转基因蚕丝蛋白生物膜及其拓扑形貌在眼表损伤修复中的作用研究

Ocular surface damage and diseases increase year by year and have become important ocular disorders with the rate of blindness second only to cataract. The research of good biocompatible artificial film simulating the structure and function of amniotic membrane provides an efficient way in the treatment of ocular surface diseases. This project is based on advanced concept and technology of Ophthalmology, Molecular biology, Biomaterials and Biophysics. On the basis of silk gland bioreactor constructed by transgenesis technology, the project aims to synthesize a silk fibroin material containing a variety of cell growth factors and adhesion molecules. Moreover, different surface topography of silk fibroin biomembrane will be achieved by using technologies such as electrospinning and template casting. Through the in vitro cell biological and animal acute corneal alkali burn model research, we hope to explore the influence of the surface topography of materials on the biological function of critical cells participating in ocular surface repair, such as corneal epithelial cells and corneal limbal stem cells, and provide certain scientific basis for the development of biomimetic membrane used in ocular surface repair.

眼表损伤和疾病逐年增多，已成为致盲率仅次于白内障的重要致盲性眼病。研究生物学效应优良的仿羊膜人工薄膜材料，是治疗眼表疾病的有效途径。本项目基于眼科学、分子生物学、生物材料学、生物物理学的先进理论和方法，拟采用转基因技术构建家蚕丝腺生物反应器，以期合成一种含多种细胞生长因子、粘附分子的丝素蛋白材料；并借助静电纺丝和模板浇筑等技术，制备具有多种不同拓扑形貌的丝素蛋白生物膜，通过体外细胞生物学和角膜急性碱烧伤模型研究，探索材料表面的拓扑形貌对于角膜上皮细胞、角膜缘干细胞等眼表修复关键细胞的生物学功能的影响，为研制眼表修复用生物仿生膜提供一定的科学基础。

本研究根据预期研究方案和路线，基于眼科学、分子生物学、生物材料学、生物物理学的先进理论和方法，构建了蚕丝中稳定表达hEGF的转基因家蚕新品系，完成了转基因蚕丝的微纳形态、蛋白质二级结构以及相应生物活性的鉴定。在此基础上，制备了不同拓扑形貌的蚕丝蛋白生物膜，明确了生物膜的基本材料表征和表面拓扑形貌对细胞行为的影响。此外，优化设计并研制具有活性氧清除功能的电纺膜-水凝胶（PUTK /RH）复合膜材料，并通过建立大鼠角膜急性碱烧伤模型，进一步评估材料促进角膜修复的能力，探索了仿羊膜生物膜材料用于角膜急性化学烧伤修复的总设计方案。

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