Supplementation of GelMA with Minimally Processed Tissue Promotes the Formation of Densely Packed Skeletal-Muscle-Like Tissues.

We present a simple and cost-effective strategy for developing gelatin methacryloyl (GelMA) hydrogels supplemented with minimally processed tissue (MPT) to fabricate densely packed skeletal-muscle-like tissues. MPT powder was prepared from skeletal muscle by freeze-drying, grinding, and sieving. Cell-culture experiments showed that the incorporation of 0.5-2.0% (w/v) MPT into GelMA hydrogels enhances the proliferation of murine myoblasts (C2C12 cells) compared to proliferation in pristine GelMA hydrogels and GelMA supplemented with decellularized skeletal-muscle tissues (DCTs). MPT-supplemented constructs also preserved their three-dimensional (3D) integrity for 28 days. By contrast, analogous pristine GelMA constructs only maintained their structure for 14 days or less. C2C12 cells embedded in MPT-supplemented constructs exhibited a higher degree of cell alignment and reached a significantly higher density than cells loaded in pristine GelMA constructs. Our results suggest that the addition of MPT incorporates a rich source of biochemical and topological cues, such as growth factors, glycosaminoglycans (GAGs), and structurally preserved proteins (e.g., collagen). In addition, GelMA supplemented with MPT showed suitable rheological properties for use as bioinks for extrusion bioprinting. We envision that this simple and cost-effective strategy of hydrogel supplementation will evolve into an exciting spectrum of applications for tissue engineers, primarily in the biofabrication of relevant microtissues for in vitro models and cultured meat and ultimately for the biofabrication of transplant materials using autologous MPT.

Biofabrication of muscle fibers enhanced with plant viral nanoparticles using surface chaotic flows.

Multiple human tissues exhibit fibrous nature. Therefore, the fabrication of hydrogel filaments for tissue engineering is a trending topic. Current tissue models are made of materials that often require further enhancement for appropriate cell attachment, proliferation and differentiation. Here we present a simple strategy, based on the use of surface chaotic flows amenable to mathematical modeling, to fabricate continuous, long and thin filaments of gelatin methacryloyl (GelMA). The fabrication of these filaments is achieved by chaotic advection in a finely controlled and miniaturized version of the journal bearing system. A drop of GelMA pregel is injected on a higher-density viscous fluid (glycerin) and a chaotic flow is applied through an iterative process. The millimeter-scale hydrogel drop is exponentially deformed and elongated to generate a meter-scale fiber, which was then polymerized under UV-light exposure. Computational fluid dynamic (CFD) simulations are conducted to determine the characteristics of the flow and design the experimental conditions for fabrication of the fibers. GelMA fibers were effectively used as scaffolds for C2C12 myoblast cells. Experimental results demonstrate an accurate accordance with CFD simulations for the predicted length of the fibers. Plant-based viral nanoparticles (i.e.Turnip mosaic virus; TuMV) were then integrated to the hydrogel fibers as a secondary nano-scaffold for cells for enhanced muscle tissue engineering. The addition of TuMV significantly increased the metabolic activity of the cell-seeded fibers (p* < 0.05), strengthened cell attachment throughout the first 28 d, improved cell alignment, and promoted the generation of structures that resemble natural mammal muscle tissues. Chaotic two-dimensional-printing is proven to be a viable method for the fabrication of hydrogel fibers. The combined use of thin and long GelMA hydrogel fibers enhanced with flexuous virions offers a promising alternative for scaffolding of muscle cells and show potential to be used as cost-effective models for muscle tissue engineering purposes.