The Effect of Exercise on the Efficacy of Neural Innervation in the Treatment of Volumetric Muscle Loss

 描述（由申请人提供）： 严重创伤可导致肌肉体积损失（VML），导致终身残疾。目前的治疗方法非常有限，几乎没有希望实现有意义的功能恢复。我们采用再生康复方法治疗VML，方法是将干细胞移植技术与身体活动结合起来，以自愿跑轮子的形式促进移植细胞的植入和成熟。在VML的鼠模型中，我们一直在开发用于将肌肉干细胞（MuSC）沿着与通过FACS从肌肉分离的其他单核细胞一起移植到VML病变中的方法。基于组织学和生理学评估，我们能够获得肌肉结构和功能的显著恢复。然而，即使存在如通过离体肌肉的直接电刺激评估的力的显著恢复，通过刺激坐骨神经的体内力评估也揭示了最小的恢复，这表明新形成的肌肉的神经支配是有限的。同样，通过身体活动增强再生反应，例如增强血管分布和减少纤维化，并不伴随着肌纤维横截面积的显著增加，这表明这种形式的运动确实产生了显著更大量的轴突再生和神经肌肉接头（NMJ）形成和成熟。 基于不同形式的运动对轴突再生的影响的证据，本提案的研究重点是检查不同的运动模式，以通过增强轴突再生和NMJ形成来优化VML的MuSC治疗。研究分为两个具体目标。在目标1中，我们将首先比较两种类型的跑步机训练（低强度训练和高强度间歇训练）与自主轮跑，即简单的神经再支配模型-单独的VML损伤（即无移植）和组合的肌肉/神经损伤模型（注射肌毒素心脏毒素和神经毒素蛙毒素）。前者提供了VML加移植后早期反应的证据，后者提供了广泛肌肉环境中神经再支配的模型。 再生我们将从组织学上评估NMJ形成和坐骨神经刺激产生的力。在目标2中，我们将采用导致最佳结果的运动模式来研究运动对MuSC移植治疗VML的影响。我们将在VML手术后使用我们优化的移植方案，然后应用优化的运动方案三周。届时，我们将通过组织学和生理学分析再次评估肌肉结构和功能的恢复。 长期目标是优化用于治疗人类VML的再生康复策略，该策略涉及以干细胞疗法形式的再生医学方法与以身体活动形式的康复医学方法相结合，以优化组织再生和功能恢复。因此，所有这些方法都被设计为可扩展到人类，并尽可能接近人类使用VML所面临的挑战。

 DESCRIPTION (provided by applicant): Major trauma can cause volumetric muscle loss (VML) resulting in life-long disability. Current therapies are extremely limited and offer little hope of meaningful functional recovery. We have pursued a Regenerative Rehabilitation approach to VML therapy by combining stem cell transplantation technology with physical activity in the form of voluntary wheel running to promote the engraftment and maturation of the transplanted cells. In a murine model of VML, we have been developing methodologies for the transplantation of muscle stem cells (MuSCs) along with other mononucleated cells isolated from muscle by FACS into VML lesions. Based on both histological and physiological assessment, we are able to obtain significant recovery of muscle structure and function. However, even though there is significant recovery of force as assessed by direct electrical stimulation of the muscle ex vivo, force assessment by stimulation of the sciatic nerve in vivo has revealed minimal recovery, suggesting that innervation of the newly formed muscle is limited. Likewise, the enhancement of regenerative responses by physical activity, such as enhanced vascularity and reduced fibrosis, are not accompanied by significant increases in muscle fiber cross-sectional area, suggesting that this form of exercise does yield a significantly greater amount of axon regrowth and neuromuscular junction (NMJ) formation and maturation. Based on evidence of effects of different forms of exercise on axon regrowth, the focus of the studies of this proposal is to examine different exercise paradigms to optimize MuSC therapy for VML by enhancing axon regrowth and NMJ formation. The studies are divided into two Specific Aims. In Aim 1, we will initially compare two types of treadmill training (low intensity training and high intensity interval training) to voluntary wheel running i simple models of re- innervation - VML injury alone (i.e. no transplantation) and a combined muscle/nerve injury model (injection of the myotoxin cardiotoxin and the neurotoxin batrachotoxin). The former provides evidence as to early responses seen after VML plus transplantation, and the latter provides a model of re-innervation in a setting of extensive muscle regeneration. We will assess NMJ formation histologically and force generation from sciatic nerve stimulation. In Aim 2, we will apply whichever exercise paradigm leads to the best outcome to the study of effects of exercise on MuSC transplantation for VML. We will use our optimized transplantation protocol following VML surgery and then apply the optimized exercise protocol for three weeks. At that time, we will again assess the restoration of muscle structure and function by histological and physiological analyses. The long-term goal is to optimize Regenerative Rehabilitation strategies for the treatment of VML in humans that involve a combination of regenerative medicine approaches, in the form of stem cell therapeutics, with rehabilitation medicine approaches, in the form of physical activity, to optimize tissue restoratio and functional recovery. As such, all of the approaches are designed to be scalable to humans and to model as closely as possible the challenges faced by humans with VML.