氧诱导小鼠视网膜新生血管的实验研究

Study on oxygen-induced retinopathy in a mouse model of retinopathy of prematurity

目的制作氧诱导视网膜新生血管的小鼠动物模型并了解其视网膜血管内皮生长因子(VEGF)的变化。方法将出生后第7d的C57BL/6J小鼠置于75%的高氧环境中5d,再置于普通空气中5d。在空气中饲养的小鼠为对照组。两组进行视网膜铺片,ADPase染色,组织切片染色,ELISA测定视网膜VEGF蛋白含量。结果实验组新生血管形成率为100%,对照组未见新生血管。实验组小鼠生后第17d时突破视网膜内界膜的血管内皮细胞核数目达(46.7±11.1)个,对照组不足2个。第12d实验组视网膜VEGF蛋白水平比对照组下降,第17d比对照组升高。结论该动物模型是研究早产儿视网膜病变(ROP)发病机制及治疗的合适模型。VEGF是造成视网膜新生血管发生的主要机制之一。

Objective This study was to establish an animal model of oxygen-induced retinopathy（OIR） and investigate the expression and signification of vascular endothelial growth factor（VEGF） in oxygen-induced retinopathy. Methods Eighty 7-day-old C57BL/6J mice were divided into two groups. Mice in hyperoxic group were exposed to 75% oxygen for 5 days and then to room air for another 5 days. Mice were exposed to room air as normoxic control group. Experimental and control mice were sacrificed at postnatal day 12 and 17 respectively. The proliferative neovascular response was estimated by observing the vascular pattern in adenosine diphosphatease （ADPase） stained flat-mounted retinas. The new vascular cell nuclei extending into the internal limiting membrane was examined in the cross-section of retinas of the left eyes of mice. Enzyme linked immunosorbent assay（ELISA） was used for the determination of VEGF expression. Results Hyperoxia-induced neovascularization appeared at the junction zone between vascular and avascular retinas in the mid-periphery in all mice exposed to hyperoxia. No neovascularization were seen in the control group. The numbers of new vascular cell nuclei extending into the internal limiting membrane were 46. 7 ± 11.1 in hyperoxic group and 1.2 ± 1.4 in normoxic control group,showing a significant difference between them（P 〈 0.01 ）. The expression level of VEGF protein in oxygen-induced retinopathy group was 63% of control group in postnatal 12-day mice. With the development of new blood vessel,expression levels of VEGF protein in retina in oxygen-induced retinopathy group increased by 2. 8 fold. Conclusion This animal model of OIR is reproducible. It is useful for the study of pathogenesis of retinal neovascularization and therapeutic intervention for ROP. One of the mechanisms for development of retinal neovascularization may be related to up-regulation of VEGF.