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High velocity impact injuries can often result in loss of large skeletal muscle mass, creating defects devoid of matrix, cells, and vasculature. Functional regeneration within these regions of large volumetric muscle loss (VML) continues to be a significant clinical challenge. Large cell-seeded, space-filling tissue-engineered constructs that may augment regeneration require adequate vascularization to maintain cell viability. However, the long-term effect of improved vascularization and the effect of addition of myoblasts to vascularized constructs have not been determined in large VMLs. Here, our objective was to create a new VML model, consisting of a full-thickness, single muscle defect, in the rat biceps femoris muscle, and evaluate the ability of myoblast-seeded vascularized collagen hydrogel constructs to augment VML regeneration. Adipose-derived microvessels were cultured with or without myoblasts to form vascular networks within collagen constructs. In the animal model, the VML injury was created in the left hind limb, and treated with the harvested autograft itself, constructs with microvessel fragments (MVF) only, constructs with microvessels and myoblasts (MVF+Myoblasts), or left empty. We evaluated the formation of vascular networks in vitro by light microscopy, and the capacity of vascularized constructs to augment early revascularization and muscle regeneration in the VML using perfusion angiography and creatine kinase activity, respectively. Myoblasts (Pax7+) were able to differentiate into myotubes (sarcomeric myosin MF20+) in vitro. The MVF+Myoblast group showed longer and more branched microvascular networks than the MVF group in vitro, but showed similar overall defect site vascular volumes at 2 weeks postimplantation by microcomputed tomography angiography. However, a larger number of small-diameter vessels were observed in the vascularized construct-treated groups. Yet, both vascularized implant groups showed primarily fibrotic tissue with adipose infiltration, poor maintenance of tissue volume within the VML, and little muscle regeneration. These data suggest that while vascularization may play an important supportive role, other factors besides adequate vascularity may determine the fate of regenerating volumetric muscle defects.

Skeletal Myoblast-Seeded Vascularized Tissue Scaffolds in the Treatment of a Large Volumetric Muscle Defect in the Rat Biceps Femoris Muscle

^{Skeletal Myoblast-Seeded Vascularized Tissue Scaffolds in the Treatment of a Large Volumetric Muscle Defect in the Rat Biceps Femoris Muscle}