Hyaluronan Scaffold for Regenerating Elastin Matrices

描述(申请人提供)：由于机械损伤或获得性疾病(如动脉瘤)或其先天缺失或畸形导致的血管弹性蛋白基质的破坏，可能严重损害血管弹性和弹性蛋白介导的细胞信号，这些细胞信号对形态发生、损伤反应和炎症至关重要。弹性蛋白在去弹力化血管和组织工程化构建物中的再生受到成人血管平滑肌细胞(VSMCs)弹性蛋白产量低的限制。此外，细胞支架材料可以上调弹性蛋白的合成，并提供必要的生物线索，以再生忠实的模拟天然弹性蛋白基质尚未确定。先前我们已经确定，含有透明质酸(HA)的高度生物相容性的交联型支架(HYLANS)能刺激培养的VSMC合成与天然弹性蛋白相近的仿制品。透明质酸是ECM中的一种糖胺聚糖。有证据表明，是HA片段，而不是天然的长链HA引发了这些弹性反应，尽管长链HA可能有助于基质的稳定。然而，含有长链HA和HA片段混合物的弹性支架的进一步制备取决于以下方面：(I)表面拴系的HA或/和HA片段对成年VSMC的弹性形成的不同影响，这对于优化支架组成是重要的；(Ii)在预先排列的3D HA支架内的细胞培养比在2D表面上的细胞培养对模拟血管弹性蛋白超微结构的潜在好处；以及(Iii)同时提供的机械和生化信号对合成的弹性蛋白基质的质量(数量、结构、耐用性和力学)的影响。目的：本项目的目的是阐明上述参数对成年大鼠VSMC培养模型中弹性蛋白合成的影响。目的1根据HA对成人RVSMCs合成弹性蛋白的个别影响，优化HA表面拴系混合物中HA片段的大小分布。目标2将研究含有目标1中确定的优化成分的管状、电纺HA纳米纤维支架的独立和综合益处，循环机械应变和生长因子(转化生长因子-b、胰岛素样生长因子-1)补充基质质量。意义：研究结果将创造一种工具，可以与现有的血管设备集成，按需制造忠实的天然弹性蛋白仿制品。这种弹性蛋白模拟物将有助于增强和修复变性血管中的弹性蛋白，也将使研究弹性形成的体外模型变得可行。

DESCRIPTION (provided by applicant): The disruption of vascular elastin matrices due to mechanical injury or acquired diseases (e.g., aneurysms), or their congenital absence or malformation can severely compromise vessel elasticity and elastin-mediated cell signaling important to morphogenesis, injury response, and inflammation. Regeneration of elastin within de-elasticized vessels and within tissue-engineered constructs is limited by the poor elastin output by adult vascular smooth muscle cells (VSMCs). Also, cell scaffold materials that can upregulate elastin synthesis and provide biologic cues necessary to regenerating faithful mimics of native elastin matrices are yet to be identified. Previously we determined that that highly biocompatible, crosslinked scaffolds (hylans) containing hyalruonan (HA), a glycosaminoglycan in the ECM, stimulates cultured VSMCs to synthesize close mimics of native elastin. Evidence suggests that HA fragments, not native long-chain HA elicit these elastogenic responses, although long-chain HA possibly facilitates matrix stabilization. The further fabrication of elastogenic scaffolds containing a mixture of long-chain HA and HA fragments, is however contingent on elucidating (i) the differential effects of surface tethered HA or/ and HA fragments on elastogenesis by adult VSMCs, important to optimizing scaffold composition, (ii) the potential benefits of cell culture within prealigned 3D HA scaffolds than on 2D surfaces, to mimicking vascular elastin ultrastructure, and (iii) the impact of concurrently provided mechanical and biochemical cues on the quality (amount, structure, durability, and mechanics) of the synthesized elastin matrix. AIMS: The objective of this project is to clarify the impact of the above parameters on elastin synthesis in an adult rat VSMC culture model. Aim 1 will optimize the HA-fragment size distribution within surface-tethered mixtures of HA based on their individual effects on elastin synthesis by adult RVSMCs. Aim 2 will investigate the standalone and combined benefits of tubular, electrospun HA nanofiber scaffolds containing the optimized composition determined in Aim 1, cyclic mechanical strain, & growth factor (TGF-b, IGF-1) supplements to matrix quality. SIGNIFICANCE: The study outcomes will create a tool that can be integrated with existing vascular devices to fabricate faithful mimics of native elastin on demand. Such elastin mimics will be useful to augument and repair elastin in degenerated vessels, and also make available an in vitro model to study elastogenesis.