Engineering vascularized tissue in vivo using postnatal progenitor cells

其目标是开发一种基于细胞的技术，通过以下方式在体内设计血管组织 使用可以通过非侵入性方式从患者身上获得的出生后祖细胞。特别是，这一点 提案将重点放在联合使用骨髓间充质祖细胞(MPC)和 人血内皮祖细胞在体内形成脂肪组织。要测试的概念 含有高度纯化和定义的MPC和EPC的植入物是否会导致 体内植入后的血管化组织。我们假设，为了实现这一点，1)MPC和 EPC首先需要创建一个血管网络，允许通过植入物进行血液灌流；以及2)这 血管化将促进尚未组装到血管网络中的MPC经历适当的 通过采用植入部位周围组织的表型来进行组织发育。在这 建议，我们将确定关键参数，以加速血管生成过程的时间框架24-48 使用我们用人内皮祖细胞和巨噬细胞建立的体内模型。接下来，我们将确定 植入物中发现的脂肪细胞是否起源于人类，或者它们是否被招募到 一旦功能性血管网络被修复，来自宿主的植入作为内源性组织修复过程的一部分 如果是这样的话。有了这些信息，我们将实施增强体内分化过程的策略 以获得功能齐全的组织为目标。最后，我们将通过评估来测试这种基于细胞的技术 我们是否能创造出持久的脂肪组织垫，随着时间的推移呈现低体积减少。这个 解决这些目标的实验方法包括：1)植入物中血液灌流的活体成像 用基于荧光素酶的生物发光2)GFP标记移植细胞来追踪它们的特定贡献 3)共聚焦显微镜通过免疫荧光染色对种植体进行评价。 我们设想这个两个单元、两个步骤的系统是一种使能技术，可以应用于许多不同的 功能血管必不可少的组织。

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.