Use of Proteoglycan-Genes to Engineer Vascular Tissue

描述(由申请人提供)： 血管组织工程的主要限制之一是工程化组织不能形成功能完整的弹性纤维。我们实验室最近的工作发现，通过基因工程使动脉平滑肌细胞过度表达特定亚型的细胞外基质蛋白多糖，当转导细胞种植到损伤的动脉中时，在体外和体内都可以形成弹性纤维。我们假设这项技术可以用来制造具有完整和功能弹性纤维的血管替代物和血管移植物。为了解决这一假设，这一建议有以下具体目标：1.优化基因传递系统，以便能够以受控的方式实现最佳表达水平，以最大限度地形成弹性纤维。2.在体外用完整的弹性纤维制造工程细胞的“薄片”，用来形成具有最大弹性特性的小直径管。3.评估将工程细胞种植到包裹在框架周围的基于微纤维的可生物降解聚合物支架上以创建夹杂有功能弹性纤维的平滑肌细胞层的可行性；以及4.确定将这些弹性纤维生成细胞整合到胶原基管状支架中是否会导致形成具有适当拉伸强度和弹性特性的管。这项应用的总体目标是创建一种血管中层，它具有模仿天然血管的适当的细胞外基质成分的混合物，并包含完整的和功能性的弹性纤维。

DESCRIPTION (provided by applicant): One of the major limitations in vascular tissue engineering is the failure of the engineered tissue to form functional and intact elastic fibers. Recent work in our laboratory has found that by genetically engineering arterial smooth muscle cells to overexpress a specific isoform of the extracellular matrix proteoglycan, versican, elastic fibers form in vitro and in vivo when the transduced cells are seeded into injured arteries. We hypothesize that this technology can be used to create blood vessel substitutes and vascular grafts with intact and functional elastic fibers. To address this hypothesis, this proposal has the following specific aims: 1. Optimize the gene delivery system so that optimal levels of expression can be achieved in a controlled fashion to maximize elastic fiber formation. 2. Create "sheets" of engineered cells in vitro with intact elastic fibers to be used to form small diametric tubes which exhibit maximal elastogenic properties. 3. Assess the feasibility of seeding engineered cells onto micro-fiber based biodegradable polymer scaffolds wrapped around a frame to create layers of smooth muscle cells interspersed with functional elastic fibers and; 4. Determine if integrating these elastic fiber generating cells into collagen based tubular scaffolds will result in the formation of tubes with appropriate tensile strength and elastogenic properties. The overall goal of this application is to create a vascular medial layer that possesses an appropriate mixture of extracellular matrix components that mimic the natural vessel and contain intact and functional elastic fibers.