Modulating Cell-fate to Promote Regenerative Tendon Healing

肌腱损伤通过慢性沉积过多、紊乱的细胞外基质以纤维化的方式愈合。 肌成纤维细胞是许多组织纤维化的关键驱动力，新出现的证据表明 肌成纤维细胞的异质性。也就是说，具有与变化相对应的独特分子特征的肌成纤维细胞 肌成纤维细胞的功能。重要的是，肌成纤维细胞来源的成纤维细胞谱系也起到了 在决定肌成纤维细胞功能中起主要作用。然而，人们对肌成纤维细胞的动力学知之甚少。 在纤维肌腱愈合过程中，包括它们的成纤维细胞/腱细胞系，它们的功能如何随着时间的推移而变化， 这些不同的肌成纤维细胞亚型的分子图谱是什么，以及肌成纤维细胞是如何与 其他细胞，如巨噬细胞，介导愈合和纤维化。我们已经确定了硬化轴的血统和 S100a4系细胞是肌腱愈合过程中肌成纤维细胞的主要前体细胞群。 有趣的是，S100a4-细胞的耗尽损害了早期的肌腱愈合，而SCX-细胞的耗竭在后期得到改善 肌腱愈合提示这些细胞可能形成功能不同的肌成纤维细胞群。在……里面 此外，我们的初步数据确认巨噬细胞是腱细胞-肌成纤维细胞的关键驱动细胞。 过渡，我们观察到在晚期愈合过程中巨噬细胞的存在时间延长，伴随着核因子B- 肌成纤维细胞中介导的促生存信号。这些数据与促纤维化反馈环一致 巨噬细胞和肌成纤维细胞之间的相互作用，以维持许多组织的纤维化。因此，在本研究中，我们 将使用急性肌腱损伤和修复的小鼠模型严格定义肌腱细胞系特异性 在愈合过程中对肌成纤维细胞命运的贡献和确定肌成纤维细胞的异质性。我们将测试 抑制巨噬细胞介导的肌成纤维细胞分化和核因子-B介导的中心假说 谱系特异性肌成纤维细胞的存活促进了再生肌腱的愈合。我们将检验这一假设 通过以下具体目标：目标1：定义颞肌和腱细胞血统依赖 纤维肌腱愈合过程中肌成纤维细胞的免疫表型。目标2：确定以下要求 外源性巨噬细胞通过调节肌腱细胞-肌成纤维细胞转变促进纤维化愈合 抑制巨噬细胞募集的翻译潜力。目标3：证明扰乱肌成纤维细胞 存活或巨噬细胞持续存在可抑制持续性纤维化，促进再生肌腱愈合。 这些研究的成功完成将建立肌成纤维细胞的谱系、分子图谱和激活 在纤维化愈合过程中随时间变化的机制，并确定持续性肌成纤维细胞和 巨噬细胞作为促进肌腱愈合的新手段。

Tendon injuries heal in a fibrotic manner via chronic deposition of excessive, disorganized extracellular matrix. Myofibroblasts are a critical driver of fibrosis in many tissues, and emerging evidence demonstrates myofibroblast heterogeneity. That is, myofibroblasts with unique molecular profiles that correspond to changes in myofibroblast function. Importantly, the fibroblast lineage from which myofibroblasts are derived also plays a major role in dictating myofibroblast function. However, very little is known about myofibroblast dynamics during fibrotic tendon healing, including their fibroblast/tenocyte-lineage, how their functions change over time, what the molecular profiles of these different myofibroblast subtypes are, and how myofibroblasts interact with other cells, such as macrophages, to mediate healing and fibrosis. We have identified Scleraxis-lineage and S100a4-lineage cells as the predominant myofibroblast precursor populations during tendon healing. Interestingly, depletion of S100a4-cells impairs early tendon healing, while Scx-cell depletion improves late tendon healing suggesting these cells may give rise to functionally distinct myofibroblast populations. In addition, our preliminary data identifies macrophages as a critical driver of the tenocyte-myofibroblast transition, and we observed prolonged macrophage presence during late healing, concomitant with NFB- mediated pro-survival signaling in myofibroblasts. These data are consistent with a pro-fibrotic feedback loop between macrophages and myofibroblasts to sustain fibrosis in many tissues. Thus, in the present study we will use a murine model of acute tendon injury and repair to rigorously define tenocyte lineage-specific contributions to myofibroblast fate and define myofibroblast heterogeneity during healing. We will test the central hypothesis that inhibiting macrophage-mediated myofibroblast differentiation and NF-B-mediated survival of lineage-specific myofibroblasts promotes regenerative tendon healing. We will test this hypothesis through the following specific aims: Aim 1: Define the temporal and tenocyte lineage-dependent immunophenotype of myofibroblasts during fibrotic tendon healing. Aim 2: Establish the requirement for extrinsic macrophages in fibrotic healing via modulation of the tenocyte-myofibroblast transition and test the translational potential of inhibiting macrophage recruitment. Aim 3: Demonstrate that disrupting myofibroblast survival or macrophage persistence inhibits sustained fibrosis and promotes regenerative tendon healing. Successful completion of these studies will establish myofibroblast lineage, molecular profile and activation mechanisms over time during fibrotic healing and define disruption of persistent myofibroblasts and macrophages as a novel means to improve tendon healing.