Tissue Engineered Motor Units for Neuromuscular Modeling and Repair

PROJECT SUMMARY Innervation plays a key role in muscle development, function, and regeneration. Therefore, tissue engineering strategies to develop biomimetic skeletal muscle to study developmental mechanisms as well as for regenerative medicine applications should consider the implications of neural inputs during the biofabrication process.

Moreover, tissue regeneration following severe musculoskeletal injuries like volumetric muscle loss (VML) is hindered by a lack of appropriate motor innervations, which diminishes regenerative capacity and often results in minimal functional recovery. Hence, there is an unmet clinical need for an intervention strategy that augments

reinnervation and promotes a pro-regenerative environment following severe musculoskeletal trauma. Over the last several years, our group has shown that pre-innervated tissue engineered muscle scaffolds can effectively enhance muscle regeneration, revascularization, and functional recovery following VML. Building on these

findings, the goal of the current program is to advance the first three-dimensional tissue engineered motor units (3D-TEMUs) comprising dense bundles of centimeter-scale myofiber fascicles innervated by preformed axonal networks projecting from discrete pools of motor and/or sensory neurons. The 3D-TEMUs will be validated as

an in vitro testbed to study the role of innervation in facilitating muscle development as well as a composite soft tissue to augment functional regeneration following implantation in a rodent model of VML. Notably, 3D-TEMUs will be generated using human induced pluripotent stem cell (iPSC) derived myocytes and motor/sensory

neurons to perform in-depth characterization using a clinically-relevant and potentially translatable biomass. These efforts will be carried out across 3 Specific Aims. First, human 3D-TEMUs will be optimized to recapitulate the native myofascicular architecture of skeletal muscle tissue in order to provide a biofidelic testbed to better

understand neuromuscular development and sensorimotor function in the context of myofiber formation and maturation as well as to refine tissue engineering strategies for augmenting muscle regeneration (Aim 1). Next, 3D-TEMU neurons will be transduced to contain optogenetically controlled (i.e., light-activated) motor inputs that

allow for spatiotemporal control of muscle-fiber contraction to demonstrate refinement of motor units over time and to measure sensorimotor function (Aim 2). Then, 3D-TEMUs will be implanted in a rat model of VML to optimize acute graft cell survival (Aim 3A) followed by chronic studies assessing the ability of 3D-TEMUs in

facilitating bulk muscle replacement, reinnervation, revascularization, and overall functional restoration (Aim 3B). Successful execution of these studies will significantly advance the development of 3D-TEMUs as both an in vitro testbed to study mechanisms of neuromuscular development as well as an implantable composite soft

tissue to enable functional regeneration following currently intractable VML. With further advancement and clinical translation, 3D-TEMUs could potentially change the surgical paradigm for muscle repair and fundamentally alter clinical expectations for functional recovery following major musculoskeletal trauma.