Independent of physical activity, volumetric muscle loss injury in a murine model impairs whole-body metabolism.

Volumetric muscle loss (VML) injuries result in a non-recoverable loss of muscle tissue and function due to trauma or surgery. Reductions in physical activity increase the risk of metabolic comorbidities over time, and it is likely that VML may reduce whole-body activity. However, these aspects remain uncharacterized following injury. Our goal was to characterize the impact of VML on whole-body physical activity and metabolism, and to further investigate possible muscle-specific metabolic changes. Adult male C57Bl/6J (n = 28) mice underwent a standardized VML injury to the posterior compartment of the hind limb, or served as injury naïve controls. Mice underwent longitudinal evaluation of whole-body physical activity and metabolism in specialized cages up to three times over the course of 8 weeks. At terminal time points of 4- and 8-weeks post-VML in vivo muscle function of the posterior compartment was evaluated. Additionally, the gastrocnemius muscle was collected to understand histological and biochemical changes in the muscle remaining after VML. The VML injury did not alter the physical activity of mice. However, there was a noted reduction in whole-body metabolism and diurnal fluctuations between lipid and carbohydrate oxidation were also reduced, largely driven by lower carbohydrate utilization during active hours. Following VML, muscle-specific changes indicate a decreased proportion of fast (i.e., type IIb and IIx) and a greater proportion of slow (i.e., type I and IIa) fibers. However, there were minimal changes in the capillarity and metabolic biochemical activity properties of the gastrocnemius muscle, suggesting a miss-match in capacity to support the physiologic needs of the fibers. These novel findings indicate that following VML, independent of changes in physical activity, there is whole-body diurnal metabolic inflexibility. Supporting future investigations into the chronic and overlooked co-morbidities of VML injury.

Pharmacological Mitigation of Fibrosis in a Porcine Model of Volumetric Muscle Loss Injury.

Volumetric muscle loss (VML) resulting from extremity trauma presents functional deficits and fibrosis, ultimately manifesting disability. The extensive fibrotic accumulation is expected to interfere with neural, trophic, vascular, and mechanical connectivity of any possible regenerative medicine approaches. Our objective was to quantify the muscle properties and stiffness following injury and investigate if the fibrotic deposition could be mitigated using an antifibrotic agent; we hypothesized that antifibrotic treatment would prevent the overwhelming fibrotic response. Yorkshire Cross pigs (n = 10) were randomized to sham or a nontreated ∼20% VML injury. Immediately following surgery, injured animals were further randomized to nintedanib (Ofev; 300 mg/day) or no treatment for 30 days. Longitudinal analysis of muscle function via peroneal nerve stimulation, compartment volume, and quantitative muscle stiffness using shearwave elastography were conducted. Terminally comprehensive histopathologic, biochemical, and genetic investigations were conducted on the skeletal muscle and fibrosis. Through 4 weeks post-VML, nontreated muscles presented a significant deficit (23%) in maximal torque compared to the sham operated (p < 0.01). The stiffness in the VML defect area increased significantly (7-fold) in the VML-nontreated leg than the VML antifibrotic-treated legs by 4 weeks postinjury, which was coupled with the nontreated muscle having ∼40% more hydroxyproline per mg of tissue than those receiving antifibrotic treatment (p = 0.01). This work indicates that VML injury progressively induces fibrosis and muscle stiffness. Antifibrotic treatment can mitigate the pathologic development of fibrosis. Future work should evaluate optimal timing and duration of treatments combined with regenerative medicine approaches in efforts to improve function. Impact statement This work primarily evaluated the effect of a clinically available antifibrotic therapy (nintedanib) on the development of fibrosis after volumetric muscle loss (VML) injury in a large animal model. As a primary outcome measure of fibrosis, skeletal muscle stiffness was repeatedly measured in vivo and noninvasively using a quantitative ultrasound device with shearwave elastography capability. The most salient finding of the study is that the antifibrotic nintedanib significantly reduced the development of VML injury-induced fibrous tissue deposition and stiffness.