The role of myosin II in tendon repair under glucose control

肌腱损伤和紊乱严重影响了经济成本和满足 患者的职业、娱乐和健康活动。尽管不断努力改进治疗方法 肌腱修复的方式，真正有效和有效的治疗方法尚未建立。我们的长期合作 目的是了解肌腱再生潜力有限的潜在机制，并开发新的 促进肌腱修复的方法。与胚胎和新生肌腱不同，损伤后新形成的肌腱 不具备固有的肌腱结构和力学性能。愈合异常瘢痕的肌腱 机械性能。损伤肌腱的结构和功能恢复的关键挑战之一是如何 为了增强参与肌腱愈合的肌腱前体细胞的修复能力，包括如何 刺激它们的肌腱分化，以及如何使它们重建肌腱基质结构。结果来自于 临床和翻译研究表明，葡萄糖代谢和肌腱愈合能力之间存在密切联系。 我们发现，葡萄糖的抑制剂2-脱氧葡萄糖(2DG)可以促进胶原纤维的恢复 在受伤的肌腱中排列。2DG在肌腱愈合中作用的蛋白质组学研究表明 2DG刺激损伤肌腱的肌动蛋白细胞骨架信号。出乎意料的是，我们发现 肌腱物质表达肌型肌球蛋白II成分，但表达强烈下调 而2DG则恢复了这些变化。 此外，损伤的肌腱来源的祖细胞对2DG有反应，上调了肌肉类型的表达 肌球蛋白轻链和重排的肌动蛋白细胞骨架。此前的研究表明，肌动蛋白 细胞骨架组织和肌球蛋白II活性是胶原纤维排列所必需的。总而言之，我们 损伤肌腱中肌球蛋白II分子上调可能刺激胶原纤维的假说 对齐。此外，葡萄糖控制着肌型肌球蛋白II分子和肌动蛋白的表达。 细胞骨架组织。为了检验这些假设，我们提出了两个目标。在目标1中，我们将确定更改 肌球蛋白II分子在肌腱损伤、愈合和再生过程中的变化。肌球蛋白的时空变化 II组分(肌球蛋白重链和轻链)将在损伤的肌腱中使用四个小鼠跟腱进行定义 肌腱损伤模型。在目标2中，我们将确定肌球蛋白II在肌腱损伤中的作用。 细胞(在TPC中)。在单一或三维培养中，用2DG处理InTPC，并观察肌球蛋白表达的变化 II分子、RhoA/RAC活性、胶原纤维排列和机械性能将被检测。此外, 在同一模型中，将确定肌肉肌球蛋白II分子的功能增强和功能丧失的影响。 就我们所知，目前还不知道肌肉和非肌肉肌球蛋白II分子是否具有 在肌腱中具有独特的功能。因此，该项目符合R21的范围，结果提供了第一个 回答这个问题的线索。

Tendon injury and disorders provide a severe negative impact on the economic cost and the ability to meet the patients' occupational, recreational, and health activities. Despite continuous efforts to improve the therapeutic modalities for tendon repair, truly effective and efficient therapies have not been established yet. Our long-term goal is to understand the mechanisms underlying the limited regenerative potential of tendons and develop new methods for stimulating tendon repair. Unlike embryonic and neonatal tendons, neo-formed tendons after injury do not possess native tendon structure and mechanical properties. Healing tendons from scar with abnormal mechanical properties. One of the key challenges for recovery of structure and function of injured tendons is how to potentiate the reparative capacity of the tendon progenitor cells involved in tendon healing, including how to stimulate their tenogenic differentiation and how to make them rebuild tendon matrix structure. The results from clinical and translational studies indicate a close link between glucose metabolism and tendon healing capacity. We have found that 2-deoxy glucose (2DG), an inhibitor of glucose, stimulated recovery of collagen fiber alignment in injured tendons. Proteomics studies on the actions of 2DG on tendon healing demonstrated that 2DG stimulated the actin cytoskeletal signaling in injured tendons. Unexpectedly, we found that the mid- substance of tendons expressed muscle type myosin II components but a strongly down-regulated expression of these molecules when injured, whereas 2DG restored these changes. Furthermore, injured tendon-derived progenitor cells responded to 2DG, up-regulated expression of muscle type myosin light chains, and rearranged actin cytoskeletons. Previous studies have demonstrated that actin cytoskeletal organization and myosin II activity are required for collagen fiber alignment. Taken together, we hypothesize that upregulation of muscle type myosin II molecules in injured tendons may stimulate collagen fiber alignment. Furthermore, that glucose controls the expression of muscle type myosin II molecules and the actin cytoskeletal organization. To test these hypotheses, we propose two aims. In Aim 1, we will determine alterations of myosin II molecules in tendons during injury, healing, and regeneration. Spatiotemporal changes in the myosin II components (myosin heavy and light chains) will be defined in injured tendons using four mouse Achilles tendon injury models. In Aim 2, we will determine the roles of muscle type of myosin II in injured tendon-derived cells (inTPCs). InTPCs will be treated with 2DG in mono- or 3D-culture, and changes in the expression of myosin II molecules, RhoA/Rac activity, collagen fiber alignment, and mechanical properties will be examined. In addition, in the same model, the effects of gain- and loss-of-function of muscle myosin II molecules will be determined. Best of our knowledge, it has not been known whether muscle and non-muscle myosin II molecules have an exclusive function in tendons. Thus, this project fits into the scope of R21, and the outcome provides the first clues to answer this question.