摘要
背景寻求有效的血管内皮生长因子（VEGF）抑制剂是治疗和预防新生血管性眼病的关键。胰岛素样生长因子结合蛋白相关蛋白-1（IGFBP—rP1）是一种新发现的抑血管新生因子，推测其在眼内有抑制VEGF的作用。目的探讨IGFBP—rP1对VEGF体外诱导视网膜新生血管形成的抑制作用及其机制。方法使用含质量分数10％FBS的DMEM对猕猴视网膜／脉络膜血管内皮细胞株（RF／6A）进行扩增培养，利用免疫荧光细胞化学染色法观察RF／6A细胞表达IGFBP—rP1的情况。RF／6A细胞血清饥饿法培养24h后分为对照组、10mg／LVEGF组，50、100、200mg／LIGFBP—rP1＋10mg/L VEGF组进行干预，分别利用MTS比色法、Transwell实验和流式细胞术比较IGFBP—rP1（0、50、100、200mg／L）联合VEGF（10mg／L）作用后，RF／6A细胞在增生、移行和凋亡等生物学行为方面的变化。结果RF／6A细胞用不同质量浓度的IGFBP—rP1培养后细胞质呈FITC激发后的绿色荧光，细胞核呈PI激发后的红色荧光，而对照组细胞仅见细胞核的红色荧光。10mg／LVEGF组RF／6A细胞的A490值、移行细胞数与对照组相比明显增加，差异均有统计学意义（t=-15．191，P=0．000；t=-21．274，P=0．000），细胞凋亡率明显下降，差异有统计学意义（t=10．228，P=0．000）。与10mg／LVEGF组相比，IGFBP—rP1（50、100、200mg／L）＋10mg／LVEGF组RF／6A细胞的A490值、移行细胞数明显下降（均P〈0．05）。50、100、200113geLIGFBP—rPl＋10mg／LVEGF组RF／6A细胞的细胞凋亡率分别提高了（1．26±0．04）％、（1．50±0．07）％和（1．93±0．27）％，各组的总体差异有统计学意义（F=274．273，P=0．000）。结论IGFBP—rP1作为一种内源性因子，通过促细胞凋亡机制抑制VEGF诱导的视网膜血管的生成。
Background Antagonists against vascular endothelial growth factor （VEGF） play key roles in treating and preventing neovascular ophthalmopathy. As a novel anti-angiogenic factor, insulin-like growth factor binding protein-related protein 1 （IGFBP-rP1） might be an antagonist against VEGF in eye. Objective This study was to explore the inhibitory effect of IGFBP-rP1, a novel anti-angiogenic factor, on VEGF-induced retinal angiogenesis in vitro. Methods The retina-choroid endothelial cell line （RF/6A） was cultured in DMEM containing 10% fetal bovine serum. Culture cells were divided into control group（free-serum culture group） , 10 mg/L VEGF culture group and different concentrations of IGFBP-rP1 （50,100,200 mg/L） ＋ 10 mg/L VEGF group. The expression of IGFBP-rP1 in the cells was detected by immunofluorescence assay. The proliferation and migration of RF/6A ceils were evaluated using MTS colorimetric assay and the chemotactic motility assay, respectively. Flow cytometry was used to detect the apoptosis of RF/6A cells. Results The immunofluorescence assay RF/6A cells showed the green fluorescence in cytoplasm and red fluorescence in nuclei after cells were exposed to any concentration of IGFBP-rP1 ,but only red fluorescence was seen in nuclei in control cells. After stimulation of 10 mg/L VEGF,the proliferation value （A490） was elevated and the numbers of ceil migration were increased in comparison with control group （ t = - 15.191, P = 0. 000 ; t = -21. 274, P = 0. 000 ）, but the cellular apoptosis rate was lower than the control group （ t = 10. 228, P = 0. 000）. After treated with various concentrations of IGFBP-rP ＋ 10% VEGF, the proliferation and migration of RF/6A cells were significantly decreased in comparison with only 10% VEGF group （F= 534. 158 ,P=0. 000 ;F= 2742. 323, P = 0. 000, respectively）, and the inhibitory effects were gradually enhanced with the increase of IGFBP-rP1 levels （P 〈0.05）. The apoptosis rate of RF/6A cells in 50,100 and 200 mg/L＋ 10 mg/L VE
出处
《中华实验眼科杂志》
CAS
CSCD
北大核心
2011年第2期113-117,共5页
Chinese Journal Of Experimental Ophthalmology
基金
国家自然科学基金重点项目（30930097）、国家自然科学基金青年基金（30801269）、上海交通大学医学院搏士创新基金（BXJ0935）、上海市眼底病重点实验室基金（09-10）
