Large extremity injuries that involve damage to both muscle and bone pose a significant challenge in the clinic. Even with gold standard treatment, covering the exposed bony defect with a vascularized muscle flap, these injuries often do not recover to full function, affecting patients' quality of life and their contributions to society. Patients recovering from large extremity wounds suffer from pain and mobility issues years after limb salvage, and some patients ultimately opt for late stage amputation. Thus, there is a need for tissue engineering strategies to facilitate functional recovery of large composite defects of the limb.

Despite this pressing need, few tissue engineering strategies exist and have been tested in composite defects preclinically, limiting the number of translatable treatment options. The overall goal of this dissertation is to examine the regenerative potential of engineered matrix constructs and stem cells on composite bone & muscle defects. To accomplish this goal, we first established a volumetric muscle loss model in the quadriceps and a composite muscle and bone injury model in the quadriceps and femur of the rat, designed to allow for functional measurements of tissue function. We found that a large VML in the quadriceps caused sufficient damage that an autograft within the defect space was not able to significantly recover muscle function. In the composite injury model, the addition of the muscle injury on top of the segmental bone defect compounded functional deficits and attenuated rhBMP-2 mediated bone regeneration. In the characterization of these models, we found that the VML in the quadriceps defect was complicated by multi-muscle and vessel/nerve damage and that early revascularization was delayed in the composite injury group compared to the bone injury only group. To address these issues, we designed a separate VML model that is more suited for testing for muscle regeneration within a single, planar muscle, the biceps femoris, and further tested a vascular treatment, microvascular constructs with or without myoblasts, within this VML model. We found that though this treatment allowed for an early vascular response, the microvascular constructs treatment did not lead to a full recovery of the muscle. Third, we tested the effects of this vascular treatment in the composite injury model, either surrounding the bone or within the muscle defect. We found that the microvascular constructs surrounding the bone resulted in an early revascularization of the limb but led to decreased bone regeneration. Microvascular constructs with myoblasts, on the other hand, were implanted into the muscle and demonstrated a small increase in both bone and muscle regeneration despite a lack of increased vascularization. Taken together, these results suggest that while vascularization may play a role in regenerating tissues, the full recovery of complex limb injuries likely depend on other cellular and biochemical factors as well.

The studies presented here move the field of composite injuries forward by presenting new platforms on which to test therapeutics and measure functional outcomes. This work has led to some insights into the revascularization of large defects, and by analyzing the effects of vascular treatment on these complex injuries, we have demonstrated a potential therapeutic that may warrant further research. Overall, this work will aid in the development of tissue engineering treatments to facilitate the full functional recovery of complex limb trauma.