Highly resilient, hydrophilic bioelastomers for engineering vocal fold tissue

声音产生受损对个人健康和福祉，社会和职业功能以及社会生产力具有重大影响;据估计，仅教师的声音问题的社会成本在美国每年就高达25亿美元。然而，用于治疗声带疾病的材料的开发受到声带的严格机械要求的阻碍，这些要求包括在高达1000 Hz的频率下维持变形的能力，以及在高达200%的瞬时拉伸后完全弹回的能力。迄今为止，尽管在声带组织工程材料支架的开发方面进行了广泛的努力，但尚未确定具有所需机械性能的材料。我们提出了一个全面的生物工程方法来解决这个问题。我们将采用基于昆虫蛋白质的新型弹性支架，弹性蛋白是昆虫发声，跳跃和飞行器官的主要能量储存，并在高达4000 Hz的频率下变形后表现出无与伦比的弹性（拉伸后恢复）。我们将采用模块化重组方法来产生节枝弹性蛋白样多肽（RLP），其可以被工程化以携带生物活性结构域而不损害节枝弹性蛋白结构域的机械性质，并且其中可以独立调节这些基质的多种性质，包括机械性质、细胞结合和降解。我们将培养人骨髓间充质干细胞（hMSCs）在这些矩阵在静态和动态条件下，并采用一套振荡流变学，拉伸测试，和高频扭转波的方法来表征细胞封装的结构的机械性能。采用组织学、免疫组化、蛋白质印迹和基因表达技术来证实hMSCs的分化和声带细胞外基质的产生。这些研究将为我们选择用于注射到兔声带中以改善声带瘢痕的细胞/材料构建体提供信息。因此，我们的调查将有助于开发方法来表征和培养材料在高频率，以及产生一类新的材料，可以优化声带组织的再生。我们的方法最终将作为一个通用平台，在机械要求苛刻的再生医学应用的材料设计。

DESCRIPTION (provided by applicant): Impaired voice production holds significant implications for individual health and wellness, social and occupational function, and societal productivity; the societal costs of voice problems in teachers alone have been estimated to be of the order of $2.5 billion annually in the United States. The development of materials for the treatment of vocal fold disorders, however, has been hampered by the stringent mechanical requirements of the vocal fold, which include the ability to both sustain deformation at frequencies as high as 1000Hz, and also completely recoil after transient stretch up to 200%. To date, despite widespread efforts in the development of materials scaffolds for the tissue engineering of the vocal fold, no materials with the required mechanical properties have been identified. We propose a comprehensive bioengineering approach to this problem. We will employ new elastomeric scaffolds based on the insect protein, resilin, which is the primary energy store in the sound-producing, jumping, and flight organs of insects, and demonstrates unmatched resilience (recovery after stretch) after deformation at frequencies up to 4000Hz. We will employ modular recombinant methods to generate resilin- like polypeptides (RLPs) that can be engineered to carry biologically active domains without compromising the mechanical properties of the resilin domain, and in which independent tuning of multiple properties of these matrices, including mechanical properties, cell binding, and degradation, is possible. We will culture human mesenchymal stem cells (hMSCs) in these matrices under both static and dynamic conditions, and employ a suite of oscillatory rheology, tensile testing, and high-frequency torsional-wave methods to characterize the mechanical properties of cell-encapsulated constructs. Histological, immunohistological, western blot, and gene expression techniques will be employed to confirm the differentiation of hMSCs and the production of vocal fold extracellular matrix. These studies will inform our choices of cell/materials constructs for injection into the vocal folds of rabbits to ameliorate vocal fold scarring. Our investigations wil thus contribute to the development of methods to characterize and culture materials at high frequencies, as well as yield a new class of materials that may optimize the regeneration of vocal fold tissue. Our approaches ultimately will be useful as a general platform in the design of materials for mechanically demanding regenerative medicine applications.