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)： 严重的创伤会导致体积性肌肉丧失(VML)，导致终身残疾。目前的治疗方法极其有限，对有意义的功能恢复几乎没有希望。我们通过干细胞移植技术与自愿车轮跑步形式的体力活动相结合，以促进移植细胞的植入和成熟，从而追求再生康复治疗VML的方法。在VML的小鼠模型中，我们一直在开发将肌肉干细胞(MuSCs)与FACS从肌肉中分离的其他单核细胞一起移植到VML病变中的方法。根据组织学和生理学评价，我们能够获得肌肉结构和功能的显著恢复。然而，即使在体外通过直接电刺激肌肉来评估力量有显著的恢复，通过刺激体内坐骨神经进行的力量评估也显示出最小的恢复，这表明新形成的肌肉的神经支配是有限的。同样，体力活动对再生反应的增强，如血管的增加和纤维化的减少，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多的轴突再生和神经肌肉接头(NMJ)的形成和成熟。基于不同形式的运动对轴突再生影响的证据，本研究的重点是考察不同的运动方案，以促进轴突再生和NMJ的形成，从而优化MUSC治疗VML的方法。这些研究分为两个具体目标。在目标1中，我们将初步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)与自愿车轮跑I简单的再神经-VML损伤模型(即不移植)和肌肉/神经联合损伤模型(注射肌毒素心脏毒素和神经毒素巴曲霉毒素)。前者提供了VML+移植后早期反应的证据，后者提供了在广泛肌肉环境中重新支配神经的模型。 再生。我们将评估NMJ的组织学形成和坐骨神经刺激产生的力量。在目标2中，我们将应用任何一种运动范式来研究运动对骨髓间充质干细胞移植治疗VML的影响。我们将在VML手术后使用我们优化的移植方案，然后应用优化的锻炼方案三周。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情况。长期目标是优化人类VML治疗的再生康复策略，包括以干细胞疗法为形式的再生医学方法与以体力活动形式的康复医学方法相结合，以优化组织恢复和功能恢复。因此，所有方法都被设计为可扩展到人类，并尽可能接近地模拟人类使用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.