Hyaluronan Scaffold for Regenerating Elastin Matrices

描述（由申请人提供）：由于机械损伤或获得性疾病（如动脉瘤）或其先天性缺失或畸形导致血管弹性蛋白基质的破坏，可严重损害血管弹性和弹性蛋白介导的细胞信号传导，这对形态发生、损伤反应和炎症很重要。弹性蛋白在去弹性血管和组织工程结构中的再生受到成人血管平滑肌细胞（VSMCs）弹性蛋白输出不足的限制。此外，细胞支架材料可以上调弹性蛋白的合成，并为再生天然弹性蛋白基质的忠实模拟物提供必要的生物线索，但尚未确定。先前我们发现，含有透明质酸（HA） （ECM中的一种糖胺聚糖）的高度生物相容性交联支架（hylan）刺激培养的VSMCs合成天然弹性蛋白的接近模拟物。有证据表明，尽管长链HA可能有助于基质稳定，但HA片段，而不是天然的长链HA引发了这些弹性反应。然而，含有长链透明质酸和透明质酸片段混合物的弹性支架的进一步制造取决于阐明(i)表面系住的透明质酸或/和透明质酸片段对成人VSMCs弹性形成的不同影响，这对优化支架成分很重要；（ii）在预排列的3D透明质酸支架内培养细胞比在2D表面培养细胞对模拟血管弹性蛋白超微结构的潜在益处。（iii）同时提供的机械和生化线索对合成弹性蛋白基质质量（数量、结构、耐久性和力学）的影响。目的：本项目旨在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将基于HA对成人RVSMCs合成弹性蛋白的个体影响，优化HA片段大小在HA表面捆绑混合物中的分布。Aim 2将研究含有Aim 1中确定的优化成分、循环机械应变和生长因子（TGF-b, IGF-1）补充剂的管状、电纺丝HA纳米纤维支架的单独和综合效益。意义：研究结果将创造一种工具，可以与现有的血管装置集成，根据需要制造天然弹性蛋白的忠实模拟物。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，也为研究弹性发生提供了一种体外模型。

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.