Controlled Spatiotemporal Delivery of miRNA Anatgomir for Promoting Vascular Self

描述（由申请人提供）： 先前的工作已经证明，分化良好的人内皮细胞（EC）植入免疫缺陷小鼠宿主后，将在体外和体内的蛋白凝胶中自组装成血管管道。仅含有EC的移植物中的血管成熟需要募集宿主壁细胞，例如血管平滑肌细胞或周细胞（PC）。当EC与人PC共植入时，血管的成熟加速并增强。血管自组装也可以通过持续递送作用于EC或PC的促血管生成蛋白来增强，特别是当EC导向剂血管内皮生长因子（VEGF）与PC导向剂单核细胞趋化蛋白-1（MCP-1）组合时。然而，血管自组装和成熟仍然显得太慢，无法优化实质细胞存活，需要至少10天。促血管生成蛋白的作用可以分别通过靶细胞内涉及microRNA（miRNA）的正反馈环和负反馈环来增强或限制。miRNA是短的非编码RNA，通过减少特定mRNA的半衰期或翻译来调节多种发育过程。单个miRNA可以减少通常在同一途径中的多个基因的表达。miRNAs的作用可以被称为miRNAs的互补短RNA序列抑制。当miRNA以细胞特异性方式表达时，拮抗剂以细胞特异性方式起作用。该项目测试了一种假设，即控制递送Escheromir可以增强血管生成蛋白（如VEGF）在血管自组装中的治疗益处。这一假设将通过两个具体目标进行检验。在目标1中，聚合物纳米颗粒（NP）将用于找到在3D培养中提供对miRNA和Eschomir递送至EC细胞质的空间和时间控制的最佳方法。在目标2中，将测试这些NP递送系统控制将西莫司递送至miR-17/20的空间和时间递送的能力-已知其增强VEGF对通过将EC和PC悬浮在胶原蛋白和纤连蛋白的凝胶中产生的3D细胞培养物的作用。 (End摘要）

DESCRIPTION (provided by applicant): Previous work has demonstrated that well differentiated human endothelial cells (ECs) will self assemble into vascular conduits in protein gels both in vitro and in vivo after implantation into immunodeficient mouse hosts. Vessel maturation in grafts containing only EC requires recruitment of host mural cells, such as vascular smooth muscle cells or pericytes (PCs). The maturation of vessels is accelerated and enhanced when ECs are co-implanted with human PCs. Vessel self assembly can also be enhanced by sustained delivery of pro- angiogenic proteins that act on ECs or PCs, especially when an EC-directed agent, vascular endothelial growth factor (VEGF), is combined with a PC-directed agent, monocyte chemotactic protein -1 (MCP-1). However, vessel self-assembly and maturation still appears too slow to optimize parenchymal cell survival, requiring at least 10 days. The actions of pro-angiogenic proteins may be augmented or limited by positive and negative feedback loops, respectively, within the target cells that involve microRNAs (miRNAs). miRNAs are short, non-coding RNAs that regulate a variety of development processes by reducing specific mRNA half lives or translation. A single miRNA can reduce the expression of multiple genes often in the same pathway. The effects of miRNAs can be inhibited by complementary short RNA sequences referred to as antagomirs. Antagomirs act in a cell-specific manner when the miRNA is expressed in a cell specific manner. This project tests the hypothesis that controlled delivery of an antagomir can enhance the therapeutic benefits of angiogenic proteins such as VEGF in vascular self-assembly. This hypothesis will be tested through two specific aims. In Aim 1, polymer nanoparticles (NP) will be used to find the optimal approaches for providing spatial and temporal control over miRNA and antagomir delivery to the cytoplasm of ECs in 3D culture. In Aim 2, these NP delivery systems will be tested for their ability to control the spatial and temporal delivery of antagomirs to miR-17/20-which is known to augment the effects of VEGF-to 3D cell cultures produced by suspending ECs and PCs in gels of collagen and fibronectin. (End of Abstract)