飞秒激光角膜凹透镜光学效应及创伤修复机制研究

Femtosecond laser small incision lenticule extraction (SMILE) is safe, mini-invasive for myopia correction (convex lenticule excision). However, the use of SMILE for the correction of hyperopia has not been reported yet. This research is designed to explore a mini-invasive way of treatment for hyperopia by changing laser scan mode to make corneal concave lenticule and to investigate its refractive effects and wound healing mechanism. By applying 3-dimensional scanning mode of femtosecond laser on rabbits’ corneas, the corneal concave lenticule in different diopters will be made and extracted as well as be used for allotransplantation or autotransplantation. Corneal refractive parameters and regularity will be measured using corneal topography postoperatively, corneal biomechanical parameters will be tested using Ocular Response Analysis (ORA) and Corvis ST, both the post-operative process of corneal nerve wound healing and corneal morphological changes will be assessed using an in vivo confocal microscopy. Light Microscopy and Transmission Electron Microscope will be applied to detect the changes in corneal ultrastructure. Cell death will be analyzed using TdT-mediated dUTP Nick-End Labeling (TUNEL). Various cytokines (TGF-β, IL-1,etc) changes will be evaluated using immunohistochemical techniques. The corneal fluorescein staining and tear film break up time will be taken; the neuro-nutrientsubstance will be detected, changes of structure between corneal layers will be observed using myelin staining with Gold Chloride, all those information and relationship with neurosensory restoring will be investigated. The outcomes of this project will provide theoretical basis to innovative corneal refractive surgery for the correction of hyperopia. Besides, it will convert the traditional “subtraction” principle of corneal refractive surgeries (to correct refractive errors by cutting or ablation corneal tissue) to a novel “addition” one (to correct refractive errors by thickening and strengthening corneal tissue) by using the concave lenticule with transplantation. It will provide a new, mini-invasive way of treatment for myopia correction in thin corneal and keratoconus eyes and be beneficial to the broad masses of the ametropic patients.

飞秒激光小切口透镜取出术（凸透镜切除）矫正近视安全、微创，但远视矫正尚无报告。本研究改变激光扫描模式制作角膜凹透镜以实现远视矫正的微创模式，探索其光学效应和创伤修复机制。通过对兔角膜进行三维扫描制作不同屈光度的凹透镜切除及自体或异体移植，用角膜地形图测量屈光力及形态规则性；ORA、Corvis ST测量角膜生物力学变化；活体共聚焦显微镜观察角膜组织结构改变、神经修复规律；光镜和透射电镜观察角膜超微结构变化；TUNEL法检测细胞凋亡，免疫组化技术检测 TGF-β、IL-1等细胞因子变化；角膜荧光素染色和泪膜破裂时间、神经营养物质检测和氯化金染色观察角膜层间结构变化与神经知觉修复的关系。本研究可为推进微创的远视矫正角膜屈光手术提供理论依据，并将取出的凹透镜移植再利用，将角膜激光手术由削薄变弱的“减法”转为加厚变强的“加法”，为薄角膜或圆锥角膜患者提供新的治疗途径，使更多屈光不正患者受益。

本项目围绕飞秒激光角膜基质凹透镜制作、取出和移植的可行性、安全性、有效性研究展开，先在动物模型眼上成功构建手术模型，探讨该技术对角膜结构生物力学、光学效应以及创伤愈合反应的影响，并转化初步应用于临床。主要研究内容和重要结果： 1）评估飞秒激光角膜凹透镜显微结构及表面形态特征，结果显示远视矫正制作凹透镜表面质量良好，可见组织桥和凹槽及腔面结构；2）成功建立飞秒激光小切口基质透镜取出和异体移植的动物模型，结果显示其具有可行性、安全性和有效性，透镜移植可有效增加角膜基质厚度，术后愈合过程中呈现从中重度反应到轻度反应变化特点，在术后半年内达稳定，术后2年屈光度呈现低度回退趋势；飞秒激光凹透镜异体移植安全、可行，透镜与周围组织具有较好的融合度，不同过渡区对手术后创伤愈合无显著影响； 3）评估飞秒激光角膜基质凹透镜制作及取出后治疗远视患者的临床效果及视觉质量，结果显示矫正远视安全、有效，术后眼内散射变化少，术后彗差及球差改变可能会影响早期的视觉效果。同时进一步拓展，将透镜移植成功应用于有疾患的人眼，治疗浅层角膜病变、圆锥角膜、角膜激光术后并发症。项目团队共发表SCI文章16篇（基金标注），参加国内相关领域眼科年会做发言报告10余人次，参加国际会议作大会发言及壁报交流5人次，培养研究生3名，学习班讲课30余次，累计培训400余名医生。研究成果获得上海市科技进步奖二等奖等，并在临床转化应用。本项目研究成果可为推进微创的远视矫正角膜屈光手术提供理论依据，并将取出的凹透镜移植再利用，将角膜激光手术由削薄变弱的“减法”转为加厚变强的“加法”，为薄角膜圆锥角膜患者、角膜营养不良等患者提供新的治疗途径，使更多屈光不正及角膜病患者受益。