兔眼玻璃体腔连续注射不同剂量更昔洛韦视网膜毒性研究

Retinal toxicity study of different-dose intravitreal ganciclovir in albino rabbit

目的初步探索健康兔眼玻璃体腔单次注射更昔洛韦的安全剂量范围,并观察兔眼玻璃体腔连续注射不同剂量更昔洛韦的视网膜毒性。方法将10只健康新西兰无色素兔平均分为5组,每组各2只兔。每组分别往兔眼玻璃体腔注射更昔洛韦1 mg/0.025 ml、2 mg/0.025 ml、5 mg/0.025 ml、10 mg/0.025 ml及0.025 ml生理盐水(对照组)。干预1周后对兔眼行超广角眼底照相、光相干断层扫描(OCT)及全视野视网膜电图(ERG)检查,暗适应条件下测定视杆细胞-视锥细胞最大混合反应(Max-R),明适应条件下测定视锥细胞反应(Cone-R)和30 Hz闪烁反应(30 Hz-R)。将24只健康新西兰无色素兔随机分为小剂量实验组、小剂量对照组和大剂量实验组、大剂量对照组,每组6只兔,以右眼为实验眼。大剂量实验组兔眼连续注射更昔洛韦2 mg/0.025 ml,1次/周,共4次。小剂量实验组兔眼注射1 mg/0.025 ml更昔洛韦,诱导期为2次/周,共4次;维持期为1次/周,共2次。大剂量对照组、小剂量对照组兔眼玻璃体腔分别注射0.025 ml生理盐水。每次注射前1 d及末次注射后1周行全视野ERG检查,暗适应条件下测定Max-R,明适应条件下测定Cone-R和30 Hz-R;注射前1 d及末次注射后1周行OCT检查;末次注射后1周处死各组实验兔行苏木精-伊红染色,光学显微镜下观察视网膜结构。不同时间Max-R、Cone-R的a波和b波振幅以及30 Hz-R振幅比较行两独立样本非参数检验。结果健康兔眼玻璃体腔单次注射1 mg或2 mg更昔洛韦1周后未见超广角眼底照相、OCT、ERG异常结果。健康兔眼玻璃体腔单次注射5 mg更昔洛韦1周后可见眼底血管堵塞、视网膜前出血,ERG可见Max-R、Cone-R的a波和b波振幅以及30 Hz-R振幅略有下降。健康兔眼玻璃体腔单次注射10 mg更昔洛韦1周后还可见视网膜坏死、渗出性改变,OCT提示坏死区域视网膜组织水肿、层次不清,ERG可见Max-R、Cone-R的a波和b波振幅以及30 Hz-R振幅明显下降。连续注射更昔洛韦1周后,小剂量实验组、大剂量实验组兔眼视网膜均未见出血、渗出;视网膜各层结构清晰连续,未见视网膜水肿或其他结构异常。与大剂量对照组、小剂量对照组比较,连续注射更昔洛韦1周后大剂量实验组、小剂量实验组兔眼ERG的暗适应Max-R a波(Z=-0.160、0.000)和b波(Z=-0.321、0.000)、明适应Cone-R a波(Z=-0.641、-0.641)和b波(Z=-0.321、-0.160)振幅以及30 Hz-R振幅(Z=-0.321、-0.160)差异均无统计学意义(P>0.05)。光学显微镜观察发现,与对照组相比,大剂量实验组、小剂量实验组兔眼视网膜各层组织结构均无明显差异。结论玻璃体腔连续注射更昔洛韦1、2 mg均未对健康兔眼造成明显视网膜毒性。

Objective:To explore safe dosage of single intravitreal injection of ganciclovir(IVG)in healthy rabit eyes,and to explore retinal toxicity of different dosage of ganciclovir after continues intravitreal injection into the vitreous cavity of healthy albino rabbit eyes.Methods:Ten healthy New Zealand albino rabbits were divided into 5 groups with 2 rabbits in each group.Each group was injected with 1 mg/0.025 ml,2 mg/0.025 ml,5 mg/0.025 ml,10 mg/0.025 ml ganciclovir or 0.025 ml saline(control group).After 1 week of intervention,rabbits were examined by ultra-wide-angle fundus photography,optical coherence tomography(OCT)and full field electroretinogram(ERG).The maximum mixed response of rod and cone cells(Max-R)was measured under dark adaption conditions,cone response(Cone-R)and 30 Hz flicker response(30 Hz-R)were measured under light adaption conditions.Twenty-four healthy New Zealand albino rabbits were randomly divided into a low-dose experimental group,a low-dose control group,a high-dose experimental group,and a high-dose control group,with 6 rabbits in each group,with the right eye as the experimental eye.The rabbits in the high-dose experimental group were continuously injected with ganciclovir 2 mg/0.025 ml,once a week,for a total of 4 times.The rabbits in the low-dose experimental group were injected with 1 mg/0.025 ml ganciclovir,the induction period was 2 times/week,a total of 4 times;the maintenance period was 1 time/week,a total of 2 times.The rabbits in the high-dose control group and the low-dose control group were injected with 0.025 ml normal saline into the vitreous cavity respectively.Full-field ERG examination was performed 1 day before each injection and 1 week after the last injection.Max-R was measured under dark-adapted conditions,and Cone-R and 30 Hz-R were measured under light-adapted conditions.OCT was recorded before the first injection and one week after the last injection.One week after the last injection,the experimental rabbits in each group were sacrificed for hematoxylin-eosin staining,and the retinal structure was observed under a light microscope.The comparison of a-wave and b-wave amplitude of Max-R,Cone-R and 30 Hz-R amplitude at different time was performed by two independent sample nonparametric test.Results:There were no abnormal results of fundus photography,OCT and ERG after single intravitral injection of 1 mg or 2 mg ganciclovir.One week after single 5 mg IVG,fundus photography of rabbits showed vascular occlusion and preretinal hemorrhage and ERG showed slight decrease of amplitude of Max-R,Cone-R and 30 Hz-R.One week after single 10 mg IVG,retinal necrosis and exudative changes were also observed.OCT showed edema and unclear retinal structure in the necrotic area.ERG showed significant decrease of amplitude of Max-R,Cone-R and 30 Hz-R.After continuous IVG in high dose and low-dose experimental group,the amplitude of Max-R a wave(Z=-0.160,0.000)and b wave(Z=-0.321,0.000),Cone-R a wave(Z=-0.641,-0.641)and b wave(Z=-0.321,-0.160),and 30 Hz-R(Z=-0.321,-0.160)showed no difference compared to control group.No histologic evidences of retinal microstructure abnormalities were found in both groups.OCT and fundus photography before and after the intervention did not show any difference,either.Conclusion:There was no retinal toxicity of continuous 1 mg or 2 mg IVG recorded in albino rabbits.