High-throughput Spheroid Bioprinting Technology for Scalable Fabrication of Tissues

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ROJECT SUMMARY/ABSTRACT The ability to bioprint cellular aggregates, such as spheroids, in a high-throughput manner into desired patterns or cellular microenvironments is crucial to facilitate fabrication of scalable constructs with cell densities similar to that of native tissues and organs. Despite the progress in spheroid bioprinting technologies, the major shortcomings associated with them, such as poor positioning of spheroids, significant loss of viability and structural integrity, poor repeatability of the process when using non-uniform size spheroids, inability to form complex 3D shapes, and most importantly, the lack of scalability, limit their translation. In this project, we propose a highly unique technology, henceforth referred as “high-throughput spheroid (HTS) bioprinting,” that enables simultaneous bioprinting of several spheroids with an order of magnitude size range and minimal cellular damage, at a high positional precision and an unprecedented speed. The proposed technology is highly versatile and enables the bioprinting of complex structures either (1) onto the surface of gel substrates (i.e., hydrogels) in a scaffold-based manner or (2) within support baths (i.e., sacrificial microgels) in a scaffold-free manner for scalable fabrication of tissues. In Specific Aim 1, we propose to develop HTS bioprinting, which has the capability of depositing several spheroids simultaneously on 3D gel substrates, thus bioprinting a complete layer of the 3D tissue at once in a rapid fashion (i.e., 100 spheroids can be bioprinted in <20 sec). We will couple HTS bioprinting with extrusion-based bioprinting of gel substrates and explore the spheroid-gel interactions, across a wide range of hydrogels, during the bioprinting process. To exemplify the technology, we will demonstrate bioprinting intraoperatively via depositing osteogenically-committed bone spheroids for the repair of craniomaxillofacial bone defects in a rat model. In Specific Aim 2, we will reconfigure the HTS bioprinting technology for freeform positioning of spheroids within sacrificial support baths. Here, we will bioprint spheroids sequentially (one after the other) in a rapid manner and pattern them according to the target design. We will explore the gel-spheroid- bioprinting process interactions, where the effectiveness of the technology will be tested for multiple support baths, including alginate microgels to be fabricated using the air-jet assisted coaxial flow technique along with a commercially available benchmark. We will exemplify the utilization of the technology for fabrication of anatomically-relevant complex-shaped human bronchopulmonary segments. In this regard, we have formed a complementary collaboration that merges essential domain knowledge in bioprinting, bioprinting process and instrument development, biomaterials, craniofacial surgery, and bone and lung tissue engineering with the depth necessary to propel the proposed work towards meaningful advances that would otherwise not be possible. Successful completion of the proposed work is anticipated to give rise to an advanced bioprinting technology for HTS bioprinting and thereby provide a novel tool for fabrication of scalable tissues and organs.