缺氧对人RPE细胞表达HIF-1α的影响及T7-siRNAs对VEGF的干涉作用

发布时间：2017-12-30 17:25

本文关键词：缺氧对人RPE细胞表达HIF-1α的影响及T7-siRNAs对VEGF的干涉作用　出处：《吉林大学》2005年博士论文　论文类型：学位论文

【摘要】：膜新生血管(choroidal neovascularization,CNV)可引起黄斑出血、渗出,最终瘢痕形成而导致患者视力丧失。视网膜色素上皮(retinal pigment epithelium,RPE)在CNV的形成过程中具有重要地位。我们首先利用胰蛋白酶消化的方法分离并原代培养了人视网膜色素上皮细胞,同时进行了细胞鉴定。建立了hRPE细胞缺氧模型,并利用RT-PCR方法和免疫荧光方法检测了不同缺氧时相hRPE细胞中HIF一1a转录和表达情况。RNA干涉作用(RNAi)是新发现的可以高效特异抑制基因表达途径。我们根据VEGFl-5外显子序列确定了3处靶位点,并通过T7RNA聚合酶体外转录合成3对siRNA,转染RPE细胞后,高效特异地干涉了hRPE细胞中VEGF的表达。这些实验为研究CNV发生病理机制和基因治疗CNV奠定了基础。
[Abstract]:Choroidal neovascularization (CNV) may cause macular hemorrhage and exudation. Eventually scar formation results in visual loss. Retinal pigment epithelium (RPE) retinal pigment epithelium. RPE) plays an important role in the formation of CNV. Firstly, human retinal pigment epithelial cells were isolated and cultured by trypsin digestion. The anoxic model of hRPE cells was established. RT-PCR and immunofluorescence methods were used to detect the transcription and expression of HIF 1a in hRPE cells under different hypoxia conditions. We have identified three target sites according to the exon sequence of VEGFl-5. Three pairs of siRNAs were synthesized by T7 RNA polymerase transcription in vitro and transfected into RPE cells. These experiments provide a basis for studying the pathogenetic mechanism of CNV and gene therapy CNV.
【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2005
【分类号】：R363

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