细胞外囊泡在间充质干细胞/内皮细胞共培养成血管化中的调控机制研究

Blood vessel regeneration is one of the critical fields in tissue engineering. Recent evidence has shown that a co-culture model of mesenchymal stem cells (MSCs) and endothelial cells (ECs) displayed stimulated angiogenesis and vascularization, which were highly promising findings for blood vessel engineering applications. Therefore, a precise understanding of the molecular mechanisms of communication between MSCs and ECs in co-culture induced vascularization will serve as the principle for future blood vessel tissue engineering designs..Extracellular vesicles (EVs) were recently demonstrated to have a fundamental role in the cross-talk between stem cells and ECs in various situations both in vivo and in vitro, and in some cases to participate in angiogenic induction. In particular, a recently discovered subpopulation within the EV family, termed migrasome, might contribute to this dynamic process. With this funding application, we aim to interrogate the precise molecular mechanism of EV-mediated vascularization using both 2D and 3D in vitro co-culture models, as well as an in vitro animal ischemia model. We will integrate single-cell “omics”, single-molecule RNA in situ hybridization, as well as advanced bioinformatics analysis to precisely depict the dynamic remodeling process during vascularization stimulated by MSC-EC communication, and use this knowledge to understand how EVs (particularly the migrasome population) serve as the signaling mediator which regulates this process. The proposed regulation mechanism will be further validated experimentally using a mouse hindlimb ischemia model, and this deepened understanding will contribute to future blood vessel regeneration and complex tissue engineering applications.

血管再生是组织工程及再生医学中重要的研究领域之一。近年来体内和体外的研究表明以间充质干细胞和血管内皮细胞为混合种子细胞的共培养模型可促进血管再生，有望为血管修复提供可能。因此，深入了解干细胞与内皮细胞间相互调控的作用机理，是今后实现有效、可控血管修复的理论基础。.现有研究表明干细胞分泌的外囊泡在干细胞和内皮细胞间通讯中起关键作用，并最终激活了血管化的发生，而细胞外囊泡家族中的迁移小体可能起到了关键的调控作用。本项目将基于体外二维体系及三维凝胶体系干细胞/内皮细胞共培养形成类血管环状结构为模型，重点利用单细胞基因组、单分子手段精细化的在单细胞水平分析干细胞与内皮相互作用产生血管化的过程，并尝试阐释由细胞外囊泡中的迁移小体介导的体外共培养成血管的信号分子调控机制，并通过小鼠后肢动脉结扎缺血致血管再生模型进行体内的验证，为今后可控地进行血管修复及组织工程血管构建提供理论基础。

干细胞介导的血管再生是解决血管梗死，血管壁结构破坏等病理发生的主要研究手段。近年来，关于干细胞与血管发生内皮细胞之间的交流方式众说纷纭，在我们的项目研究中，旨在探究细胞外囊泡或迁移小体对细胞间通讯的积极作用，利用单细胞转录组学精准化分析干细胞介导的血管发生动态过程，并阐明细胞外囊泡在成血管信号分子中的调控机制。值得注意的是，根据MSCs与ECs的体外共培养模型以及单细胞转录组学分析结果，细胞外囊泡或不是影响细胞间通讯最直接且主要的因素。相反，细胞外基质的分泌与降解是能够形成类血管化网状结构的必要充分条件；通过对这一过程提出的动态迁移假说，建立双态玻尔兹曼数学模型，利用该模型中参数的性质进一步验证了细胞外基质动态调控内皮细胞迁移行为的观点，与此同时本课题还结合合成生物学底盘细胞改造思想，对细胞表达水平进行lncRNA水平调控，为今后可控的进行血管修复以及组织工程血管的建立提供理论基础。本课题组研究期间，共在 FASEB journal，Molecular Biology，Synthetic and Systems Biotechnology, Applied Materials Today 等SCI杂志发表4篇文章，并有3篇文章已被Bioactive materials, Cancer Biology & Medicine, Quantitative Biology杂志接收；2篇文章在投。