血管抑素降低ROP幼鼠视网膜及虹膜血管渗漏性的作用机制研究

目的：探讨血管抑素（angiostatin）玻璃体注射对氧诱导的早产儿视网膜病变（retinopathy of prematurity，ROP）的视网膜及虹膜血管渗漏的作用及其机制。方法：将出生7d（P7）的Brown Norway鼠置高氧环境（750mL／L O2）5d后再置正常氧环境诱导ROP，建立ROP动物模型，并以年龄相匹配的正常鼠作为正常对照。所有ROP鼠（P14）及正常鼠均右眼玻璃体腔注射血管抑素，左眼注射相同剂量的PBS（磷酸盐缓冲生理盐水）作为对照。用Evans蓝微血管渗透性检测法及总蛋白标准化分别于注射后1，2和3d检测视网膜和虹膜的血管渗透性；用Western blot蛋白印迹分析和免疫组化方法检测注射24h后血管内皮生长因子（vascular endothelial growth factor，VEGF）在视网膜的表达。结果：ROP鼠视网膜及虹膜的血管渗透性明显增加（P〈0．01）；中剂量（3．75ug/眼）和高剂量（7．5ug/眼）血管抑素降低ROP鼠视网膜血管渗透性（P〈0．05，P〈0．01），而低剂量组（1．88ug／眼）没有引起明显改变，呈现剂量依赖型；三种不同剂量的血管抑素玻璃体注射后ROP鼠的虹膜均未发生明显的血管渗透性的改变。血管抑素注射后第1d和第2d视网膜血管渗透性明显降低（P〈0．05。P〈0．01），而第3d无明显降低，其作用呈现出时间进程。Western blot蛋白印迹和免疫组化分析表明血管抑素显著降低了ROP鼠视网膜的VEGF水平，但对正常鼠无影响。结论：血管抑素可以降低ROP鼠视网膜的病理性血管渗漏，其血管渗透性下降可能与血管抑素下调VEGF的表达有关。血管抑素可能对ROP等其他视网膜血管渗漏性疾病具有潜在的治疗作用。

Mechanism of angiostatin induced reduction of vascular leakage in retina and iris of rats with retinopathy of prematurity

·AIM: To study the effect of an intravitreal injection of angiostatin on vascular leakage in the retina and iris of oxygen-induced retinopathy of prematurity (ROP).·METHODS: Brown Norway rats at postnatal day 7 (P7) were exposed to hyperoxia (750mL/L O2 )for 5 days (P7-12) and then returned to normoxia to induce retinopathy. Angiostatin was reconstituted in sterile Phosphate Buffered Saline(PBS) and diluted to desired different concentrations. Angiostatin solution was injected into the vitreous of the right eye of the ROP rats at P14 and the age-matched normal rats through pars plana using a glass capillary, and the left eye received the same volume of sterile PBS as the control. Vascular permeability was quantified at 1, 2 and 3 days after the injection by measuring albumin leakage from blood vessels into the retina and iris using the Evans blue method and normalized by total protein concentrations. The expression of vascular endothelial growth factor (VEGF) in retina was evaluated using the Western Blot analysis and immuno-histochemistry 24 hours following the injection.·RESULTS: ROP rats showed significant increases of vascular permeability in the retina and iris (P<0.01). Angio-statin reduces vascular permeability in a dose-dependent manner in the retina of ROP rats. The reduction showed a time course trend. [Angiostatin injection reduced retinal vascular permeability by approximately 1.5 and 2-fold at P15 (P<0.05) and P16 (P<0.01), respectively.] Angiostatin injection significantly reduced VEGF levels in the retina of ROP rats but did not affect retinal VEGF levels in normal rats.·CONCLUSION: Angiostatin significantly decreases pa-thological vascular permeability in the retina and iris of ROP rats but not in normal rats. Angiostatin down-regulates VEGF expression in retina of ROP rats. These results suggest that angiostatin may have a therapeutic potential in the treatment of ROP and other diseases with vascular leakage.