Controlled Spatiotemporal Delivery of miRNA Anatgomir for Promoting Vascular Self

描述（由申请人提供）： 先前的工作表明，分化良好的人内皮细胞（EC）在植入免疫缺陷小鼠宿主后，无论在体外还是在体内，都会在蛋白质凝胶中自组装成血管导管。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 与人类 PC 共同植入时，血管的成熟会加速并增强。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，无法优化实质细胞的存活，至少需要 10 天。在涉及 microRNA (miRNA) 的靶细胞内，促血管生成蛋白的作用可能分别受到正反馈环和负反馈环的增强或限制。 miRNA 是短的非编码 RNA，通过缩短特定 mRNA 半衰期或翻译来调节多种发育过程。单个 miRNA 可以降低同一途径中多个基因的表达。 miRNA 的作用可以被称为 antagomir 的互补短 RNA 序列抑制。当 miRNA 以细胞特异性方式表达时，Antagomirs 以细胞特异性方式发挥作用。该项目测试了以下假设：antagomir 的受控递送可以增强血管生成蛋白（例如 VEGF）在血管自组装中的治疗效果。这一假设将通过两个具体目标进行检验。在目标 1 中，聚合物纳米粒子 (NP) 将用于寻找最佳方法，以在 3D 培养中对 miRNA 和 antagomir 递送至 EC 细胞质提供空间和时间控制。在目标 2 中，将测试这些 NP 递送系统控制 miR-17/20 的空间和时间递送的能力，已知 miR-17/20 可以增强 VEGF 对通过将 EC 和 PC 悬浮在胶原和纤连蛋白凝胶中产生的 3D 细胞培养物的影响。 （摘要完）

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)