Regulation of renal and bone marrow injury by extracellular vesicle non-coding RN

描述（由申请人提供）：最近的研究表明，细胞源性囊泡可以通过RNA物种的转移改变单独靶细胞的基因组表型。进一步的研究表明，间充质干细胞衍生囊泡（MSC-dv）可以通过microRNA的转移介导损伤肾和骨髓组织的愈合。一个关键的需要是确定MSC-dv是否可以用来逆转人类的肾脏或骨髓损伤。目前的建议是确定分离和表征携带“愈合”microRNA的MSC-dv的最佳方法，这些microRNA可用于甘油暴露或缺血/再灌注损伤的肾组织或照射损伤的骨髓组织（干细胞）。“愈合”的MSC-dv中的microRNA将通过深度测序确定，具体的“愈合”microRNA将通过其递送到受损肾和骨髓细胞系模型并恢复增殖和减少凋亡的能力来确定。这将通过对受损细胞系进行脂肪转染和分析增殖和凋亡来实现。我们还将描述用特定的“愈合”microRNA装载“愈合”MSC-dv的最佳方法。预计这些研究将产生以下预期结果：对微RNA在组织修复中的作用的理解，2)对负责细胞命运变化的特定微RNA或微RNA亚群的定义，以及3)开发一种递送MSC囊泡微RNA以修复受损肾脏或骨髓组织的方法。这是建议的第一阶段。在第二阶段，在确定MCS-dv中包装最佳的“愈合”microRNA后，我们下一步需要优化组织修复的递送时间表，评估早期和晚期毒性，以及注入剂的分布。此外，我们还需要评估储存的富含microRNA的MSC-d囊泡“愈合”表型的稳定性。最佳愈合水平将在时间过程实验中评估，在我们的模型中，不同的时间和频率的输注被评估为愈合效果。在这些研究过程中，我们还将确定输注动物的早期和晚期毒性。还将评估动物总器官组织学和生存率。

DESCRIPTION (provided by applicant): Recent studies indicate that cellular derived vesicles can alter the genomic phenotype of separate target cells by transfer of RNA species. Further work has indicated that mesenchymal stem cell-derived vesicles (MSC-dv) can mediate healing of injured renal and marrow tissue by transfer of microRNA. A critical need is to determine whether MSC-dv can be utilized to reverse kidney or marrow damage in humans so afflicted. The present proposal is to define the optimum approach for separating and characterizing MSC-dv carrying "healing" microRNA to renal tissue injured by glycerol exposure or by ischemia/reperfusion or marrow tissue (stem cells) injured by irradiation. The microRNA in "healing" MSC-dv will then be determined by deep sequencing and the specific "healing" microRNA determined by its capacity to be delivered to injured renal and marrow cell line models and restore proliferation and reduce apoptosis. This will be carried out by lipofecting injured cell lines and analyzing proliferation and apoptosis. We will also characterize optimal approaches to loading "healing" MSC-dv with specific "healing" microRNA(s). It is anticipated that these studies will yield the following expected outcomes: 1.) An understanding of the role of micro RNAs in tissue restoration, 2.) a definition of the specific micro RNA or subset of microRNAs responsible for cell fate change and 3.) the development an approach for the delivery of MSC vesicle microRNA to restore injured renal or marrow tissue. This constitutes Stage 1 of the proposal. In Stage 2, after the determination of a "healing" microRNA packaged optimally in MCS-dv for delivery to injured marrow or renal tissue, we next need to optimize delivery schedule for tissue repair, evaluate early and late toxicity, and distribution of the infued agent. In addition, we need to evaluate the stability of "healing" phenotype of the microRNA rich MSC-d vesicles with storage. Optimal healing levels will be assessed in time course experiments where different times and frequencies of infusion are evaluated as to healing effects in our models. During the course of these studies we will also determine early and late toxicities in the infused animals. Total animal organ histology and survival will also be assessed. Next we will track tissue distribution after an optimal infusion schedule and determine the stability of MSC-dv over time under different conditions. The totality of the studies outlined in Stage 1 and 2 are unique and innovative. The delivery of microRNA in MSC-dv to heal tissue injuries is designed to prepare the basis for therapeutic interventions in different renal and marrow diseases which have not been previously attempted.