作者:黄玲 惠延年 徐磊 王丽丽
关键词: 玻璃体视网膜病,增生性;玻璃体;肝细胞生长因子;酶联免疫吸附测定 摘 要:目的 定量研究肝细胞生长因子(HGF)在增生性玻璃体视网膜病变(PVR)患者玻璃体中的水平. 方法 采用双夹心酶联免疫吸附测定法检测正常对照组10只眼、PVR-A,B组7只眼、PVR-C组13只眼、PVR-D组16只眼. 结果 玻璃体内HGF的含量:正常对照组1.8~7.4μg・L -1 ;PVR-A,B组3.8~12.8μg・L-1 ;PVR-C组7.7~20.1μg・L-1 ;PVR-D组10.3~24.6μg・L-1 .PVR各组玻璃体中HGF的含量比正常对照组显著升高(P<0.01);PVR-C,D组玻璃体中HGF的含量较PVR-A,B组的含量升高(P<0.05or P<0.01);PVR-D组玻璃体中HGF的含量与PVR-C组比较含量也升高(P<0.05),玻璃体中HGF的含量随着PVR的发展而呈上行性升高. 结论 PVR患者玻璃体中HGF含量升高,HGF可能参与了PVR的病理过程. Changes of hepatocyte growth factor levels in vitre┐ous body with proliferative vitreoretinopathy HUANG Ling HUI Yan-Nian XU Lei WANG Li-Li
1 Department of Ophthalmology,Xijing Hospital,Fourth Military Medical University,Xi'an710033,China, 2 Section of Neurobiology Research,Medical School of Xi'an Jiaotong University,Xi'an710061,China, 3 Department of Ophthalmology,Fourth Mu-nicipal Hospital of Xi'an,Xi'an710004,China Keywords:vitreoretinopathy,proliferative;vitreous body;hepatocyte growth factor;enzyme-linked im-munosorbent assay Abstract:AIM To determine the mass concentration of hepatocyte growth factor(HGF)in the vitreous body of pa-tients with proliferative vitreoretinopathy(PVR).METH┐ODS The high sensitive sandwich enzyme-linked im-munosorbent assay(ELISA)was used to measure the level of HGF in vitreous bodies of10normal eyes,7PVR-A,B eyes,PVR-C13eyes,and16PVR-D16eyes.RESULTS The mean value of HGF level in the vitreous body was3.3(range1.8~7.4)μg・L-1 in the normal group;7.4(range3.8~12.8)μg・L-1 in the PVR-A,B group;12.0(range7.7~20.1)μg・L-1 in the PVR-C group,15.6(range10.3~24.6)μg・L-1 in PVR-D group.HGF levels in the PVR groups were significantly higher than the level in the normal group(P<0.01);the PVR-C,D groups showed a higher mean vitreous HGF concentration than that in PVR-A,B group(P<0.05or P<0.01);the level of HGF in the PVR-D group was also higher than that in the PVR-C group(P<0.05).From grade A,B PVR to grade D PVR,a progres-sive increase was observed in the amount of HGF.CONCLU┐SION HGF levels in the vitreous body complicated with PVR were significantly higher than the level in normal group.HGF might be involved in the pathogenesis of PVR. 0 引言 增生性玻璃体视网膜病变(proliferative vitreo-retinopathy,PVR)是重要的致盲性眼病,是裂孔源性视网膜脱离(rhegmatogenous retinal detachment,RRD)手术失败的最主要原因.PVR的主要特点是视网膜色素上皮细胞(retinal pigment epithelial cell,RPE)在视网膜表面和玻璃体腔内形成具有收缩能力的增殖膜,属眼内发生的过度损伤修复反应.肝细胞生长因子(HGF)的研究主要在肝肾脑疾病及肿瘤生物行为等领域;在眼科方面,特别是增生性视网膜疾病等方面未见相关研究.我们以孔源性视网膜脱离PVR为研究对象,通过对PVR患者玻璃体中肝细胞生长因子进行定量分析从而观察其在PVR中的变化规律,从而明确HGF是否参与了PVR的病理过程.
1 对象和方法 1.1 对象 ①患者组:标本来源于我科1999-03/2000-11确诊PVR并进行玻璃体手术的住院患者36例共36只眼,男20例,女26例,年龄(42±6)岁.按照1983年国际视网膜命名委员会的诊断标准进行PVR分类,其中PVR-A,B级7例(含单纯黄斑裂孔性视网膜脱离5例),PVR-C级13例,PVR-D级16例;36只眼在作睫状体扁平部三切口玻璃体显微手术,插入眼内注液管而尚未注液入玻璃体腔时于玻璃体切割前经睫状体平坦部在显微镜直视下先取出原玻璃体液0.2~0.3mL,并移至eppendorf管中,样本取出后1000r・min-1 离心10min,取上清液均置于-70℃冰箱保存,实验前样本置于室温下.②对照组:10只非正常死亡者的眼球均于死后6h内作角膜移植取出角膜植片后,用10mL一次性注射器经睫状体平坦部垂直于眼球壁向心性穿刺入玻璃体腔,抽取正常玻璃体液0.5~1.0mL.样本取出后1000r・min-1 离心10min,取上清液均置于-70℃冰箱保存,实验前样本置于室温下. 1.2 方法 人HGF的ELISA试剂盒为美国R$D公司产品,灵敏度<40ng・L-1 .试剂盒内附有纯的人HGF标准品.对HGF具有特异性的mAb粘附在ELISA检测板的微孔底面,加入样品和标准品后室温下孵育2h,使HGF与mAb结合.洗涤液洗板4次后,每孔加入200μL对HGF有特异性酶标记的多克隆抗体,室温下孵育2h,产生抗体-HGF-抗体复合物.每孔加入200μL过氧化氢和四甲基对二氨基联苯显色液,室温下孵育30min.最后加入50μL终止液.采用酶联免疫测定仪(DG-3022型酶联免疫检测仪为中国华东电子管厂生产)测定样本处在A450nm 波长的条件下的光密度值.用HGF标准品做出一条从0~8000ng・L-1 与值相对应的标准曲线.通过将测得的每个样品的A值与标准曲线相比较,计算出样品中的HGF含量. 统计学处理:采用SPLM统计软件处理数据,各组间比较采用t检验.
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