Cai Aijia, Zheng Zengming, Müller-Seubert Wibke, Biggemann Jonas, Fey Tobias, Beier Justus P, Horch Raymund E, Frieß Benjamin, Arkudas Andreas
Department of Plastic and Hand Surgery and Laboratory for Tissue Engineering and Regenerative Medicine, University Hospital of Erlangen, Friedrich-Alexander University of Erlangen-Nürnberg (FAU), Krankenhausstraße 12, 91054 Erlangen, Germany.
Department of Materials Science and Engineering, Institute of Glass and Ceramics, Friedrich-Alexander-University Erlangen-Nürnberg (FAU), Martensstr. 5, 91058 Erlangen, Germany.
J Pers Med. 2022 Mar 11;12(3):442. doi: 10.3390/jpm12030442.
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.
去细胞化的全肌肉构建体是肌肉组织工程的理想支架，因为它们保留了骨骼肌组织细胞外基质的网络和蛋白质。血管蒂的存在使得基于灌注的去细胞化方案更有效，并允许随后在体内对肌肉构建体进行再细胞化和移植。本研究的目的是通过建立一个动物模型来为去细胞化全肌肉构建体的移植创建一个基线，该模型用于研究在血管化和功能方面从其蒂上分离出的完整天然肌肉。对5只雄性Lewis大鼠的左内侧腓肠肌进行制备，并将其从肌床上提起以进行原位肌肉刺激。刺激方案包括抽搐、强直刺激、疲劳测试和肌肉拉伸。测定了峰值力、最大收缩和舒张速率、达到最大收缩和舒张的时间以及最大收缩和舒张速率。之后，将肌肉取出并通过微血管吻合术异位移植到同基因大鼠的隔离室中。2周后，暴露移植的腓肠肌并再次进行刺激，随后用造影剂进行血管内灌注以进行μCT分析。然后将肌肉构建体石蜡包埋用于免疫组织学染色。肌肉移植后，峰值抽搐和强直力值均显著下降，而两组之间的疲劳指数和被动拉伸特性没有差异。血管分析显示保留了灌注血管，其中大多数血管半径较小，最大可达20μm和45μm。在本研究中，建立了一种在隔离室中进行异位微血管肌肉移植的新型大鼠模型。通过评估原位肌肉收缩特性以及移植2周后的血管分布，该模型为未来的研究奠定了基础，包括灌注去细胞化肌肉构建体的移植。
