兔眼玻璃体腔注射重组人血管内皮抑制素的安全性研究

Safety of intravitreal injection of recombinant human endostatin in rabbits

目的评估不同质量浓度重组人血管内皮抑制素(rh-endostatin)兔眼玻璃体腔注射的安全性。方法取健康成年新西兰白兔30只,以右眼为实验眼,按照计算机数字随机分配法随机平均分为0.125 mg rh-endostatin组、0.250 mg rh-endostatin组、0.500 mg rh-endostatin组、生理盐水组和正常对照组,每组6只。各注射组玻璃体腔注射100μl不同剂量的rh-endostatin或生理盐水,正常对照组不做处理。玻璃体腔注射前,注射后1、3、7、14、30和60 d,裂隙灯显微镜和间接检眼镜下分别检查术眼眼前节和眼底,测量术眼眼压;于注射前,注射后7、30和60 d分别进行兔眼光相干断层扫描(OCT)检查;于注射前,注射后14和60 d行闪光视网膜电图检查;于注射后60 d摘除眼球,各组取其中3只眼球制作石蜡切片行组织病理学观察,另取3只眼球,分离视网膜组织并采用透射电子显微镜观察视网膜超微结构变化。结果各注射组玻璃体腔注射后各时间点眼前节、后节均未见明显改变,其中0.125 mg rh-endostatin组、0.500 mg rh-endostatin组于注射后1 d各有1眼前房出现絮状渗出,分别于注射后7 d和14 d渗出完全吸收。OCT图像显示未出现异常光反射及形态改变。观察期间,各组注射前后各时间点间眼压值、a波振幅、b波振幅比较,差异均无统计学意义(F分组=0.134、0.101、0.476,F时间=1.709、2.479、1.706,均P>0.05)。光学显微镜及透射电子显微镜检查各组视网膜未见明显异常。结论兔眼玻璃体腔注射0.125~0.500 mg rh-endostatin是安全的。

Objective To investigate and evaluate the safety of intravitreal injection of recombinant human endostatin(rh-endostatin)with different concentrations in rabbit eyes.Methods Thirty healthy adult New Zealand white rabbits were enrolled with the right eyes selected as experimental eyes,and were randomly divided into five groups by random distribution of computer numbers,with 6 eyes in each group.The rabbits in the normal control group were given no treatment,and the rabbits in the normal saline group,0.125 mg rh-endostatin group,0.250 mg rh-endostatin group and 0.500 mg rh-endostatin group were treated with 100μl of normal saline,0.125 mg/100μl,0.250 mg/100μl and 0.500 mg/100μl rh-endostatin according to grouping,respectively.The anterior segment and fundus of the experimental eyes were examined using slit lamp biomicroscope and indirect ophthalmoscope,and the intraocular pressure(IOP)of the experimental eyes were measured with iCARE handheld tonometer before injection and 1 day,3,7,14,30 and 60 days after injection.Optical coherence tomography(OCT)examination was performed before the intravitreal injection and 7,30,and 60 days after injection,respectively.Flash electroretinogram was performed before intravitreal injection and 14 days and 60 days after injection.The rabbits were sacrificed by euthanasia at 60th day after injection.Three experimental eyes of each group were dissected and made into paraffin section,and histopathological staining was used to detect the retinal structural changes.The retinal tissue was separated from the other three study eyes in each group,and the transmission electron microscope was employed to observe the ultrastructural changes of the retina.All animal experiments were performed in adherence to the Regulations of the State and the Animal Center of Yangzhou University Medical College for the Use of Animals in Research.Results After intravitreal injection,no obvious anterior or posterior chamber change was observed by slit lamp microscopy in all groups at any time point.Flocculent seepage was observed in one eye of the 0.125 mg and 0.500 mg rh-endostatin group,respectively,which was then absorbed completely on the 7th and 14th day.OCT examination showed no abnormal light reflection or morphological changes in fundus of day after injection in all the groups.There was no significant difference in IOP,a-wave and b-wave amplitude among all the groups at different time points(F group=0.134,0.101,0.476;F time=1.709,2.479,1.706;all at P>0.05).Neither light nor electron microscopy showed any retinal damage in any group.Conclusions Intravitreal injection of rh-endostatin is safe at the dosage of 0.125-0.500 mg in rabbits.