Cell Therapy for Neuroprotection in Congenital Heart Disease

项目总结 严重的神经延迟正在成为儿童面临的最重要的当前挑战之一 先天性心脏病(CHD)，但目前可用的治疗选择很少。在我们的第一期资金中， 我们提出将体外循环(CPB)作为一种细胞输送系统应用于婴幼儿先心病 改善冠心病患者神经功能损害的途径。我们已出版和未出版的作品有 证明了这种方法的有效性和实用性，确定了交付骨髓的全身影响- 体外循环诱导间充质干细胞及其对脑白质和脑室下的影响 开发区(SVZ)开发。最值得注意的是，第一个奖项成功地导致了一期临床试验的发展。 在国家少年儿童协会被称为“MedCap”。使用我们的翻译仔猪模型，我们已经演示了细胞， 通过CPB交付BM-MSC并生成关键信息后，结构、行为和行为得到改善 用于从长椅到床边的翻译。然而，BM-MSCs治疗作用的潜在机制仍然 在很大程度上仍然不为人知。此次R01续订将解决关键的知识差距，目标是进一步 加强我们基于细胞的治疗，以保护CHD人群的神经。Exosome是一类 胞外囊泡，内含微小核糖核酸(MiRNA)等生物活性分子。源自BM的外体- 骨髓间充质干细胞(BM-MSCexo)可以在对周围细胞和组织的作用中发挥重要作用，并诱导出有利于 对各种疾病的反应。我们已经建立了一条细胞后传递整合转录的管道 骨髓间充质干细胞和宿主组织外体miRNAs的分析。我们的初步研究已经确定 BM-MSCexo来源的miRNAs可能是减少神经元凋亡和小胶质细胞激活的关键驱动因素 观察BM-MSC治疗后大脑皮层的变化。这项更新提案的首要目标是 从关键细胞群中建立用于组织修复和再生的详细分子特征 BM-MSC交付后的细胞分辨率；然后我们将使用这些分子签名作为路线图来识别新的 BM-MSCexo内解释组织损伤后疾病修饰生物活性的分子实体 儿科心脏手术。更新研究将检验我们的中心假设，即特定的外体货物 BM-MSCs的成分通过神经前体细胞和 调节性T细胞，从而改善神经预后和术后病程。要追溯识别 关键的外体生物活性分子，我们将确定三个转录和染色质景观 特殊细胞群：1)SVZ神经干细胞和祖细胞；2)WM少突胶质细胞；3)调节性T细胞 细胞。再加上我们正在进行的基于前一个奖项的临床试验，确定了分子 BM-MSC治疗和挖掘特异性BM-MSCexo用于独特的CPB病理将具有重要意义 提高我们对这种基于细胞的治疗的理解，并将为潜在的 CHD儿童神经保护的无细胞间充质干细胞治疗。

PROJECT SUMMARY Significant neurological delay is emerging as one the most important current challenges for children with congenital heart disease (CHD), yet few treatment options are currently available. In our first period of funding, we proposed the use of cardiopulmonary bypass (CPB) as a cell delivery system in infants with CHD as a novel approach for improving the neurological impairments in CHD. Our published and unpublished work has demonstrated the efficacy and utility of this approach, determining the systemic effects of delivery bone marrow- derived mesenchymal stromal cell (BM-MSC) via CPB and the effect on white matter (WM) and sub-ventricular zone (SVZ) development. Most notably, the first award successfully led to development of a phase 1 clinical trial termed “MeDCaP” at Children’s National. Using our translational piglet model, we have demonstrated cellular, structural, and behavioral improvements after BM-MSC delivery through CPB and generated critical information for bench-to-bedside translation. However, the mechanisms underlying the therapeutic action of BM-MSCs still remain largely unknown. This R01 renewal will address the key knowledge gaps with the goal of further enhancing our cell-based treatment for neuroprotection in the CHD population. Exosome is a class of extracellular vesicles loaded with bioactive molecules such as microRNA (miRNA). Exosomes derived from BM- MSCs (BM-MSCexo) can play a major role in the effects on surrounding cells and tissues and elicit favorable responses in various diseases. We have established a pipeline for post-cell delivery integrated transcriptomic analysis of exosomal miRNAs from BM-MSCs and host tissue mRNA. Our preliminary studies have identified the BM-MSCexo-derived miRNAs as putative key drivers of reduced neuronal apoptosis and microglial activation observed after BM-MSC treatment in the cerebral cortex. The overarching goal of this renewal proposal is to establish detailed molecular signatures from critical cell populations for tissue repair and regeneration at single cell resolution after BM-MSC delivery; we will then use those molecular signatures as roadmaps to identify novel molecular entities within the BM-MSCexo that account for the disease-modifying bioactivity in tissue injury after pediatric cardiac surgery. The renewal studies will test our central hypothesis that specific exosomal cargo constituents from BM-MSCs promote repair and regenerative processes both through neural progenitors and regulatory T cells, thereby improving neurological outcomes and post-operative course. To retroactively identify key exosomal bioactive molecules, we will determine the transcriptional and chromatin landscape of three specific cell populations: 1) SVZ neural stem and progenitor cells; 2) WM oligodendrocytes; and 3) regulatory T cells. Together with our ongoing clinical trial established based on the previous award, identifying molecular signatures of BM-MSC treatment and mining specific BM-MSCexo for unique CPB pathology will significantly improve our understanding of this cell-based treatment and will provide a new therapeutic paradigm for potential cell-free MSC-based therapies for neuroprotection in children with CHD.