行政院國家科學委員會補助專題研究計畫成果報告:色素上皮細胞與感光細胞於高度近視網膜變性時凋亡現象之偵測與其基底質調整治療(2/2)

High myopic maculopathy are one of the major causes of blindness. In the past several years retinal pigment epithelium transplantation has become the major direction of researches in the treatment of macular degeneration. Our previous studies demonstrated that photoreceptor-retinal pigment epithelium-choriocapillaris complex became ruptured in the elastic membrane of Bruch’ s membrane in form-deprivation myopic chicks. The staining of extracellular matrix has also decreased. This is similar to the lacquer crack formation in Bruch’s membrane in high myopia in human. These are the pathologic changes found in the substrates attached by retinal pigment epithelium. Accordingly, the successfulness of retinal pigment epithelium depends on the remodeling of the substrates. The gelatin-matrix transfer technique we developed is able to transfer extracellular matrix with or without cells. Our study demonstrates that immunolabeling of the exoplants after enzymatic treatments showing decreased finonectin, laminin, vitronectin and collagen IV staining.Transferred extracellular matrix was able to increase the efficiency of retinal pigment epithelial cells to substrates although they were previously treated by trypsin, dispase or collagenase. Our results suggest that the modification of substrates can be achieved by gelatin matrix technique transfer of extracellular matrix. In addition we found that DBA/2J mouse developed myopic changes with myopic fundus tessellation. In the present study, the retinal pigment epithelium and photoreceptors developed apoptosis, and combined RPE-extracellular matrix trasplantation significantly protected photoreceptor cell from apoptotic cell death in DBA/2J mouse and form-deprivation myopia chicks

行政院國家科學委員會補助專題研究計畫成果報告:感光細胞-色素上皮細胞-脈絡膜複合體之退化與高度近視黃斑部變性之關係 (3/3)

高度近視黃斑部變性會導致失明，同時也是國人視力障礙的主因。然而針對這 個高度近視最嚴重的併發症，仍有下列問題有待解決：（1）成黃斑部變性的病 理機轉至今不明。（2）目前沒有一個良好的高度近視網膜退化的動物模式。（3） 迄今仍沒有有效的治療方式。 面對這些問題，我們第一步要以體外實驗方式，配合臨床上的循血綠攝影檢查 的發現，探求造成黃斑部變性的基本原因。最近一、二年對於不同原因的黃斑部 變性的致病機轉有嶄新的突破出現，我們計畫利用細胞生物學的方法，分層的探 討感光細胞層。色素上皮細胞與脈絡膜微細血管層發生病變的原因及其相關性。 第二步將使用遮蔽性近視動物，來研究眼球構造上的變化。我們將本著第一步 研究的發現使用器官體外培養的方法來探討近視眼球的功能上的缺陷，例如：色 素上皮細胞在其基質上附著、生長與繁殖的能力等，期望由此建立第一套的高度 近視黃斑部變性的動物模式。 第三年計畫的目的在完成動物感光細胞，色素上皮細胞移植的活體實驗。最近 幾年，眼球內網膜細胞移植正逐漸受到重視，尤其是對於針對增齡性黃斑部變性 網膜下新生血管夾除後的色素上皮細胞層缺損的課題，有許多可喜的成就產生， 我們初步發展出來的網膜下細微手術方法，可以提供一個有效而直接的治療方 向，配合前兩年的研究成果，可將需要的細胞組織與間質融合到一個易於操作的 戴體上，植入眼球內的病變部位，然後在活體的環境下，可觀察其對高度近視黃 斑部變性感光細胞退化的治療效果，並且對於黃斑變性原因可以得到更進一步的 認識，期望由此可以找到一條通往高度近視黃斑部變性的光明之路。high myopic maculopathy, macular degeneration, retinal pigment epithelium, extracellular matrix, retinal transplantation high myopic maculopathy may cause the loss os central vision and is the leading cause of blindness in taiwan. Unfortunately, there are still several topics need to be resolved in dealing with this most serious complication of high myopia: (1) The pathogenesis of high myopic maculopathy remains unknow. (2) The need for an animal model of high myopic maculopathy exists. (3) Untill now there is no effective treatment for high myopic maculopathy. First of all, we would start by using an in vitro method to study the cause of it. There has been tremendous breakthroug in the study of the pathogenesis of macular degeneration in recent years. We planned to study the defects and relationships of the these cellular layess in the photoreceptors-retinal pigment epithelium-choriocapillais complex. Secondly, we would study the structural changes in the eyeball of form- deprivation myopia. On the basis of the findins the previous steps, we would study the functional defect of the high myopia eyes by an organ culture technique, e.g. the reattachment, growth and proliferation of retinal pigment epithelium on the substrates, The aim is to establish an animal model for high myopic maculopathy. Thirdly, we would transplant retinal pigment epithelium to the subretinal space of high myopic eyes by a microsurgical delivery technique in combination with the necessary factors found in the previous studies, e.g. extracellular matrix and/or growth factors. Our results showed that the rupture of elastic membrane is the first sign of Bruch’s membrane degeneration by 8 weeks, however the reattachment efficiency of RPE to Bruch’s membrane were decreased since 2 weeks after form- deprivation. In the rescue of macular degeration, the combined RPE-Extracellular matrix transplautation technique we developed and published peuiously serue as a novel method in the rescue of photoreceptor cells in large myopic eyes of DBA/2J mice