基于球模多孔基质和干细胞来源内皮的3D打印人工角膜构建策略及机制研究

Corneal transplantation is the most effective method currently to treat corneal disease, but it is confronted with the difficulty of the extreme shortage of donor corneas. Artificial cornea as a substitute for human cornea is expected to solve this dilemma, but there are still two major bottlenecks limiting its development: 1. Lack of personalization – unable to meet the diversity treatment needs of patients with corneal diseases. 2. Immaturity of functional endothelium cell reconstruction. The applicants have taken the lead in the preparation of artificial cornea with the combination of sphere-templated porous scaffold and dual network polymer technology, which is free of immune rejection. We also got 3D technology platform to convert the artificial cornea into 3D printing model, and make it possible to be personalized. At the same time, in order to reconstruct corneal endothelial layer, we used stem cells to transform to endothelial like cells in vitro, then plan to use cell surface engineering technology to induce directional modification and in vivo transformation. Through 3D printing, corneal data acquisition, calculation and model transformation, this study aims to explore the rules of endothelial cell differentiation, and influence on the mechanism of artificial cornea biological properties, thus provide scientific basis for the realization of personalized and bionic artificial cornea. This study also aims at exploring the method of inducing the transformation of corneal endothelium cells by stem cells in situ, to illustrate the regulation and related mechanisms by aqueous vasoactive intestinal peptide cytokines and intraocular micro environment. This study will be initial of 3D printing preparation for individualized artificial cornea production, the first to clarify its mechanism and provide the feasibility. Also provide the basis to clarify the mechanism of stem cells in situ inducing functional endothelial cells formation.

角膜移植为治疗角膜病的最佳方式，但面临人供体角膜匮乏的困境。人工角膜作为替代物可望解决该问题，但尚有两大瓶颈亟待突破：一、缺乏个性化而无法满足患者多样性治疗需求；二、难以重建功能化角膜内皮。申请人以球模多孔支架与双网络聚合物技术制备人工角膜，并获抗免疫排斥功能的3D打印模型，为个性化制造奠定基础；经体外诱导干细胞为角膜内皮样细胞，并完成细胞表面原位细胞因子修饰。本项目通过研究3D打印过程角膜数据采集、运算和转模，探索其对内皮细胞分化作用规律和对人工角膜生物性能影响机制，为实现人工角膜个性化和仿生化制备提供科学依据；同时探索干细胞体内原位诱导制备功能完善角膜内皮的方法，力图阐明房水中血管活性肠肽等细胞因子及眼内微环境对该过程的调控规律与相关机制。本项目将开创3D打印制备个性化人工角膜的先河，率先阐明其机制、明确构建策略可行性，并为干细胞原位诱导功能化内皮机制阐明提供转化医学研究学术依据。

角膜移植为治疗角膜病的最佳方式，但面临人供体角膜匮乏的困境。人工角膜作为替代物可望解决该问题，但尚有两大瓶颈亟待突破：一、缺乏适宜地人工角膜去满足患者多样性治疗需求；二、难以重建功能化角膜。在之前的研究基础上，申请人拟逐步实现人工角膜构建和干细胞应用于角膜内皮和基质再生的技术。本项目重点聚焦于人工角膜相关构建地关键技术、以及适用于角膜再生修复地目标干细胞这个两个目标。发展了一种新型的基于角膜伤口修复的原位层层自组装技术，并在大鼠角膜上做了动物验证实验，显示了对角膜伤口的明显加速愈合作用。同时，本实验从自体眼睑提取脂肪干细胞，通过血液注射的方式，观察其在角膜伤口的定位和聚集，以及对角膜基质伤口的愈合的促进作用；显示了眼睑来源的脂肪干细胞下一步作为目标干细胞的可能性，阐明干细胞促进角膜基质再生修复的效应和作用机制。最终为进一步人工角膜地构建提供依据、并为干细胞原位诱导功能化内皮机制阐明提供转化医学研究学术依据。