高氧诱导小鼠视网膜病变模型中血管内皮生长因子和色素上皮衍生因子的表达及意义

目的 研究高氧诱导小鼠视网膜病变模型中血管内皮生长因子(VEGF)和色素上皮衍生因子(PEDF)的表达和意义,并探讨两者在视网膜新生血管形成过程中的作用.方法 对照实验研究.对新生C57BL-6N系小鼠给予高氧后,置相对低氧环境中饲养,诱导产生视网膜新生血管.在小鼠生后第12、14及17天摘除眼球,通过逆转录聚合酶链反应和免疫印迹法及荧光素灌注造影,分别检测不同时间点全视网膜新生血管组织对VEGF与PEDF的mRNA和蛋白质的表达水平的差异,并观察视网膜新生血管的分布与形态变化,通过血管内皮细胞计数对新生血管进行量化.应用SPSS11.5统计学软件,采用析因设计方差分析,分别比较实验组与对照组mRNA与蛋白质表达水平的差异,以P<0.05作为差异有统计学意义.结果 在高氧环境中(出生第12天小鼠),OIR模型鼠VEGF蛋白质表达A值(0.47±0.12)较正常鼠(1.81±050)下降,PEDF蛋白质表达A值(5.35±0.94)较正常鼠(0.68±0.17)明显升高;而相对低氧环境中(出生第14和17天小鼠),VEGF蛋白质表达A值(2.15±0.46,5.49±0.97)较正常鼠(0.90±0.05,0.88±0.91)明显升高,PEDF蛋白质表达A值(2.07±0.35,1.37±0.48)较正常鼠(2.62±0.68,5.30±0.59)明显下降,尤以第17天下降显著.对不同时间点VEGF和PEDF蛋白质表达水平进行析因方差分析,结果显示各时间点的VEGF蛋白质表达水平差异有统计学意义(F=70.450,P=0.000),各时间点的PEDF蛋白质表达水平差异有统计学意义(F=160.237,P=0.000).出生第12、14及17天小鼠视网膜VEGF蛋白质的表达与正常对照组比较,差异有统计学意义(P=0.009,0.010,0.000),PEDF蛋白质的表达差异也有统计学意义(P=0.002,0.046,0.000);出生第12、14及17天小鼠视网膜VEGF mRNA的表达与正常对照组比较,差异有统计学意义(P=0.001,0.000,0.001),PEDF mRNA的表达差异也有统计学意义(P=0.000,0.001,0.000).伴随着视网膜组织的缺氧,出现了VEGF蛋白质表达水平升高和PEDF蛋白质表达水平降低,导致视网膜组织中VEGF与PEDF蛋白质表达失衡;VEGF与PEDF的mRNA表达亦发生类似变化,且较蛋白质改变提前发生.上述改变均伴随着同期视网膜新生血管的发生、发展及其严重程度而逐渐加重.结论 VEGF和PEDF的mRNA和蛋白质表达失衡可能足造成视网膜新生血管发生的机制之一.(中华眼科杂志,2008,44:734-740)

Expression of vascular endothelial growth factor and pigment epithelium derived factor In mouse oxygen-induced retinopathy and its significance

Objective To investigate the expression and significance of vascular endothelial growth factor (VEGF) and pigment epithelium derived factor (PEDF) in oxygen-induced mouse retinopathy. Methods This experiment was a control experiment study. Newborn C57BL-6 mice were exposed to hyperoxia, and then returned to normoxia to induce retinal neovascularization. Mice were sacrificed at postnatal days 12, 14 and 17 and the retina were processed for RT-PCR, Western Blot and fluorescein angiography. Analysis of variance and Dunnett's t3 was used to compare the mRNA and protein level of VEGF and PEDF between experiment and control groups. Statistical difference was considered significant at a P value less than 0. 05. Results At any of the time points tested, there was significant difference at VEGF protein level (A value) between the experiment (0.47±0.12,2.15±0.46,5.49±0.97) and control(1.81±0. 50,0.90±0.05,0.88±0.91) groups(P=0. 009,0. 010,0. 000, respectively);the same situation occurred at PEDF protein level (P=0. 002,0. 046,0. 000,respectively) . At postnatal day 12,14 and 17 ,a significant difference at VEGF protein level was observed among the different experiment groups (P =0. 002,0. 001,0. 000, respectively); the same situation occurred at PEDF protein level( P =0.009,0.010,0.000, respectively). There was significant difference at VEGF mRNA level between the experiment and control groups (P= 0. 001,0. 000,0. 001);at postnatal day 12 and 14, the same situation occurred at PEDF protein level(P=0.001,0.000, respectively) ,but there was no significant difference at postnatal day 17(P=0.612). At postnatal day 12,14 and 17,a significant difference at VEGF mRNA level was observed among different experiment groups (P=0.000, 0.001,0.000, respectively); the same situation occurred at PEDF mRNA level (P=0.000,0.001,0.000, respectively). The time course of the decrease of PEDF was consistent with the increase of VEGF expressiorL VEGF/PEDF ratio change was correlated with the development and progression of retinal neovascularization. The time course of PEDF mRNA down-regulation was consistent with the VEGF mRNA up-regulation, similar with the changes of the protein. The change of VEGF and PEDF mRNA was prior to that of the protein. Conclusions One of the mechanisms for development of retinal neovascularization is the changes of VEGF\PEDF level in the retina.(Chin J Ophthalmol, 2008,44:734-740)