Engineering biomimetic corneal constructs

描述（由申请人提供）：在美国，每年有超过45,000例角膜移植手术。该手术的成功率相当高（两年后达到90%），尽管目前获得供体组织是足够的，但组织质量差异很大，这影响了手术结果。此外，LASIK手术的普及使角膜不适合移植，在不久的将来可能会减少角膜供体的可用性。近年来，通过组织工程制造角膜等效物的尝试值得称赞。这些结构证明了类似于上皮细胞、角质细胞和内皮细胞的三层细胞可以培养成胶原基质的概念。然而，这种结构只遇到了有限的成功，因为基质基质，它提供了角膜独特的（和至关重要的）机械和光学特性，还没有被复制。随机导向的胶原蛋白凝胶是组织工程角膜的典型起点，但对于临床应用来说，它可能不够牢固或不够清晰。此外，预计在体内有显著的效果

DESCRIPTION (provided by applicant): Every year in the United States, over 45,000 corneal transplants are performed. The success rate for this procedure is fairly high (90% after two years) and although current access to donor tissue is adequate, the quality of tissue varies significantly which influences surgical outcomes. In addition, the proliferation of LASIK procedures, which disqualifies a cornea for transplantation, threatens to reduce the availability of donor corneas in the near future. In recent years, laudable attempts have been made to produce corneal equivalents by tissue engineering. These constructs have proven the concept that three layers of cells, resembling the epithelium, keratocytes and endothelium may be cultured into a collagen matrix. However, such constructs have only met with limited success because the stromal matrix, which provides the cornea with its unique (and critically important) mechanical and optical properties, has not been reproduced. Randomly oriented collagen gels, which represent the typical starting point for tissue-engineered corneas, are not likely to be strong enough or clear enough for clinical use. In addition, expecting a significant in vivo remodeling response to integrate a partially functioning artificial cornea is not acceptable. The artificial construct should be functional at the time of implantation. For these reasons, we propose a stromal-centric approach toward the generation of an artificial cornea. By combining engineering, biology, biomechanics and biochemistry, two different routes, both designed to produce biomimetic stromal lamellae (the building blocks for an artificial cornea) will be attempted. The first method employs microfluidics to influence the self-assembly of thin, aligned lamellar sheets comprising collagen and proteoglycans. This approach is acellular and is designed to produce a biomimetic stromal scaffold de novo. The second approach utilizes mechanical cues such as contact guidance (provided by the de novo lamellae) and strain to "direct" the synthesis of matrix by ascorbic acid stimulated human corneal fibroblasts. An extensive characterization of the synthetic response to the imposed stimuli will be conducted to gain a fundamental understanding of synthesis, remodeling and homoeostasis of highly structured matrix. Completion of both parts of this proposal will provide insight critical to achieving our ultimate goal, which is the ex vivo generation of a functional, biomimetic artificial cornea from natural components.