Therapeutic Mechanisms of Human CD34 Exosomes

描述（由申请人提供）：基于干细胞的疗法，包括最近的CD 34+细胞疗法，是改善心脏再生和功能的有前景的治疗方法。CD 34+细胞移植的益处似乎主要是通过缺血心肌中CD 34+细胞分泌的旁分泌因子增加血管新生而发生的。然而，导致CD 34+细胞诱导的血管生长和治疗恢复的确切机制知之甚少。这种缺乏机制的洞察力是心脏干细胞治疗成功的关键障碍，因为移植干细胞的再生功效受到其在缺血心肌中的不良存活力和保留的限制。因此，为了解决这些局限性并开发新的替代方法，我们必须寻求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，来自人CD 34+细胞的旁分泌物含有称为外泌体（即CD 34+外泌体）的膜结合纳米囊泡，其具有与细胞相似的血管生成和治疗作用。此外，CD 34+外泌体携带并转移促血管生成miRNA（如miR-126）至心肌中的缺血内皮细胞并诱导其血管生成活性。我们建议的基本基础是利用CD 34+外泌体的再生潜力和通信能力来增强治疗方法。我们的中心假设是，通过人CD 34+细胞的旁分泌释放的外泌体通过将miRNA直接转移到心脏中的细胞来介导心肌修复。我们的目标是建立CD 34+干细胞来源的外泌体作为一种新的无细胞治疗实体的缺血性心肌修复，并发展外泌体诱导的心脏血管生成和治疗恢复的分子机制的全面理解。这项创新研究对于理解CD 34+细胞治疗的机制和解锁祖细胞衍生的外泌体的转化潜力非常重要。我们将在三个具体目标中检验我们的假设：1）确定无细胞的CD 34+外泌体在心肌缺血的鼠模型中的治疗功效，并确定CD 34+外泌体分泌是否是CD 34+细胞治疗的关键机制之一; 2）通过研究CD 34+外泌体诱导血管生成的分子机制来表征CD 34+外泌体诱导血管生成的分子机制。外泌体在缺血性心脏中的运输和信号传导机制; 3）在MI的鼠模型中建立由miR-126介导的CD 34+外泌体的有益作用的程度。本提案中描述的实验将通过表征CD 34+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。我们的研究将通过使用来自人类干细胞的外泌体作为合适的无细胞替代物，为新的治疗方法奠定基础。它有可能通过利用外泌体相对于细胞的许多实际和技术优势来推进基于细胞的治疗，以应用于心血管再生医学。

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.