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Scaffold-free, label-free and nozzle-free biofabrication technology using magnetic levitational assembly

Tissue spheroids have been proposed as building blocks in 3D biofabrication. Conventional magnetic force-driven 2D patterning of tissue spheroids requires prior cell labeling by magnetic nanoparticles, meanwhile a label-free approach for 3D magnetic levitational assembly has been introduced. Here we present first-time report on rapid assembly of 3D tissue construct using scaffold-free, nozzle-free and label-free magnetic levitation of tissue spheroids. 
Chondrospheres of standard size, shape and capable to fusion have been biofabricated from primary sheep chondrocytes using non-adhesive technology. Label-free magnetic levitation was performed using a prototype device equipped with permanent magnets in presence of gadolinium (Gd3+) in culture media, which enables magnetic levitation. 

Mathematical modeling and computer simulations were used for prediction of magnetic field and kinetics of tissue spheroids assembly into 3D tissue constructs. First, we used polystyrene beads to simulate the assembly of tissue spheroids and to determine the optimal settings for magnetic levitation in presence of Gd3+. Second, we proved the ability of chondrospheres to assemble rapidly into 3D tissue construct in the permanent magnetic field in the presence of Gd3+.

Thus, scaffold- and label-free magnetic levitation of tissue spheroids is a promising approach for rapid 3D biofabrication and attractive alternative to label-based magnetic force-driven tissue engineering.

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Bioprinting of a functional vascularized mouse thyroid gland construct

Bioprinting can be defined as additive biofabrication of three-dimensional (3D) tissues and organ constructs using tissue spheroids, capable of self-assembly, as building blocks. The thyroid gland, a relatively simple endocrine organ, is suitable for testing the proposed bioprinting technology. Here we report the bioprinting of a functional vascularized mouse thyroid gland construct from embryonic tissue spheroids as a proof of concept. Based on the self-assembly principle, we generated thyroid tissue starting from thyroid spheroids (TS) and allantoic spheroids (AS) as a source of thyrocytes and endothelial cells (EC), respectively. Inspired by mathematical modeling of spheroid fusion, we used an original 3D bioprinter to print TS in close association with AS within a collagen hydrogel. During the culture, closely placed embryonic tissue spheroids fused into a single integral construct, EC from AS invaded and vascularized TS, and epithelial cells from the TS progressively formed follicles. In this experimental setting, we observed formation of a capillary network around follicular cells, as observed during in utero thyroid development when thyroid epithelium controls the recruitment, invasion and expansion of EC around follicles. To prove that EC from AS are responsible for vascularization of the thyroid gland construct, we depleted endogenous EC from TS before bioprinting. EC from AS completely revascularized depleted thyroid tissue. The cultured bioprinted construct was functional as it could normalize blood thyroxine levels and body temperature after grafting under the kidney capsule of hypothyroid mice. Bioprinting of functional vascularized mouse thyroid gland construct represents a further advance in bioprinting technology, exploring the self-assembling properties of tissue spheroids.

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The Scalable Standardized Biofabrication of Tissue Spheroids from Different Cell Types Using Nonadhesive Technology

We suggest a straightforward procedure for biofabrication and initial characterization of tissue spheroids with optimal controllable parameters prepared from four different cell types using nonadhesive technology. Applying different immortalized and primary cells, namely HEK293, primary human fibroblasts (HF), primary sheep chondrocytes, and primary sheep osteoblasts, we have demonstrated the reproducibility and scalability of spheroid generation, the strong dependency of ultimate spheroid diameter on initial cell seeding density, and cell type. In addition, the spheroid viability is governed by cell derivation. In this study, we suggest a decision procedure to apply for any cell type one starts to work with to prepare a new type of tissue spheroids with predictable controllable optimal features suitable for high-quality standards in biofabrication and drug discovery. 

Публикации команды

Patterning of tissue spheroids biofabricated from human fibroblasts on the surface of electrospun polyurethane matrix using 3D bioprinter

Organ printing is a computer-aided additive biofabrication of functional three-dimensional human tissue and organ constructs according to digital model using the tissue spheroids as building blocks.

Публикации команды

Распластывание тканевых сфероидов, сформированных из первичных фибробластов человека, на поверхности микроволокнистого электроспиннингового полиуретанового матрикса

Тканевые сфероиды, сформированные из первичных фибробластов человека с использованием неадгезивных агарозных
форм, были размещены с помощью трехмерного биопринтера на поверхности микроволокнистого полиуретанового матрик-
са, полученного методом электроспиннинга.