摘要
目的探讨携带血管生成素1(Ang-1)基因的骨髓间充质干细胞(BMSCs)对氧诱导视网膜病模型小鼠视网膜新生血管的作用。方法将出生7天的昆明小鼠随机分成空气对照组和高氧模型组(其中包括注射Ang-1+BMSCs组、单纯BMSCs组及高氧PBS组),每个亚组12只。除空气对照组之外,其余组均放入氧分压65%±5%的氧箱内饲养5天后取出。将携带Ang-1的BMSCs和不携带Ang-1的BMSCs各2 ml(内含细胞总数为1×105)在鼠龄第7、12、17日分别腹腔注入高氧模型组中的Ang-1+BMSCs组和单纯BMSCs组小鼠体内,空气对照组和高氧PBS组注入相同剂量的PBS缓冲液。于鼠龄17天时取小鼠眼球做标本固定,分别用免疫组织化学法检测Ang-1蛋白、qRT-PCR法检测Ang-1蛋白mRNA、计数突破视网膜内界膜的血管内皮细胞数及明胶墨汁灌注眼底视网膜血管,比较小鼠视网膜新生血管情况。结果高氧模型各组每只眼球均可见突出内界膜的新生血管内皮细胞。其中Ang-1+BMSCs组、单纯BMSCs组与高氧PBS组相比较,突破视网膜内界膜的血管内皮细胞核明显减少,差异有统计学意义(P<0.05);与高氧PBS组比较,Ang-1+BMSCs组及单纯BMSCs组血管走形较直,Ang-1蛋白表达量增加,免疫组织化学染色被黄染的胞浆较丰富;Ang-1+BMSCs组与单纯BMSCs组相比,前者血管走行较直,周边新生血管覆盖率降低,突破内界膜新生血管细胞数目减少,Ang-1蛋白的表达量增多。结论高氧能诱导新生鼠视网膜新生血管生成;Ang-1基因与BMSCs结合与单纯应用BMSCs相比,前者能更有效地改善视网膜新生血管的增生状况。
Objective To investigate the role of Angiogenin-1 (Ang-1) expression inbone marrow mesenchymal stem cells (BMSCs) in angiogenesis of oxygen induced retinopathy of prematurity.Methods 7-day-old kunming mice were divided into a room air control group,and 3 hyperoxia groups (Ang-1 + BMSCs group,BMSCs group and PBS hyperoxia control group) with 12 mice in each group.Mice in the hyperoxia groups were exposed to(65 ± 5)% oxygen for 5 days prior to living in room air.On day 7,12 and 17 days of life,while mice in the Ang-1 + BMSCs and BMSCs groups received 2 ml of bone marrow (1 × 105 cells) either with or without Ang-1 expression via intra-abdominal injections; those mice in the room air and hyperoxia control groups received 2 ml of PBS at the same corresponding ages and through the same route.Eyes were removed on postnatal day 17.Specimens were studied using immunohistochemical and qRT-PCR methods to count the number of breakthrough endothelial cells through the retina boundary membrane.Gelatin ink perfusion of retinal blood vessels was also performed to investigate the new retinal blood vessel formation in mice.Results Neo-vascular endothelial cells were seen extending into the inner boundary membrane of retina in mice eyes of all three groups of mice exposed to high-concentration of oxygen.The numbers of the neo-vascular endothelial cells werefewer in the two treatment groups (Ang-1 + BMSCs group and BMSCs group) than in the hyperoxia PBS control group(P <0.05);Compared to the samples in the hyperemia PBS control group,the two intervention groups demonstrated straighter neo-blood vessels and higher expression level of the the Ang-1 target gene,as well as more intense cytoplasmic signals in immunohistochemical stain.Comparing to mice in the BMSCs group,those in the Ang-1 + BMSCs group has formed straighter neo-blood vessels,decreased neoblood vessel formation in the surrounding area,fewer endothelial cells breaking through the inner boundary of retina and higher expression level of the the Ang-1 target gene.Conclusions High concentration oxygen exposure can induce retinal angiogenesis in mice; BMSCs with Ang-1 over expression,when compared to BMSCs only,is more effective in improving the growth of retinal blood vessels.
出处
《中国新生儿科杂志》
CAS
2015年第1期62-68,共7页
Chinese Journal of Neonatology
基金
国家自然基金青年基金(81000265)
关键词
早产儿视网膜病
血管生成素1
间质干细胞移植
视网膜新生血管化
Retinopathy of prematurity
Angiopoietin-1
Mesenchymal stem cell transplantation
Retinal neovascularization