角膜论文：阿瓦斯汀不同的给药途径对兔角膜新生血管及角膜超微结构的影响.doc

角膜论文：阿瓦斯汀不同的给药途径对兔角膜新生血管及角膜超微结构的影响【中文摘要】通过研究阿瓦斯汀不同的给药途径分别对兔角膜新生血管及超微结构的影响,探索阿瓦斯汀治疗角膜新生血管较安全持久有效的给药途径,为临床应用阿瓦斯汀治疗角膜新生血管性疾病提供实验依据。方法：选用健康新西兰大白兔55只,兔龄2.5-3个月,雌雄不限。随机选50只兔左眼用1mol/L氢氧化钠溶液烧伤,建立碱烧伤角膜新生血管动物模型,造模成功后选取48只随机分为A点眼液组12只(5mg/ml,3次/日)、B结膜下注射组12只(25mg/ml,1次/周),C角膜基质注射组12只(25mg/ml,1次/周),D模型对照组12只(不用药),选取未造模2只兔为空白对照,于碱烧伤术后1天、4天、给药后一周、二周、四周观察造模兔眼结膜充血、角膜浑浊、新生血管生长情况及角膜组织的完整性,并进行眼前节照相,并测量各试验兔眼角膜新生血管的长度及分布钟点位数以计算角膜新生血管面积,与给药后一周、二周、四周三个时间点后用空气栓塞法每组各处死4只试验用兔,处死后即刻抽取试验眼房水约0.1m1用于检测房水中的VEGF蛋白的含量,并取新生血管生长最旺盛的角膜组织分别固定,待做电镜、免疫组化(角膜组织中CD31)等检测。结果：1、在碱烧伤后各观察时间点房水中VEGF的浓度均高于正常兔眼水平,碱烧伤后11天内房水中VEGF蛋白含量增加迅速,约18天左右时房水中VEGF蛋白含量达到最高水平,之后有所下降,到32天时达到一个较稳定的水平,不同处理方法之间相比较在三个时间点差别均有统计学意义。2、碱烧伤角膜新生血管在碱烧伤后12天内生长迅速,14天时有所消退,之后新生血管趋于相对稳定状态,不同处理方法之间相比较在三个时间点差别均有统计学意义。3、免疫组化染色结果示：CD31在正常兔眼角膜组织中未有表达,各治疗组阳性细胞数明显少于模型对照组,差异有统计学意义。4、电镜检查结果：ABD组角膜超微结构在一周、二周、四周时除碱烧伤损伤外无明显其它改变,在给药后第二周、四周时C组角膜超微结构损伤要重于ABD组。结论：1、兔眼房水中VEGF蛋白含量在碱烧伤后11天内增长较快,至18天左右时达到高峰水平,之后VEGF有所下降,最后趋于较稳定水平。2、点眼液(5mg/ml,3次/日),结膜下注射(25mg/ml,1次/周),角膜基质注射(25mg/ml,1次/周)三种给药途径都对碱烧伤角膜新生血管增殖期动物模型有较好的抑制作用,同时也能减轻角膜的混浊。3、点眼液与结膜下注射给药途径简单安全、效果相对稳定,在观察期内未见对角膜超微结构的产生明显影响。4、角膜基质注射给药途径虽短期效果显著,但在观察期内对角膜超微结构产生较明显的影响。【英文摘要】:This article is to explore the safe and effective approach of avastin to corneal neovascnlarization,through studying the influence of different administration route of avastin to corneal neovascnlarization and ultrastructure,and then provid theoretical evidence to clinication application of avavtin.Method:55 healthy adult New Zealand white rabbit, male and female merger,50 rabbit were selected to establish corneal new vessel animal model with alkali bure,established rabbits were selected for four groups at random,group A(n=12) received eye drops; group B(n=12) received a subconjunctival injection of 0.1 milliliter avastin; group C(n=12) received a corneal stroma injectin of 0.02 milliliter of avastin; group D(n=12) model control group. All groups were observed the corneal opacity,NV and cornela integrity every day after alkali burn. All groups are measured the length and extent of corneal by anterior photograph at day 7,14,32 after administration,and then sacrifice four rabbits each group with aeroembol-ism,instantly draw ocular humor or so 0.1 milliliter.The content of VEGF protein in aqueous humor was deteted by Elisa.The location and espression of CD31 were determined by immunohisto-chemical method.Corneal ultrastructure morphology was observed by transmission election microscope.Results:The extent of VEGF in aqueous humor of the treatment group was less than that of control group at each time.From the point of corneal NV counted number,the NV in the treatment group was less than that of the control group after alkali burn (P0.05) with statistically significant difference.The expression of CD31 in the cornea was restricted in the treatment group.With the ultrastructure morphology, groupC had no difference with groupA. groupB at day 7,14,but groupC had created relatively obvious injury to corneal ultrastructure morphology at day 32.Conclusion:1. The extent of VEGF in aquous humor increase fast at 10 day after alkali burn,and reach a peak at about2 weeks, then descend a fairly stable level at 32 day2.Three administration can effectively restrain corneal NV in short time.3.Eye drop and subconjunctival injection administration were safe and effective in fairly long time,and had no influence to corneal ultrastructure morphology4.Corneal strome injection administration had damage to comeal ultrastructure morphology for long time.【关键词】角膜 新生血管 阿瓦斯汀 眼液 结膜下注射 基质注射【英文关键词】Corneal neovascnlarization Avastin eye-drop subconjunctival injection stroma injectin【目录】阿瓦斯汀不同的给药途径对兔角膜新生血管及角膜超微结构的影响摘要3-5ABSTRACT5-6缩略语表9-10第1章 引言10-121.1 概述101.2 研究现状10-12第2章 材料与方法12-182.1 材料12-132.1.1 实验动物122.1.2 主要试剂122.1.3 实验仪器、设备12-132.2 实验方法13-142.2.1 造兔左眼碱烧伤角膜新生血管模型132.2.2 术后处理132.2.3 实验分组132.2.4 观察指标13-142.2.5 动物处死方法、时间与取材142.3 术后观察14-172.3.1 活体观察指标142.3.2 角膜新生血管定量14-152.3.3 组织病理学观察15-162.3.4 房水VEGF检测16-172.4 统计分析17-18第3章 结果18-233.1 结膜,角膜大体情况18-193.1.1 裂隙灯