Stem Cell Oxygenation and Ischemic Tissue Regeneration

项目主任/首席调查员(末位、第一位、中位)：关建军 项目摘要 严重肢体缺血(CLI)是一种严重的外周动脉疾病，具有很高的肢体损失率和死亡率。它是 以低血液灌注率、广泛的组织缺血和骨骼肌退化为特征。快速血管化 恢复血液灌流和快速肌肉再生以恢复正常功能，代表着最佳目标 CLI治疗。目前还没有有效的治疗方法，尽管干细胞疗法是最有效的治疗方法之一。 前景看好的战略。大多数干细胞类型主要通过旁分泌促进血管形成和肌肉再生。 一些细胞也可能分化为内皮细胞和骨骼肌细胞。然而，目前的干细胞 治疗的效果很低，主要是由于极低的细胞存活率和旁分泌效应。 缺血肢体的氧合状况(1%)。在本项目中，我们提出了一种新的细胞递送系统，该系统可以连续 释放适当浓度的氧气，同时提高干细胞存活率和旁分泌效应， 导致快速的血管形成和肌肉再生。旁分泌效应同时提供多种增长 对血管形成和肌肉再生至关重要的因子，这是生长因子无法轻易实现的 心理治疗。 在我们的初步研究中，我们创造了一种基于水凝胶的细胞输送系统，可以释放氧气。当经过测试时 使用骨髓间充质干细胞(MSCs)，释放的O2增加了细胞在 体外缺血条件下不增加活性氧(ROS)含量。它还上调了MSC 旁分泌作用，特别是在分泌前血管生成/前肌肉生成生长因子方面，如PDGF和IGF-1。 在植入缺血肢体后，氧气释放细胞输送系统不仅增加了MSC的存活，而且 在4周内完全恢复血液灌流和肌肉收缩能力。骨髓间充质干细胞在血管生成中的作用 主要来自旁分泌作用，因为只有一小部分细胞分化为内皮细胞。 同时，MSC旁分泌效应和肌源性分化均有助于肌肉再生。这些 初步数据表明，增加MSC存活率和旁分泌效应可以显著增强 缺血肢体的血管形成和肌肉再生。然而，细胞存活和旁分泌效应并不总是如此。 同时增加。 基于我们的初步研究和上述讨论，我们假设干细胞输送系统与 同时提高MSC存活率和旁分泌效应的最佳O2释放谱将显著 促进缺血肢体的血管形成和肌肉再生。 目标1将检验这样一个假设，即最佳的氧气释放曲线将促进MSC的存活和旁分泌效应 在缺血条件下。 目的#2将使用后肢缺血模型测试所创建的细胞输送系统的有效性。 这个项目是创新的，因为它开发了一个安全和长期的氧气释放系统来建立作用， 控制性氧释放增强小鼠干细胞存活和旁分泌效应的机制和效果 用于加速再生的缺血组织。拟议的干细胞输送系统也是可翻译的。 OMB编号0925-0001/0002(03/16修订版批准至2018年10月31日)页面续格式页面

Program Director/Principal Investigator (Last, First, Middle): GUAN, JIANJUN Project Summary Critical limb ischemia (CLI) is a severe peripheral artery disease with high rates of limb loss and mortality. It is featured by low blood perfusion, extensive tissue ischemia, and degenerated skeletal muscle. Quick vascularization to restore blood perfusion, and fast muscle regeneration to restore normal function, represent the optimal goals for CLI treatment. Currently there is no efficient treatment available, although stem cell therapy is one of the most promising strategies. Most of stem cell types promote vascularization and muscle regeneration mainly by paracrine effects while some may also differentiate into endothelial and skeletal muscle cells. However, current stem cell therapy experiences low efficacy largely due to inferior cell survival and paracrine effects under the extremely low oxygen condition (<1%) of ischemic limbs. In this project, we propose a new cell delivery system that continuously releases appropriate concentration of O2 to simultaneously improve stem cell survival and paracrine effects, resulting in quick vascularization and muscle regeneration. Paracrine effects concurrently provide multiple growth factors critical for vascularization and muscle regeneration, which cannot be readily achieved by growth factor therapy. In our preliminary studies, we have created a hydrogel-based cell delivery system that releases O2. When tested using bone marrow-derived mesenchymal stem cells (MSCs), the released O2 increased cell survival under ischemic conditions in vitro without increasing reactive oxygen species (ROS) content. It also upregulated MSC paracrine effects especially in terms of secreting proangiogenic/promyogenic growth factors like PDGF and IGF-1. After implanting into ischemic limbs, the O2 releasing cell delivery system not only augmented MSC survival, but also fully restored blood perfusion and muscle contractility in 4 weeks. The contribution of MSCs to vascularization is mainly from paracrine effects as only a low percentage of cells were differentiated into endothelial cells. Meanwhile, both MSC paracrine effects and myogenic differentiation contributed to muscle regeneration. These preliminary data suggest that increasing both MSC survival and paracrine effects can significantly enhance vascularization and muscle regeneration in ischemic limbs. Yet, cell survival and paracrine effects do not always increase concurrently. Based on our preliminary studies and above discussion, we hypothesize that stem cell delivery systems with optimal O2 release profiles that simultaneously increase MSC survival and paracrine effects, will significantly accelerate vascularization and muscle regeneration in ischemic limbs. Aim #1 will test the hypothesis that optimal O2 release profiles will promote MSC survival and paracrine effects under ischemic conditions. Aim #2 will test efficacy of the created cell delivery systems using a model of hindlimb ischemia. This project is innovative because it develops a safe and long-term O2 release system to establish the role, mechanism, and efficacy of controlled O2 release in augmenting both stem cell survival and paracrine effects in ischemic tissues for accelerated regeneration. The proposed stem cell delivery system is also translational. OMB No. 0925-0001/0002 (Rev. 03/16 Approved Through 10/31/2018) Page Continuation Format Page