摘要
目的观察熊果酸(ursolic acid,UA)玻璃体内注射对小鼠氧诱导视网膜病变的保护作用。方法将7 d龄清洁级C57BL/6J小鼠120只,用随机数字表法随机分为6组:空白对照组、模型对照组(PBS液)、阳性对照组(曲安奈德对照)和低剂量、中剂量、高剂量UA干预组,每组20只小鼠(均取右眼为实验对象)。空白对照组小鼠在空气中喂养,其他各组小鼠制备氧诱导视网膜病变模型。造模成功后,各组立即给予相应的药物治疗:模型对照组小鼠玻璃体内注射PBS液3μL,低剂量、中剂量、高剂量UA干预组小鼠玻璃体内分别注射1.5μg·L-1、3.0μg·L-1、6.0μg·L-1UA各3μL,阳性对照组小鼠玻璃体内注射曲安奈德(1 mL∶40 mg)3μL。17 d龄时过量麻醉处死小鼠,摘除眼球制备视网膜组织切片,HE染色,计数突破视网膜内界膜的新生血管内皮细胞核数,利用免疫组织化学法检测视网膜组织中VEGF、COX-2及MMP-2的阳性表达。结果模型对照组小鼠每张切片突破视网膜内界膜的新生血管内皮细胞核数为(18.65±3.24)个,明显高于空白对照组的(0.78±0.11)个,高剂量、中剂量UA干预组分别为(7.93±1.09)个和(13.32±1.87)个,均明显低于模型对照组,差异均有统计学意义(均为P<0.05)。不同组别视网膜组织中COX-2、VEGF和MMP-2蛋白的积分光密度值比较:模型对照组明显高于空白对照组,高剂量、中剂量UA干预组均明显低于模型对照组,差异均有统计学意义(均为P<0.05)。视网膜HE染色结果显示,空白对照组内界膜结构均一,血管内皮细胞排列整齐,偶见血管内皮细胞突破内界膜;模型对照组可见大量血管内皮细胞团突破内界膜,形成新生血管腔;低剂量UA干预组与模型对照组结果基本相似;高剂量UA干预组与阳性对照组结果接近,可见少量血管内皮细胞核突破内界膜,内界膜结构较均一,新生血管腔少见。结论 UA通过降低视网膜组织中的VEGF、COX-2及MMP-2的表达,抑制氧诱导小鼠视网膜病变模型新生血管的形成,并与UA的剂量呈正相关,对视网膜具有保护作用。
Objective To observe the protection of intravitreal injection of ursol-ic acid (UA) to oxygen-induced retinopathy in rats. Methods One hundred and twenty C57BL/6J rats of pathogen-free aged 7 days were randomly and evenly divided into 6 groups:blank control group, model control group (PBS solution), positive control group (triamcinolone acetonide ), UA intervention group (low, middle and high dosa- ges). All the right eyes were the experimental objects. The rats in blank control group were fed in the air, and neovascularization model was established by the rats in other groups. After the model establishment, rats in each group were given the corresponding drug at once:rats in model control group were given intravitreal injection of 3 μL PBS solution;rats in UA intervention group (low, middle and high dosages ) were given in- travitreal injection of 3 μL UA of 1.5 μg · L- 1,3.0 μg · L- 1, and 6.0 μg · L -1, respec- tively;the positive control group was given intravitreal injection of 3 ~L triamcinolone acetonide( 1 mL : 40 mg). Rats aged 17 days were killed through excessive anesthesia and the eyes were removed to prepare retinal tissue sections. Endotheliocyte nuclei of new vessels beyond the inner limiting membrane in sections were counted with HE stai- ning, and the positive expression of VEGF, COX-2 and MMP-2 in retinal tissues was de- tected by immunohistochemistry. Results The number of endotheliocyte nuclei of new vessels beyond the inner limiting membrane in each section of model control group was 18.65 ± 3.24, significantly higher than that of blank control group ( 0.78 ± 0. 11 ) ; UA intervention group of high and middle dosages were 7.93 ± 1.09 and 13.32 ± 1.87, respectively ,both significantly lower than that of model control group. All the differ- ences were of statistical significance ( all P 〈 0.05 ). Integrated optical density compari- son of COX-2, VEGF and MMP-2 in retinal tissues of different groups: model control groups markedly higher than blank control group; UA intervention group of high and middle dosages were significantly lower than model control group. All the differences were statistically significant(P 〈 0.05 ). HE staining showed that in blank control group, inner limiting membrane structure was uniform, vascular endothelial cells were orderlyand vascular endothelial cells were occasionally beyond inner limiting membrane;in model control group, many vascular en- dothelial cells were beyond inner limiting membrane and formed new blood vessel cavity; The result of UA intervention group of low dosage was almost same with model control group; Vascular endothelial cells in UA intervention group of high dosage and positive control group were close to inner limiting membrane, and a few were beyond it, the inner limiting mem- brane structure was uniform, and new blood vessel cavity was seldom seen. Conclusion UA may inhabit the formation of new blood vessel through reducing the VEGF, COX-2 and pression has a positive correlation with UA dosage. MMP-2 expression in retinal tissues to protect retina, and the ex-pression has a positive correlation with UA dosage.
出处
《眼科新进展》
CAS
北大核心
2013年第10期918-921,926,共5页
Recent Advances in Ophthalmology
基金
河南省新乡市重点科技攻关计划基金资助(编号:ZG12013)~~