Therapeutic Mechanisms of Human CD34 Exosomes

描述(申请人提供)：干细胞疗法，包括最近的CD34细胞疗法，是一种很有前途的改善心脏再生和功能的治疗方法。CD34细胞移植的好处似乎主要是通过增加缺血心肌中CD34细胞分泌的旁分泌因子促进血管生成来实现的。然而，导致CD34细胞诱导血管生长和治疗恢复的确切机制尚不清楚。这种对机制认识的缺乏是心脏干细胞治疗成功的关键障碍，因为移植的干细胞的再生效率受到它们在缺血心肌中的低活性和保留性的限制。因此，为了解决这些局限性并开发新的替代方法，我们必须努力了解导致治疗恢复的机制。我们先前发表的数据和我们的初步数据显示，人CD34细胞的旁分泌含有膜结合的纳米囊泡，称为外体(即CD34外体)，它与细胞类似，具有血管生成和治疗作用。此外，CD34外切体携带和转移促血管生成的miRNAs，如miR-126，到心肌中的缺血内皮细胞，并诱导它们的血管生成活性。我们建议的基本基础是利用CD34外切体的再生潜力和通讯能力来增强治疗方法。我们的中心假设是，通过人CD34细胞旁分泌释放的外切体通过将miRNAs直接转移到心脏细胞来介导心肌修复。我们的目标是建立CD34干细胞来源的外切体作为一种新的无细胞治疗实体，用于缺血性心肌修复，并全面了解外切体诱导的心脏血管生成和治疗恢复的分子机制。这项创新性的研究对于理解CD34细胞治疗的机制和解锁祖细胞来源的外切体的转化潜力是重要的。我们将在三个具体目标上验证我们的假设：1)确定无细胞CD34外切体在小鼠心肌缺血模型中的治疗效果，并确定CD34外切体的分泌是否是CD34细胞治疗的关键机制之一；2)通过研究CD34外切体在缺血心脏中的转运和信号机制来表征CD34外切体诱导血管生成的分子机制；3)在小鼠心肌梗死模型中建立CD34外切体的有益作用在多大程度上是由miR-126介导的。这项提案中描述的实验将通过描述CD34干细胞治疗的主要但尚未确定的机制来探索关键的科学问题。我们的研究将为一种新的治疗方法奠定基础，即利用人类干细胞的外切体作为合适的无细胞替代品。它通过利用外切体相对于细胞的许多实用和技术优势，在心血管再生医学中的应用，有可能促进基于细胞的治疗。

DESCRIPTION (provided by applicant): Stem cell-based therapies, including the recent CD34+ cell therapy, are a promising therapeutic approach for improving cardiac regeneration and function. The benefits of CD34+ cell transplantation appear to occur primarily via increase in vascular angiogenesis by the CD34+ cell-secreted paracrine factors in the ischemic myocardium. However, the precise mechanisms that lead to CD34+ cell-induced vessel growth and therapeutic recovery are poorly understood. This lack of mechanistic insight is a critical barrier to the success of cardiac stem cell therapy, as the regenerative efficacy of the transplanted stem cells is limited by their poor viability and retention in the ischemic myocardium. Therefore, to address these limitations and to develop novel alternate approaches, we must seek to understand the mechanisms that lead to therapeutic recovery. Our previously published data and our preliminary data have revealed that the paracrine secretion from human CD34+ cells contain membrane-bound nano-vesicles called exosomes (i.e. CD34+ exosomes), which are angiogenic and therapeutic similar to the cells. Further, CD34+ exosomes carry and transfer proangiogenic miRNAs, such as miR-126, to ischemic endothelial cells in the myocardium and induce their angiogenic activity. The fundamental basis of our proposal is to harness the regenerative potential and communication power of CD34+ exosomes to augment therapeutic approaches. Our central hypothesis is that exosomes released via paracrine secretion of human CD34+ cells mediate myocardial repair by direct transfer of miRNAs to cells in the heart. Our goal is to establish CD34+ stem cell-derived exosomes as a novel cell-free therapeutic entity for ischemic myocardial repair and to develop a comprehensive understanding of the molecular mechanisms of exosomes-induced cardiac angiogenesis and therapeutic recovery. This innovative study is important to understand the mechanisms of CD34+ cell therapy and to unlock the transformative potential of progenitor cell-derived exosomes. We will test our hypothesis in three specific aims: 1) Determine the therapeutic efficacy of cell-free CD34+ exosomes in a murine model of myocardial ischemia and determine whether CD34+ exosomes secretion is one of the key mechanisms of CD34+ cell therapy; 2) Characterize the molecular mechanisms of CD34+ exosomes-induced angiogenesis by studying the CD34+ exosomes trafficking and signaling mechanisms in the ischemic heart; 3) Establish the extent to which the beneficial effects of CD34+ exosomes are mediated by miR-126 in a murine model of MI. The experiments described in this proposal will explore key scientific questions by characterizing the predominant, but as yet undefined, mechanism of CD34+ stem cell therapy. Our studies will lay foundation to a novel therapeutic approach by using exosomes from human stem cells as a suitable cell-free alternative. It has the potential to advance cell- based therapis by exploiting many practical and technical advantages of exosomes relative to cells for application in cardiovascular regenerative medicine.