A Cell-free Approach to the Engineering of Corneal Stroma

角膜基质工程的无细胞方法

• 批准号: 10222698
• 负责人: Nima Saeidi
• 金额: $ 41.62万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-30 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10222698
• 关键词: Address; Allografting; Biological; Biomimetics; Biophysics; Blindness; Caliber; Cell Culture System; Cells; Characteristics; Cicatrix; Clinical; Clinical Engineering; Collagen; Collagen Fibril; Collagen Type I; Complex; Cornea; Corneal Stroma; Crowding; Development; Endothelium; Engineering; Epithelial; Exhibits; Extracellular Matrix; Eye; Foundations; Generations; Goals; Grain; Human; In Vitro; Keratoplasty; Liquid substance; Maps; Mechanics; Modeling; Molecular; Morphology; Nature; Optics; Oryctolagus cuniculus; Outcome; Patients; Production; Property; Proteoglycan; Research; Stromal Cells; Structure; System; Techniques; Testing; Tissue Engineering; Tissues; Transplantation; Weight-Bearing state; Work; analog; base; condensed matter physics; corneal scar; crystallinity; decorin; flexibility; gain of function; immunoreaction; improved; in vivo; lumican; mechanical properties; mimetics; novel; predictive modeling; self assembly; self organization; sight restoration; simulation; stem cell fate; stem cells; success; theories; wound; wound healing

ABSTRACT. Over 25 million people worldwide suffer from corneal blindness in one or both eyes. Corneal transplantation is the only option available to restore vision. However, there is a worldwide donor shortage, and fewer than 1% of patients with corneal blindness receive a transplant each year. Despite the great promise of corneal tissue engineering research to fill this gap, efforts to develop a clinically viable corneal substitute are hindered primarily due to the inability of the existing in vitro culture systems to reproduce the intricate extracellular matrix structure (ECM) of the stroma. Approximately 95% of the stroma is composed of type I collagen-based ECM in which regularly packed collagen fibrils have a uniform diameter and are arranged as orthogonal lamellae, providing the cornea with its unique mechanical and optical properties. The predominantly acellular nature of the stroma has motivated us to pursue a cell-free approach to the engineering of the stroma. We have recently demonstrated that the planar confinement of crowded collagen molecules induces self-organization of the collagen into highly ordered structures. Our objective here is to engineer an acellular stromal analog. Our central hypothesis is that a fully functional corneal stroma can be developed by harnessing the inherent physicochemical properties of collagen molecules. Our efforts to pursue the goals of this proposal will be pursued in three specific aims: In Aim 1, the effects of confining and crowding conditions on the long and short-range organization of collagen fibrils will be determined. Next, the impact of lumican and decorin proteoglycans on collagen ultrastructure (i.e. diameter and spacing) will be elucidated. In Aim 2, we will use a combination of theoretical and numerical modeling to provide an understanding of how crowding and confining conditions, as well as interactions with proteoglycans, impact collagen organization, morphology, and self-assembly. The predictive nature of the model would also enable identifying additional experimental parameters to further optimize the stromal-mimetic collagenous structures. In Aim 3, the long-term in vivo function of the acellular stromal analogs will be delineated by transplanting them into rabbits with deep corneal scars. In addition, in a set of exploratory studies we will investigate whether the native-like physical characteristics of the constructs (e.g., fibrillary organization, diameter and mechanical properties), would improve the differentiation of corneal stromal stem cells into native corneal stromal keratocytes. If successful, the work described here is expected to result in the development of first generation acellular corneal stromal analogs which can restore corneal function upon transplantation. The acellular stroma could be used directly for lamellar transplant in vivo or they could be integrated with the epithelial and endothelial layers to produce a fully functional cornea. Furthermore, the versatile collagen organizing technique that we propose to develop could be used for the production of biomimetic substrates to be integrated into the existing in vitro platforms to mechanistically investigate how various properties of ECM (e.g., organization and diameter) impacts cellular fate and function.

抽象的。全世界有超过2500万人患有单眼或双眼的角膜失明。角膜 移植是恢复视力的唯一选择。然而，全世界的捐赠者短缺，而且 每年只有不到1%的角膜失明患者接受移植手术。尽管有很大的希望 角膜组织工程研究为了填补这一空白，开发临床可行的角膜替代物的努力是 主要是由于现有的体外培养系统无法复制错综复杂的细胞外 基质的基质结构(ECM)。大约95%的间质由I型胶原组成 其中规则堆积的胶原纤维具有均匀的直径并排列成正交片的ECM， 为角膜提供其独特的机械和光学特性。主要是脱细胞的性质 基质促使我们追求一种无细胞的方法来构建基质。我们最近做了 证明了拥挤的胶原分子的平面限制诱导了细胞的自组织 胶原蛋白形成高度有序的结构。我们的目标是设计一种脱细胞基质类似物。我们的中央 假说是，通过利用固有的物理化学物质可以形成功能齐全的角膜基质。 胶原蛋白分子的性质。我们为实现这项提案的目标所作的努力将在三个具体方面进行 目的：在目标1中，限制和拥挤条件对长程和短程组织的影响 胶原蛋白纤维将被测定。接下来，鲁米肯和核心蛋白多糖对胶原蛋白的影响 超微结构(即直径和间距)将被阐明。在目标2中，我们将结合使用理论和 数值模拟，以提供对拥挤和限制条件以及 与蛋白多糖的相互作用，影响胶原蛋白的组织、形态和自组装。预测性的 该模型的性质还将使识别额外的实验参数能够进一步优化 基质样胶原结构。在目标3中，脱细胞基质类似物在体内的长期功能 将通过将它们移植到有深角膜疤痕的兔子体内来描绘。此外，在一套探索性的 研究我们将调查结构(例如，纤维状)的天然相似的物理特征 组织、直径和力学性能)，将促进角膜基质干细胞的分化 细胞转化为天然角膜基质角质细胞。如果成功，这里描述的工作预计将导致 第一代可恢复角膜功能的脱细胞角膜基质类似物的研究进展 移植。脱细胞基质可以直接用于体内的板层移植，也可以 与上皮层和内皮层整合，形成功能齐全的角膜。此外， 我们建议开发的多功能胶原组织技术可以用于生产 将仿生底物集成到现有的体外平台中，以机械地研究如何 细胞外基质的各种性质(如组织和直径)影响细胞的命运和功能。