Enhancing the Repair of Substantial Volumetric Muscle Loss by Creating Different Levels of Blood Vessel Networks Using Pre-Vascularized Nerve Hydrogel Implants.

Volumetric muscle loss (VML), a severe muscle tissue loss from trauma or surgery, results in scarring, limited regeneration, and significant fibrosis, leading to lasting reductions in muscle mass and function. A promising approach for VML recovery involves restoring vascular and neural networks at the injury site, a process not extensively studied yet. Collagen hydrogels have been investigated as scaffolds for blood vessel formation due to their biocompatibility, but reconstructing blood vessels and guiding innervation at the injury site is still difficult. In this study, collagen hydrogels with varied densities of vessel-forming cells are implanted subcutaneously in mice, generating pre-vascularized hydrogels with diverse vessel densities (0-145 numbers/mm2) within a week. These hydrogels, after being transplanted into muscle injury sites, are assessed for muscle repair capabilities. Results showed that hydrogels with high microvessel densities, filling the wound area, effectively reconnected with host vasculature and neural networks, promoting neovascularization and muscle integration, and addressing about 63% of the VML.

Engineering large and geometrically controlled vascularized nerve tissue in collagen hydrogels to restore large-sized volumetric muscle loss.

Developing scalable vascularized and innervated tissue is a critical challenge for the successful clinical application of tissue-engineered constructs. Collagen hydrogels are extensively utilized in cell-mediated vascular network formation because of their naturally excellent biological properties. However, the substantial increase in hydrogel contraction induced by populated cells limits their long-term use. Previous studies attempted to mitigate this issue by concentrating collagen pre-polymer solutions or synthesizing covalently crosslinked collagen hydrogels. However, these methods only partially reduce hydrogel contraction while hindering blood vessel formation within the hydrogels. To address this challenge, we introduced additional support in the form of a supportive spacer to counteract the contraction forces of populated cells and prevent hydrogel contraction. This approach was found to promote cell spreading, resist hydrogel contraction, control hydrogel/tissue geometry, and even facilitate the engineering of functional blood vessels and host nerve growth in just one week. Subsequently, implanting these engineered tissues into muscle defect sites resulted in timely anastomosis with the host vasculature, leading to enhanced myogenesis, increased muscle innervation, and the restoration of injured muscle functionality. Overall, this innovative strategy expands the applicability of collagen hydrogels in fabricating large vascularized nerve tissue constructs for repairing volumetric muscle loss (∼63 %) and restoring muscle function.

Viscous Fingering as a Rapid 3D Pattering Technique for Engineering Cell-Laden Vascular-Like Constructs.

Tissues are much larger than the diffusion limit distance, so rapidly providing blood vessels to supply embedded cells inside tissues with sufficient nutrients and oxygen is regarded as a major strategy for the success of bioengineered large and thick tissue constructs. Here, a patterning technique, viscous fingering, is developed to bioengineer vascularized-like tissues within a few minutes. By controlling viscosity, flow rate, and the volume of photo-cross-linkable prepolymer, macro- and microscale vascular network structures can be precisely engineered using the Hele-Shaw cell that is designed in this study. After cross-linking, a vascular-like gel with fingering structures is formed between the bottom and top base gels, creating a sandwich-like structure. Cells can be incorporated into the fingers, bases, or both gels. The spreading and growth direction of the embedded cells are successfully controlled and guided by manipulating the physical properties of the fingering and base gels individually. Moreover, fingering is generated, connected, and surrounded prepared cell-laden microgels in base prepolymers to form prevascularized tissue-like constructs. Taken together, the 3D cell patterning technique extends the potential for modeling and fabricating large and stackable vascularized tissue-like constructs for both ex vivo and in vivo applications.