聚多巴胺辅助羟基磷灰石纳米涂层复合骨髓间充质干细胞实现人工角膜生物整合的研究

The cornea is the first line of defense for the optical system. Any material that replaces it must integrate with the surrounding tissue such that the interface provides a bacteria tight seal and a mechanically stable attachment. This is where most attempts to replace corneal function have failed. PMMA has remained a popular material of choice in KPro because of its excellent optical properties and biocompatibility, although its biointegration in cornea tissue is known to be poor. Here, polydopamine coating, which has demonstrated excellent adhesive properties, and subsequent modification with Hydroxyapatite (HAp) was applied to the PMMA surface with the prospect that it may enhance the integration of the material with the corneal tissue. To assess the feasibility of polydopamine coating in enhancing the integration with the corneal tissue, previous in vitro studies were performed using both corneal epithelial cells and the keratocytes. Polydopamine coating and subsequent modification with HAp significantly enhanced cellular proliferation of both cell types. Because of the host cornea's insufficiency and weekly proliferative capacity, autologous bone mesenchymal stem cells (BMSCs) will be combined with HAp coated PMMA surface. General cornea response to the polydopamine-HAp coating or/and combined with BMSCs in vivo will be observed by implanting the surface-treated PMMA keratoprosthesis in the rabbit's eye. ECM including the collagen, MMPs and GAGs will be investigated by western blot, immunohistochemistry, RT-PCR and flow cytometry cell staining in order to understand the surrounding tissue of the graft biological behavior and function. For deeply understand the biomechanics behavior of the keratoprosthesis in vivo, the project intends to innovative use of optical coherence tomography (OCT) based air jet indentation technology to construct the in vivo device to measure the elasticity of the cornea and the movement of the cornea tissue around KPro stem after keratoprosthesis implanted surgery. Considering the fact that polydopamine film can form on various materials, the results herein point to a possibility that polydopamine coating may improve the biointegration of a range of prosthetic devices.

各种原因导致的严重眼表损伤及角膜盲，传统角膜移植成功率很低,合成的硬质人工角膜是这类患者脱盲的希望。目前影响人工角膜发展的主要问题是其和周围组织不能生物整合，可造成严重并发症。本课题拟采用HAp纳米涂层对人工角膜材料表面改性，增加角膜细胞和细胞外基质在杂化界面的粘附、增殖，并减轻炎症反应。并拟使用BMSCs体外诱导软骨细胞，复合在涂层表面，观察角膜细胞因子和细胞外基质与干细胞的相互作用；通过动物实验动态观察植入HAp涂层人工角膜以及复合BMSCs后与周围组织的安全性和有效性；探讨其植入的可行性；通过检测细胞外基质及金属蛋白酶的表达，观察植入术后角膜周围微环境的变化；应用基于OCT气冲印压系统，比较涂层及BMSCs复合后角膜生物力学改变，以及力学刺激对人工角膜周围增殖组织蛋白的表达与调控，评价其在体效果，为解决人工角膜长期稳定性的问题提供实验基础，促进人工角膜技术的发展。

角膜病是仅次于白内障的主要致盲性眼病。据WHO统计，全球角膜盲患者已近4000万。尽管角膜移植是目前角膜盲的主要复明手段，但对于伴有多量新生血管的患者成功率不足50%，且对于严重眼外伤、角膜和眼表损害等引起的双眼严重角膜瘢痕、血管化等均无法通过角膜移植手术恢复视力。合成的硬质人工角膜是这类患者脱盲的希望。如何使人工角膜镜柱和周围角膜组织形成牢固的生物结合，减少溶解的发生，是目前所有合成人工角膜共同面临的难题。因此，本项目围绕人工角膜与周围组织不能生物整合而影响人工角膜发展的主要问题，采用纳米涂层及其联合骨髓间充质干细胞(BMSCs) 技术在人工角膜表面使用聚多巴胺辅助羟基磷灰石（HAp）纳米涂层，利用其吸引生物分子电荷的特点来改善界面，并进行了安全性、有效性、复合BMSCs的可行性及其生物力学性质等系列实验研究。.研究结果显示，聚多巴胺辅助羟基磷灰石涂层（HAp）纳米涂层可成功对钛合金人工角膜进行了表面及内部间隙的改性；确认了气浮沉积（aerosol deposition，AD）技术进行HAp涂层是对人工角膜表面改性的最稳定技术和材料；明确其可增加角膜细胞和细胞外基质在杂化界面的粘附、增殖，并减轻炎症反应；该支架能够满足作为BMSC生长及增生的支架，BMSCs在HAp涂层后的人工角膜材料表面均可诱导成骨及软骨，具有良好的安全性和组织相容性。发表论文12篇，其中SCI收录4篇；申请国家专利1项，实用新型专利2项，获得实用新型授权1项；研究成果可为人工角膜的发展和应用提供理论及实验基础，且各项专利成果具有产业化前景。