【作者】 李林；

【导师】 杨安怀；

【作者基本信息】 武汉大学 ， 眼科学， 2005， 硕士

【摘要】 目的:视网膜下膜(subretinal membranes,SRM)是视网膜外表面的增生性膜,是增生性玻璃体视网膜病变(proliferative vitreoretinopathy,PVR)的一部分,但它的病理过程及形成机制还不清楚。近年来人们发现在复杂视网膜脱离病例中视网膜前膜和视网膜下膜的发生率很高,而且它们的存在与视网膜脱离的术后复发有很重要的关系。近年研究结果提示,PVR与眼内细胞增生有关,有多种免疫因子参与,包括免疫细胞、细胞因子及细胞外基质,它们在PVR的发生、发展中起重要作用。近来有作者报道在PVR患者的玻璃体腔、视网膜前膜、视网膜下液中ICAM-1、MMP-2均呈高表达,提示两者在PVR的发生、发展中起一定作用,有关视网膜前膜在这方面的研究国内外已有较多报道,但有关视网膜下膜的报道尚不多见,本试验主要观察增生性玻璃体视网膜病变的视网膜下膜中粘附分子(ICAM-1)及基质金属蛋白酶(MMP-2)的表达,以探讨ICAM-1、MMP-2与增殖性玻璃体视网膜病变形成和发展的关系,为临床上预防和治疗增生性玻璃体视网膜病变提供理论依据。 方法:用免疫组织化学方法对15例复杂视网膜脱离行玻璃体切割术时剥离的视网膜下膜进行观察。采用常规睫状体平坦部巩膜三通道切口,先切除玻璃体,存在视网膜前膜应作视网膜前膜剥除再处理下膜,手术过程中取出的视网膜下膜组织立即放入4%多聚甲醛固定液中,4℃冰箱冷藏备用。15份SRM标本均按常规制作石蜡切片。常规清洁载皮片,将制成的5μm厚的切片贴附于用3-丙氨基三乙氧基硅烷(3-aminopropyl-triethoxysilane,APES)胶涂片的玻片上,37℃温室放置24h,在二甲苯及梯度酒精内依次脱蜡,用PBS洗3次,作免疫组织化学染色。所有切片先作HE染色,取光镜下结构完整的膜再切片进行免疫组织化学染色。采见辣根酶标记的链霉卵白素试剂盒(博士德生物工程有限公司)染色(SP法):单克隆鼠抗人ICAM-1和单抗隆羊抗人MMP-2抗体用抗体稀释液稀释至1:25和1:75滴于标本上。4℃冰箱湿房过夜,37℃恒温箱复温1h;用PBS洗3次;抗体稀释的生物素标记二抗,滴度1:150,滴于标本上。37℃恒温箱放置30min,用PBS洗3次后用1:150滴度的辣根酶标记的链酶卵白素滴于标本上,37℃恒温箱放置30min,用PBS洗3次,用1:50的3,3-二氨基联苯胺(3,3-diaminbzidine,DAB)滴于标本上,室温下放置15min,显微镜下终止显色,充分冲洗,用苏木素复染,脱水,透明,中性树胶封片。特异性第一抗体用PBS代替并平行染色作为阴性对照。更多还原

【Abstract】 Objective: The subretinal membrane was cell proliferative membrane of retinal outward apparence, subdivision of the proliferative vitreoretinopathy. Recently, people discovered the incidence of the subretinal membrane was very high in the complex retinal detachment cases;the recurrence after operation had relation with the epiretinal membrane and subretinal membrane. Several immune-molecular including immune-cell,cytokines and extracellular matrix are often observed in vitreous and retinal membranes from eyes with PVR.There are many reports about epiretinal membrane,but less about subretinal membrane,so we investigate the expression of intercellular adhesion molecules-1 (ICAM-1) and matrix metalloproteinase-2 (MMP-2) in subretinal membranes of eyes with proliferative vitreoretinopathy.Methods: Fifteen subretinal membranes from retinal detachment complicated with PVR and observed by immunohistochemical examination.Undergoing pars plana vitrectomy by PVR were studied for retinal detachment complicated,the subretinal membranes were immediately thrown into phosptat buffe containing 4% citromint and cryotubes for storing at 4℃ until use.Results: Positive expression of ICAM-1 and MMP-2 located in the membranes of some cells mainly,the others located in the cellular matrix.Expressions of ICAM-1 and MMP-2 were observed in 10 and 8 membranes respectively. Expression of both in 7 membranes.Conclusion: The subretinal membranes in complex retinal detachment were composed of pigment epithelial cells,glial cells,fibroblast and matrix.Glial cell and pigment epithelial cell had conversion and proliferation.They synthesized new collagen fiber. ICAM-1 and MMP-2 are expressed in subretinal membranes of eyes with PVR. These results indicate that adhesion molecules and matrix metalloproteinase might be involved in the development of PVR.更多还原