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Microgravity has been shown to lead to both muscle atrophy and impaired muscle regeneration. The purpose was to study the efficacy of microgravity to model impaired muscle regeneration in an engineered muscle platform and then to demonstrate the feasibility of performing drug screening in this model. Engineered human muscle was launched to the International Space Station National Laboratory, where the effect of microgravity exposure for 7 days was examined by transcriptomics and proteomics approaches. Gene set enrichment analysis of engineered muscle cultured in microgravity, compared to normal gravity conditions, highlighted a metabolic shift toward lipid and fatty acid metabolism, along with increased apoptotic gene expression. The addition of pro-regenerative drugs, insulin-like growth factor-1 (IGF-1) and a 15-hydroxyprostaglandin dehydrogenase inhibitor (15-PGDH-i), partially inhibited the effects of microgravity. In summary, microgravity mimics aspects of impaired myogenesis, and the addition of these drugs could partially inhibit the effects induced by microgravity. 

Skeletal myofibers naturally regenerate after damage; however, impaired muscle function can result in cases when a prominent portion of skeletal muscle mass is lost, for example, following traumatic muscle injury. Volumetric muscle loss can be modeled in mice using a surgical model of muscle ablation to study the pathology of volumetric muscle loss and to test experimental treatments, such as the implantation of acellular scaffolds, which promote de novo myogenesis and angiogenesis. Here we provide step-by-step instructions to perform full-thickness surgical ablation, using biopsy punches, and to remove a large volume of the tibialis anterior muscle of the lower limb in mice. This procedure results in a reduction in muscle mass and limited regeneration capacity; the approach is easy to reproduce and can also be applied to larger animal models. For therapeutic applications, we further explain how to implant bioscaffolds into the ablated muscle site. With adequate training and practice, the surgical procedure can be performed within 30 min. 

Volumetric muscle loss (VML) is associated with irreversibly impaired muscle function due to traumatic injury. Experimental approaches to treat VML include the delivery of basic fibroblast growth factor (bFGF) or rehabilitative exercise. The objective of this study was to compare the effects of spatially nanopatterned collagen scaffold implants with either bFGF delivery or in conjunction with voluntary exercise. Aligned nanofibrillar collagen scaffold bundles were adsorbed with bFGF, and the bioactivity of bFGF-laden scaffolds was examined by skeletal myoblast or endothelial cell proliferation. The therapeutic efficacy of scaffold implants with either bFGF release or exercise was examined in a murine VML model. Our results show an initial burst release of bFGF from the scaffolds, followed by a slower release over 21 days. The released bFGF induced myoblast and endothelial cell proliferation in vitro. After 3 weeks of implantation in a mouse VML model, twitch force generation was significantly higher in mice treated with bFGF-laden scaffolds compared to bFGF-laden scaffolds with exercise. However, myofiber density was not significantly improved with bFGF scaffolds or voluntary exercise. In contrast, the scaffold implant with exercise induced more re-innervation than all other groups. These results highlight the differential effects of bFGF and exercise on muscle regeneration. 

The regeneration of skeletal muscle can be permanently impaired by traumatic injuries, despite the high regenerative capacity of native muscle. An attractive therapeutic approach for treating severe muscle inuries is the implantation of off-the-shelf engineered biomimetic scaffolds into the site of tissue damage to enhance muscle regeneration. Anisotropic nanofibrillar scaffolds provide spatial patterning cues to create organized myofibers, and growth factors such as insulin-like growth factor-1 (IGF-1) are potent inducers of both muscle regeneration as well as angiogenesis. The aim of this study was to test the therapeutic efficacy of anisotropic IGF-1-releasing collagen scaffolds combined with voluntary exercise for the treatment of acute volumetric muscle loss, with a focus on histomorphological effects. To enhance the angiogenic and regenerative potential of injured murine skeletal muscle, IGF-1-laden nanofibrillar scaffolds with aligned topography were fabricated using a shear-mediated extrusion approach, followed by growth factor adsorption. Individual scaffolds released a cumulative total of 1244 ng ± 153 ng of IGF-1 over the course of 21 days in vitro . To test the bioactivity of IGF-1-releasing scaffolds, the myotube formation capacity of murine myoblasts was quantified. On IGF-1-releasing scaffolds seeded with myoblasts, the resulting myotubes formed were 1.5-fold longer in length and contained 2-fold greater nuclei per myotube, when compared to scaffolds without IGF-1. When implanted into the ablated murine tibialis anterior muscle, the IGF-1-laden scaffolds, in conjunction with voluntary wheel running, significantly increased the density of perfused microvessels by greater than 3-fold, in comparison to treatment with scaffolds without IGF-1. Enhanced myogenesis was also observed in animals treated with the IGF-1-laden scaffolds combined with exercise, compared to control scaffolds transplanted into mice that did not receive exercise. Furthermore, the abundance of mature neuromuscular junctions was greater by approximately 2-fold in muscles treated with IGF-1-laden scaffolds, when paired with exercise, in comparison to the same treatment without exercise. These findings demonstrate that voluntary exercise improves the regenerative effect of growth factor-laden scaffolds by augmenting neurovascular regeneration, and have important translational implications in the design of off-the-shelf therapeutics for the treatment of traumatic muscle injury.