Microsurgical Transplantation of Pedicled Muscles in an Isolation Chamber—A Novel Approach to Engineering Muscle Constructs via Perfusion-Decellularization

Abstract

Decellularized whole muscle constructs represent an ideal scaffold for muscle tissue engineering means as they retain the network and proteins of the extracellular matrix of skeletal muscle tissue. The presence of a vascular pedicle enables a more efficient perfusion-based decellularization protocol and allows for subsequent recellularization and transplantation of the muscle construct in vivo. The goal of this study was to create a baseline for transplantation of decellularized whole muscle constructs by establishing an animal model for investigating a complete native muscle isolated on its pedicle in terms of vascularization and functionality. The left medial gastrocnemius muscles of 5 male Lewis rats were prepared and raised from their beds for in situ muscle stimulation. The stimulation protocol included twitches, tetanic stimulation, fatigue testing, and stretching of the muscles. Peak force, maximum rate of contraction and relaxation, time to maximum contraction and relaxation, and maximum contraction and relaxation rate were determined. Afterwards, muscles were explanted and transplanted heterotopically in syngeneic rats in an isolation chamber by microvascular anastomosis. After 2 weeks, transplanted gastrocnemius muscles were exposed and stimulated again followed by intravascular perfusion with a contrast agent for µCT analysis. Muscle constructs were then paraffin embedded for immunohistological staining. Peak twitch and tetanic force values all decreased significantly after muscle transplantation while fatigue index and passive stretch properties did not differ between the two groups. Vascular analysis revealed retained perfused vessels most of which were in a smaller radius range of up to 20 µm and 45 µm. In this study, a novel rat model of heterotopic microvascular muscle transplantation in an isolation chamber was established. With the assessment of in situ muscle contraction properties as well as vessel distribution after 2 weeks of transplantation, this model serves as a base for future studies including the transplantation of perfusion-decellularized muscle constructs.