氧诱导视网膜病变小鼠模型新生血管变化规律及VEGF的动态表达

Rule of new vascular changes and dynamic expression of VEGF in mice models with oxygen induced retinopathy

目的观察氧诱导视网膜病变(oxygen induced retinopathy,OIR)小鼠模型新生血管变化规律及血管内皮生长因子(vascular endothelial growth factor,VEGF)的动态表达,利用VEGF的变化反映OIR模型视网膜组织中新生血管的发生发展。方法80只C57BL/6J新生幼鼠随机分为OIR组和对照组,每组各40只。OIR组7d龄鼠暴露在(75±2)%浓度的高氧状态下5d,随后在正常空气环境下饲养;对照组小鼠,均在正常空气环境下饲养。2组均于12d龄(P12)、14d龄(P14)、17d龄(P17)、21d龄(P21)随机抽取2组小鼠各10只(20眼)处死,利用视网膜组织切片,HE染色计数突破视网膜内界膜的内皮细胞核数,分析视网膜新生血管增生情况;实时荧光定量RT-PCR(real time RT-PCR)观察VEGF mRNA的动态表达规律。结果突破视网膜内界膜的内皮细胞核数显示对照组小鼠视网膜新生血管P12、P14、P17、P21分别为0.27±0.12、0.30±0.15、0.31±0.13、0.40±0.17,自始至终很少见到突破内界膜的内皮细胞核;OIR组小鼠视网膜新生血管P12、P14、P17、P21分别为0.41±0.16、10.25±2.68、34.14±1.79、8.12±2.13,P17达高峰,以后逐渐下降。real time RT-PCR显示对照组视网膜病变中VEGF mRNAP12、P14、P17、P21分别为9.29±0.13、9.52±0.17、10.49±0.24、10.51±0.29;P12时表达相对较高,以后逐渐下降。OIR组视网膜病变中VEGF mRNAP12、P14、P17、P21分别为10.45±0.11、7.41±0.48、8.53±0.32、10.56±0.17;P12表达较低,P14达高峰,以后逐渐下降。2组VEGF mRNA P12、P14、P17时表达差异均有统计学意义(均为P<0.05),在P21时表达差异无统计学意义(P>0.05)。结论VEGF是重要的血管生成因子,利用VEGF mRNA的动态表达可以映射视网膜新生血管的发生发展过程,为进一步研究奠定基础。

Objective To investigate rule of new vascular changes and dynamic expression of vascular endothelial growth factor（VEGF）in mice models with oxygen induced retinopathy（OIR）and to map the development and progression of new vascular changes of mice models with OIR by the changes of VEGF.Methods Eighty 7-day-old C57BL/6J newborn mice were divided into two groups,40 mice in each group.Mice in OIR group were exposed to（75±2）% oxygen for 5 days and then to room air.Mice in control group were exposed to room air. Ten mice （20 eyes） in each group were ran domly chosen and sacrificed at postnatal 12 days （P12） ,14 days （P14） ,17 days （P17） and 21 days （ P21 ） in both two groups, respectively. The proliferative neovascular responses were analyzed by counting the number of neovascular cell nucleus extending into the internal limiting membrane with retinal sections and HE staining. Dynamic expression of VEGF mRNA was observed by real-time RT-PCR. Results The number of neovascular cell nucleus extending internal limiting membrane showed that new vessels were 0.27 ±0.12,0.30±0. 15,0.31±0. 13 and 0.40±0. 17 at P12,P14,P17 and P21 in control group, respectively, and neovascular cell nucleus extending into the internal Um- iting membrane was seldom to be seen, were 0.41± 0. 16,10.25 ± 2.58,34.14 ± 1.79, and 8.12 ±2.13 at P12,P14,P17 and P21 in OIR group,respectively,went to peak at P17 and then gradually decreased. Real-time RT-PCR showed that VEGF mRNA were 10.45 ± 0. 11,7.41 ±0.48,8.53 ±0.32 and 10.56±0. 17 at P12,P14,P17 and P21 in OIR group,respectively,was lower at P12,went to peak at P14 and then gradually decreased, were 9. 29 ±0.13,9.52±0.17,10.49±0.24 and 10.51±0.29 at P12,P14,P17 and P21 in control group, respectively, was relatively higher at P12 and gradually decreased. There was statistical difference in VEGF mRNA at P12, P14 and P21 between the two groups（ all P 〈 0.05 ）, but no difference was found at P21 between the two groups（ P 〉 0.05 ）. Conelusion VEGF is an important neovascular growth factor. Dynamic expression of VEGF mRNA can reflect the development and progression of retinal new vessels and lay the foundation for further studies.