2010-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/648771摘要：眼內新生血管形成可出現於多種視網膜疾病，引起視力的嚴重受損，其中以糖尿病視網膜病變於先進國家中為引起中老年人視力受損的主因。目前的研究得知眼內新生血管形成和多種生長因子有密切的關係，最近國外的研究顯示出肝細胞生長因子(hepatocyte growth factor,簡稱HGF)和眼內新生血管形成有關。目前已知和眼內新生血管形成相關的生長因子尚包括血管內皮細胞生長因子(vascularendothelial growth factor, 簡稱 VEGF)，鹽基性芽細胞生長因子(basic fibroblast growthfactor,簡稱 bFGF )，類胰島素生長因子(insulin-like growth factor, 簡稱 IGF )和多種其他的生長因子。本研究主要目的為建立新的眼內新生血管形成的動物模式，使用生長因子HGF 注射於兔眼玻璃体腔中造成眼內新生血管，於此動物模式建立完成後，探討不同藥物和光凝固作用對其抑制作用之評估，可做為日後臨床上應用的根據。同時探討視網膜雷射光凝固作用對於兔眼眼內HGF 含量的影響。本研究動物實驗方面使用的動物為成熟有色兔子，体重為2.5 至3.5 公斤，於Zoletil和Rompam 全身麻醉後進行實驗。眼內新生血管動物模式的建立為將不同劑量的HGF注射於兔眼視神經盤前方的玻璃体腔之中，而產生眼內新生血管的形成。所有兔子在實驗組於兔子左眼注射生長因子，而對照組則於兔子右眼注射緩衝溶液。所有兔子皆於注射生長因子之前，注射後第1 週、第2 週、第3 週、第4 週和第6週做眼底照相和螢光眼底血管攝影檢查，觀察眼內新生血管的形成過程。兔子在適當的時段於麻醉作用後，做眼球摘除手術，將眼球置於固定液之中，而後以角鞏膜剪刀去除眼球前部和玻璃体，再仔細取下必要的部位，利用光學顯微鏡和免疫化學組織檢查法以觀察新生血管形成的情況。對於眼內新生血管抑制作用的研究則利用生長因子HGF 抗体做拮抗作用，使用thalidomide 注射於玻璃体中，同時也探討雷射光凝固作用對於眼內新生血管的抑制情形，同時分析雷射光凝固作用對於眼內HGF 含量的影響。這些研究工作皆為先進的研究，目前國外尚無類似的成果被發表。由於視網膜色素上皮細胞(retinal pigment epithelial cells, 簡稱 RPE 細胞)可分泌多種生長因子且對於眼內新生血管的成長過程扮演相當要的角色，本研究同時將探討HGF對於其它cytokines 表現產生的影響和thalidomide 對於RPE 細胞的作用情形，同時分析雷射光凝固作用對於RPE 細胞產生HGF 的影響。<br> Abstract: Intraocular neovascularization occurs in several retinal diseases, the mostcommon of which is diabetic retinopathy, a major cause of new blindness indeveloped nations. Occlusion of retinal vessels leading to retinal ischemia is a featureshared by diseases in which intraocular neovascularization occurs. This has led to thehypothesis that the development of intraocular neovascularization is stimulated byone or more angiogenesis factors released by ischemic retina.Recent studies demonstrated that hepatocyte growth factor (HGF) is a potentischemia-induced angiogenic factor that acts during ocular angiogenesis. The recentstudy demonstrated that sustained intravitreal release of VEGF at the retinal surfacecan induce retinal neovascularization in rabbits. In addition toVEGF, other growthfactors including bFGF and IGF were reported to be related to the development ofintraocular neovascularization.The main purposes of this study are to establish a new animal model ofintraocular neovascularization , using the methods of injecting HGF into the vitreouscavity of the rabbits. After the establishment of this animal model, we will usedifferent agents and laser photocoagulation to inhibit this model of intraocularneovascularization and evaluate the effects.The pigmented rabbits (2.5-3.5 kgs) are used in this study. A sterile solutioncontaining HGF is introduced into the vitreous cavity through a 27-gauge needleunder direct visualization. The reagent is injected in the vitreous cavity just above theoptic disc. Fundus color photographs and fluorescein angiography are taken weeklywith a fundus camera. After an adequate interval the animals are expired and the eyeballs are removed. The areas of intraocular neovascularization are then sectioned andprocessed for light microscopy and immunohistochemistry for staining factor VIIIand CD31 to evaluate the new vessel formation.The inhibition of intraocular neovascularization by intravitreal injection ofthalidomide, antibodies to HGF, or scatter laser photocoagulation is also be evaluated.Intraocular levels of HGF after retinal laser photocoagulation will also beinvestigated.Bovine RPE cell culture will also be performed and the effects of HGF on theproduction of other cytokines and apoptosis of RPE cells will be investigated. Theeffects of HGF and thalidomide on the migration and proliferation of RPE cells areevaluated. The effect of laser photocoagulation on the production of HGF by RPEcells will also be studied.