Restoration of myocardial reparative function of diabetic progenitor cells by epigenetic modulation

在缺血损伤后缺乏有效的内源性修复机制的情况下，基于细胞的疗法应运而生 作为一种潜在的治疗缺血组织修复的新方法。在推测的骨骼的初步特征之后 骨髓源性内皮祖细胞及其促进心脏和危重肢体缺血的潜能 为了新生血管和减轻缺血性损伤，十多年来紧张的临床前研究导致了 骨髓前体细胞/基于EPC的临床试验。然而，尽管早期热情高涨，基于细胞的疗法产生的效果并不明显。 临床结果。基于细胞的治疗的适度临床结果可能反映了已知的细胞功能障碍 随后在糖尿病动物和患者的EPC/祖细胞中获得。令人信服的证据表明 内皮祖细胞功能障碍是糖尿病血管修复和血管生成受损的一种机制 导致血管功能障碍。因此，了解糖尿病导致EPC功能障碍的分子基础 潜在地逆转EPC功能障碍可能代表着一种战略，以加强基于细胞的治疗 糖尿病患者的心肌/缺血肢体修复。越来越多的证据也表明，这种机制 高血糖记忆和EPC功能障碍可能涉及表观遗传机制，包括增强 血管基因上的表观遗传抑制标记导致其表观遗传沉默。此外，由于高血糖 记忆是通过细胞分裂来遗传的，糖尿病内皮祖细胞中改变的表观遗传模式可以传递给 子代细胞。了解糖尿病患者内皮祖细胞功能障碍的表观遗传学基础和表观遗传学重编程 因此，糖尿病内皮祖细胞对于糖尿病患者的细胞治疗至关重要。因此，我们的 中心假设是糖尿病内皮祖细胞中的表观遗传抑制标记使其功能障碍和表观遗传学 针对这些抑制标记的修饰剂可以将糖尿病内皮祖细胞重新编程为更具功能性和修复性的 表型。这个项目的目的是详细地研究表型、表观遗传学和分子特征。 糖尿病小鼠的重编程糖尿病内皮祖细胞以及糖尿病小鼠的人CD34+造血干细胞 患者及其外切体衍生物，并测试这些重新编程的心肌缺血修复能力 糖尿病内皮祖细胞在生理相关的心肌梗死和后肢缺血模型中的作用。这一总体目标将通过以下方式实现 在以下三个具体目标下组织实验：1)评估表型稳定性和 表观遗传重编程的糖尿病内皮祖细胞的修复潜力；2)确定特定的表观遗传学 重编程内皮祖细胞的修饰及HDAC1和G9a甲基转移酶在重编程中的作用 过程和3)建立表观遗传重编程以挽救人CD34+的功能和修复缺陷 来自2型糖尿病患者的干细胞。

In the absence of effective endogenous repair mechanisms after ischemic injury, cell-based therapies have emerged as a potential novel therapeutic approach in ischemic tissue repair. After the initial characterization of putative bone marrow-derived endothelial progenitor cells (EPC) and their potential to promote cardiac and critical limb ischemia neovascularization and to attenuate ischemic injury, more than a decade of intense preclinical research, led to the BM progenitors/EPC-based clinical trials. However, despite early enthusiasm, cell based therapies yielded modest clinical results. Modest clinical outcomes of cell-based therapies may reflect the cellular dysfunction that is known to ensue in EPC/progenitor cells obtained from diabetic animals as well as patients. Compelling evidence indicates that EPCs dysfunction represents a mechanism for impaired vascular repair and angiogenesis in diabetes, subsequently leading to vascular dysfunction. Therefore, understanding the molecular basis of diabetes-induced EPC dysfunction and potentially reversing EPC dysfunction may represent a strategy to enhance cell-based therapeutics for myocardial/ischemic limb repair in diabetic patients. Increasing evidence also indicates that the mechanism underlying hyperglycemic memory and EPC dysfunction may involve epigenetic mechanisms involving enhanced epigenetic repressive marks on vascular genes leading to their epigenetic silencing. Moreover, since hyperglycemic memory is inherited through cell division, and altered epigenetic patterns in diabetic EPCs can be transmitted to daughter cells. Understanding the epigenetic basis of EPC dysfunction in diabetics and epigenetic reprogramming of diabetic EPCs, is therefore, of paramount importance for cell based therapies in diabetic patients. Therefore, our central hypothesis is that epigenetic repressive marks in diabetic EPCs render them dysfunctional and epigenetic modifying agents targeting those repressive marks can reprogram diabetic EPCs to a more functional and reparative phenotype. This project aims to study, in detail, phenotypic, epigenetic and molecular characterization of reprogrammed diabetic EPCs from diabetic mice as well as human CD34+ hematopoietic stem cells from diabetic patients and their exosome derivatives and test the ischemic myocardial repair capacity of these reprogrammed diabetic EPCs in physiologically relevant model of MI and hind limb ischemia. This overall aim will be achieved by conducting experiments organized under the following three specific aims: 1) To evaluate phenotypic stability and reparative potential of epigenetically reprogrammed diabetic EPCs; 2) To determine the specific epigenetic modifications in reprogrammed EPCs and establish a role of HDAC1 and G9a methytransferase in reprogramming process and 3) To establish epigenetic reprogramming rescues functional and reparative deficits in human CD34+ stem cells from patients with Type 2 diabetes.