Utilization of Engineered Skeletal Muscle Units to Repair Volumetric Muscle

 描述（由申请方提供）：多种病理状况（包括肌肉创伤、手术损伤和肌病）可导致肌肉体积损失（VML），导致肌肉功能不可恢复。迄今为止，VML的治疗包括肌瓣或移植物并不总是有效的，并且受到有限的组织可用性和供体部位发病率的阻碍。具有功能性肌腱连接、腱-骨锚和神经支配的工程化肌肉组织的产生不仅将恢复复杂组织的功能，例如创伤性损伤或疾病后的小面部肌肉，而且还可以用作研究发育肌肉生物学和肌肉药理学的模型。为了满足骨骼肌替代组织的需求，我们的实验室开发了一种具有适当尺寸和功能的组织工程化功能性骨骼肌单位（SMU），用于小型VML损伤（如手部和面部）的临床应用[1-8]。我们的SMU是一种多相复合组织，由具有工程化肌腱和/或骨-肌腱锚端的工程化肌肉组织组成。在体内暴露于VML环境28天后，我们的构建体表现出SMU表型的显著改变，包括包裹在广泛的细胞外基质中的单轴排列的肌纤维、骨附着处的完全骨愈合、附着点和肌腱界面的形成、广泛的血管化和神经支配以及伴随的神经肌肉接头的形成（8）。这项令人兴奋的技术实际上会在修复区域添加新的肌肉纤维。我们的研究结果表明，我们的多相复合组织在植入后存活，在体内经受住了施加在它们身上的机械载荷，并形成了必要的界面，这些界面是将表型向成人肌肉结构和功能推进所需的。虽然我们的技术在修复受损肌肉方面表现出很大的希望，但仍有大量工作要做，以确定如何最好地利用这项技术来获得肌肉功能到受伤部位的最佳恢复。例如，我们需要更完全地恢复天然肌肉力量。关于在人类中使用这种新颖和创新技术的另一个问题是在患者中使用干细胞/前体细胞衍生组织的安全性。用于产生这些组织构建体的原代细胞或基质细胞在植入后不构成急性或长期威胁是必要的。该领域的主要关注点是确定移植构建体中没有未分化的细胞持续存在的能力，所述未分化的细胞持续存在以后可能导致异常细胞行为如癌症。因此，为了使该技术向临床应用发展，必须阐明移植组织整合、再生和修复的机制。该提案的总体目标是使用VML作为模型，用于机械地观察SMU组织在植入到适当的诱导组织环境中后的整合和再生。这项工作对干细胞生物学，肌肉骨骼生物学和再生医学以及肌肉骨骼组织损伤治疗的直接临床相关性具有广泛的意义。

 DESCRIPTION (provided by applicant): Multiple pathological conditions including muscle trauma, surgical damage, and myopathies can lead to volumetric muscle loss (VML) resulting in unrecoverable muscle function. To date, the treatments for VML including the muscle flap or graft are not always effective and are hindered by limited tissue availability and donor site morbidity. The creation of engineered muscle tissue with functional myotendinous junctions, tendon- bone anchors and innervation will not only restore the function of a complex tissue, e.g. a small facial muscle following traumatic injury or disease, but can also be used as a model for studying developmental muscle biology and muscle pharmacology. To address the need for skeletal muscle replacement tissue, our laboratory has developed a tissue-engineered functional skeletal muscle unit (SMU) of appropriate size and function for clinical use in situations of small VML injuries, such as those found in the hand and face [1-8]. Our SMU is a multi-phasic composite tissue consisting of engineered muscle tissue with engineered tendon and/or bone- tendon anchor ends. After twenty-eight days of exposure in vivo to a VML environment, our constructs exhibited significant alteration to the SMU phenotype including uniaxial aligned muscle fibers encased in an extensive extra-cellular matrix, complete bone healing at the bone attachment, formation of enthesis and myotendinous interfaces , extensive vascularization and innervation with concomitant formation of neuromuscular junctions (8). This exciting technology actually results in the addition of new muscle fibers to the repair area. Our findings show that our multi-phasic composite tissue survives implantation, survives the mechanical loads placed on them in vivo, and develops the necessary interfaces needed to advance the phenotype toward adult muscle structure and function. While our technology shows great promise for the repair of damaged muscle, a lot of work remains to determine how best to utilize this technology to obtain optimal recovery of muscle function to an injury site. For example, we need more complete recovery of native muscle forces. An additional concern regarding the use of this novel and innovative technology in humans is the safety of use of stem/precursor cell-derived tissue in patients. It is imperative that the primary or stromal cells used for the generation of these tissue constructs pose no acute or long-term threat after implantation. The main concern in the field is the ability to ascertain that no undifferentiated cels persist in the transplanted construct that might later lead to aberrant cellular behavior such as cancer. Thus, to progress this technology towards clinical application, the mechanisms of graft tissue integration, regeneration, and repair must be elucidated. The overall goal of this proposal is to use VML as a model for looking mechanistically at the integration and regeneration of our SMU tissue following implantation into an appropriate inductive tissue environment. This work has broad implications for stem cell biology, musculoskeletal biology and regenerative medicine as well as direct clinical relevance for the treatment of musculoskeletal tissue injuries.