视网膜色素上皮细胞通过p75NTR受体调控脉络膜新生血管生成的机制研究

Choroidal neovascularization (CNV) is the primary pathogenic cause of age-related macular degeneration (AMD). Until recently, the only effective treatment is to inhibit CNV generation and development. Retinal pigment epithelium (RPE) oxidative damage play a crucial role in CNV formation. Several studies have verified that under oxidative damage, the p75NTR receptor of RPE is activatied, which is one of the members of tumor necrosis factor superfamily, and promote vascular endothelial proliferation. However, the regulation mechanisms between p75NTR receptor and CNV generation is unclear. Our previous study validated that p75NTR receptor was elevated after oxidative stress in RPE, and induce the up-regulation of vascular endothelial growth factor and inflammatory cytokins. To explore the relation between p75NTR receptor in RPE and CNV generation, RPE cell lines, primary RPE and CNV animal model were used in this study. The dose-time dependent expression of p75NTR receptor was evaluated after RPE subject to oxidative damage; p75NTR receptor-target agonists/antagonists, gene overexpression/gene konck down method were used to study the behavior changes of RPE; the alteration of pro-/anti- angiogenesis factors, matrix metalloproteinase and tissue inhibitor of matrix metalloprotease, the inflammatory cytokines and JNK signal pathway were evaluated; the inhibitory efficacy and possible mechanisms of different p75NTR receptor-targeted drugs were studied in CNV animal model. This study will enrich our knowledge on the mechanisms of CNV formation and explore new strategies for CNV treatment.

脉络膜新生血管(CNV)是年龄相关黄斑变性等致盲眼病的首要致病因素，目前唯一有效的治疗途径是抑制其生成及发展。视网膜色素上皮细胞(RPE)氧化损伤在CNV生成中具有重要作用。研究发现RPE氧化损伤能激活属于肿瘤坏死因子超家族的p75NTR受体，引起血管内皮细胞增殖，但其调控CNV生成的机制未明。本课题预实验发现RPE氧化损伤后p75NTR受体、血管内皮细胞生长因子、炎症因子表达均升高。为探索RPE中p75NTR受体与CNV生成的关系，本课题拟利用RPE细胞系、原代RPE及CNV动物模型，分析不同氧化损伤下RPE中p75NTR受体表达变化；并通过受体激动剂/抑制剂、基因过表达/基因敲减方法，研究RPE行为学功能，血管生成促进/抑制因子、基质金属蛋白酶/基质金属蛋白酶组织抑制剂、炎症因子及JNK信号通路表达变化；最后探索受体靶向性药物抑制CNV生成的疗效及可能机制，以期寻找CNV治疗新靶点。

脉络膜新生血管(CNV)是年龄相关黄斑变性等致盲眼病的首要致病因素，目前唯一有效的治疗途径是抑制其生成及发展。视网膜色素上皮细胞(RPE)氧化损伤在CNV生成中具有重要作用。研究发现RPE氧化损伤能激活属于肿瘤坏死因子超家族的p75NTR受体，引起血管内皮细胞增殖，但其调控CNV生成的机制未明。本课题预实验发现RPE氧化损伤后p75NTR受体、血管内皮细胞生长因子、炎症因子表达均升高。由此，本课题利用RPE细胞系及CNV动物模型，分析不同氧化损伤下RPE中p75NTR受体表达变化；并通过受体激动剂/抑制剂、基因过表达/基因敲减方法，研究RPE行为学功能，血管生成促进/抑制因子、基质金属蛋白酶/基质金属蛋白酶组织抑制剂、炎症因子及JNK信号通路表达变化；最后探索受体靶向性药物抑制CNV生成的疗效及可能机制，以期寻找CNV治疗新靶点。.本课题通过翔实的研究证实：p75NTR受体在RPE细胞中具有表达，但其表达水平较低；p75NTR受体激动剂增加RPE细胞的损伤，引起RPE细胞增殖能力下降、凋亡增加、ROS释放、线粒体膜电位减低，导致RPE细胞Caspase 3和细胞色素c (cyto-c)表达增高。上述结果均证实p75NTR受体的激活将导致RPE细胞的形态学及功能学改变，且为损伤性改变。除外证实p75NTR受体激动剂在RPE细胞中的作用以外，我们探索了t-BHP、Ab对ARPE19细胞氧化损伤合适浓度，同时证实了t-BHP以及OAb1-42可以引起p75NTR受体及其共受体TrkA和配体NGF的表达增高，并导致内质网应激，为下一步实验提供依据。.在新生血管方面的研究中，证实在患者糖尿病视网膜病变增殖膜中，p5NTR与新生血管共定位，同时，在接受anti-VEGF治疗的患者中，p75NTR表达明显减低。在动物模型中，我们证实，p75NTR受体抑制剂能够抑制激光诱导的脉络膜新生血管发生及发展，也能够抑制氧诱导视网膜新生血管的生成。p75NTR受体在新生血管中的作用机制为开发新的抗新生血管药物提供了新的研究靶点。