Engineering vascularized tissue in vivo using postnatal progenitor cells

我们的目标是开发一种基于细胞的技术，通过以下方式在体内构建血管化组织： 使用可以通过非侵入性手段从患者获得的出生后祖细胞。这尤其 该提案将集中在骨髓来源的间充质祖细胞（MPC）和 人血液来源的内皮祖细胞（EPCs）在体内形成脂肪组织。要测试的概念 含有高度纯化和确定的MPC和EPCs的植入物是否会导致特化 在体内植入后血管化的组织。我们假设，要发生这种情况，1）MPC和 EPCs首先需要创建一个血管网络，允许血液通过植入物灌注; 2）这 血管化将促进尚未组装成血管网络的MPC进行适当的血管化。 通过在植入部位采用周围组织的表型进行组织发育。在这 我们将确定关键参数，以加速血管生成过程的时间范围为24-48 小时使用我们已经建立的人EPCs和MPCs的体内模型。接下来，我们将确定 无论在植入物中发现的脂肪细胞是人类起源的还是它们被募集到 一旦功能性血管网络被破坏，作为内源性组织修复过程的一部分， 提供了有了这些信息，我们将实施策略，以加强在体内分化的过程中， 目的是获得功能齐全的组织。最后，我们将测试这种基于细胞的技术， 我们是否可以创造出持久的脂肪组织垫，随着时间的推移，脂肪组织垫的体积减少很少。的 实现这些目标的实验性方法包括：1）植入物中血液灌注的体内成像 2）植入细胞的GFP标记以追踪它们的特定贡献 3）共聚焦显微镜，通过免疫荧光染色评价植入物。 我们设想这种两细胞，两步系统作为一种使能技术，可以应用于许多不同的领域。 其中功能性血管是必需的组织。

The goal is to develop a cell-based technology that will allow engineering vascularized tissues in vivo by using postnatal progenitor cells that can be obtained from patients by non-invasive means. In particular, this proposal will focus on the combined use of bone marrow-derived mesenchymal progenitor cells (MPCs) and human blood-derived endothelial progenitor cells (EPCs) to form adipose tissue in vivo. The concept to test is whether implants containing highly purified and defined MPCs and EPCs will result in specialized vascularized tissues upon implantation in vivo. We hypothesize that for this to happen, 1) both MPCs and EPCs need first to create a vascular network that will allow blood perfusion through the implants; and 2) this vascularization will promote MPCs that have not assembled into the vascular network to undergo appropriate tissue development by adopting the phenotype ofthe surrounding tissue at the site of implantation. In this proposal, we will determine key parameters to accelerate the vasculogenic process to a time frame of 24-48 hours using the in vivo model we have established with human EPCs and MPCs. Next, we will determine whether the adipocytes found in the implants are human in origin or whether they are recruited into the implants from the host as part of an endogenous tissue repair process once a functional vascular network is provided. With this information, we will implement strategies to enhance the process of in vivo differentiation with the aim of having fully functional tissues. Finally, we will test this cell-based technology by evaluating whether we can create long-lasting adipose tissue pads that present low volume reduction over time. The expenmental approaches to address these aims include: 1) in vivo imaging of blood perfusion in the implants using luciferase-based bioluminescence 2) GFP labeling of implanted cells to track their specific contribution to the resulting tissues; 3) confocal microscopy to evaluate implants through immunofluorescence staining. We envision this two-cell, two-step system as an enabling technology that can be applied to many different tissues wherein functional blood vessels are essential.