The mechanics of host cell repopulation of engineered tissues

项目总结/摘要 我们建议确定血液动力学环境如何调节附着， 侵入和宿主细胞分化为“现成的”脱细胞组织工程化 心脏瓣膜（TEHV）。我们假设动态机械拉伸和流体剪切应力 通过增强和排列3D基质粘附来调节TEHV基质的再增殖， 从基质中激活潜在的TGF-β。为了验证我们的假设， 在可拉伸的威尔斯孔和微流体室中浇铸成纤维细胞， 各向同性或对齐的新组织，然后原位脱细胞。然后我们将量化 血管和循环细胞在周期性拉伸下粘附并侵入基质（Aim 1)和与体内植入相关的动态流动条件（目标2）。细胞附着， 浸润、增殖、凋亡、表型和内皮-间质转化 将随时间定量监测标记物。TGF-β活化和3D基质粘附 将检查蛋白质含量和排列，以及相关的信号转导途径。 将被询问以确定控制细胞反应的机制。结果 从这个系统的研究将有直接的影响TEHV的发展，通过确定 有助于（或阻碍）瓣膜基质的宿主细胞再增殖的信号， 在复杂的体内条件下适应性重塑的瓣膜设计。

Project Summary/Abstract We propose to determine how the hemodynamic environment regulates the attachment, invasion, and differentiation of host cells into “off-the-shelf” decellularized tissue engineered heart valves (TEHVs). We hypothesize that dynamic mechanical stretch and fluid shear stress regulate repopulation of the TEHV matrix by enhancing and aligning 3D matrix adhesions and activating latent TGF-beta from the matrix. To test our hypothesis, biopolymer scaffolds seeded with fibroblasts will be cast in stretchable wells and microfluidic chambers until remodeled into isotropic or aligned neo-tissues and then decellularized in situ. We will then quantify the extent to which vascular and circulating cells adhere to and invade the matrix under cyclic stretch (Aim 1) and dynamic flow conditions (Aim 2) relevant to in vivo implantation. Cell attachment, infiltration, proliferation, apoptosis, phenotype, and endothelial-to-mesenchymal transition markers will be quantitatively monitored over time. TGF-beta activation and 3D matrix adhesion protein content and alignment will be examined, and associated signal transduction pathways will be interrogated to determine the mechanisms governing the cell responses. The results from this systematic study will have a direct impact on TEHV development by determining the signals that aid (or hinder) host cell repopulation of the valve matrix with the goal of optimizing valve design for adaptive remodeling under complex in vivo conditions.