or more cell types at high resolutions, high cell densities and high initial cell viabilities. These printing characteristics are required to pattern cells in tissue-like micro-architectures and to quickly mature physiologically relevant tissue. We developed a novel 3D bioprinter and bioprinting methodology, designed to pattern cells without compromising these printing characteristics. Initially a droplet-in-oil 3D printer was created, which assembled mm-scale aqueous droplet networks composed of thousands of picolitre volume micro-compartments. These networks were programmed to show emergent tissue-like properties, mimicking nervous tissue's rapid electrical communication or self-folding in a manner similar to muscle contractions. This printing process was then adapted to pattern HEK-293T cells within droplet networks at high droplet resolution (~50-150 μm), high initial cell viability (90±4%) and high droplet cell density (~30x106 cells/mL). Capillary-like and layered sheet cell architectures were formed with two printed fluorescent cell populations. These 3D printed constructs were transferred to culture medium with minimal loss in pattern fidelity using a novel phase transfer approach. Patterned HEK-293T cells developed into dense cell organisations which showed, high cell viabilities, high cell numbers (up to ~18,000) and mitotically active cells, when cultured over 7 days. Specialised tissues were also developed, here, patterned ovine mesenchymal stem cells were differentiated, after printing, into chondrogenic progenitor cells as affirmed by Sox9 expression. |
