Caspase 3抑制剂AcD-EVD-CHO对实验性近视眼视网膜细胞凋亡的治疗

The treatment of retinal apoptosis by Caspase 3 inhibitor Ac-DEVD-CHO in experimental myopia

目的 研究鸡近视眼视网膜细胞凋亡的分子机制及Caspase3抑制剂Ac- DEVD -CHO的治疗作用。方法 眼睑缝合法建立鸡的近视眼模型。玻璃体内注射Ac -DEVD- CHO后按预定时间检影验光,摘眼球,测量眼球外径,并拍眼底照片,用电镜、末端脱氧核糖核苷酸连接酶介导生物素化脱氧尿苷酸末端标记(TUNEL)技术及流式细胞术检测视网膜的凋亡细胞。用免疫组化、流式细胞术和色敏比色法分析视网膜Caspase3蛋白表达及其活性变化。结果 眼睑缝合12周时缝合眼与对照眼相比屈光度加深,眼球外径增大。缝合眼中出现漆裂纹样眼底改变(发生率为45 .56% ),并在缝合眼视网膜内、外核层发现凋亡细胞,凋亡率高于对照眼(P<0 .05),且视网膜Caspase3蛋白及活性均增高。玻璃体内注射Ac -DEVD- CHO后,缝合眼视网膜细胞的凋亡率和Caspase3蛋白及其活性均减少,而且Ac -DEVD -CHO抑制视网膜细胞凋亡的作用随剂量的加大而增强。结论 Caspase3在鸡近视眼视网膜细胞的凋亡过程中发挥重要作用,Ac- DEVD- CHO通过阻断Caspase3作用而有效抑制视网膜细胞凋亡。

Objective To investigate the mechanism of retinal apoptosis in chick experimental myopia and the therapy of Caspase 3 inhibitor Ac-DEVD-CHO. Methods Chick myopia was induced by lid-suture. After Ac-DEVD-CHO had been injected into vitreous, myopia was confirmed by optometry. Subsequently, chick eyeballs removed were measured its extro-dimensions, and the change of fundus were observed. Retinal apoptotic cells were determined by electron microscopy, TUNEL technique and flow cytometry. Retinal Caspase 3 proteins and its activities were measured by immunohistochemistry, flow cytometry and a colorimetric method using Ac-DEVD-pNA as a substrate. Results Ocular diopters and extro-diameters in all sutured eyes remarkly increased in comparison with its control eyes at 12 weeks after lid-suture. Lacquer crack lesions were found in 41of 90 sutured eyes(45.56%). Apoptotic cells were observed in retinal outer and inner nuclear layer of the sutured eyes, and its apoptotic rate was significantly more than in control eyes( P <0.05). Moreover, the expression of retinal Caspase 3 proteins and its activities were increased. After Ac-DEVD-CHO was injected into vitreous, the decrease of retinal apoptotic rate, Caspase 3 proteins and its activities was found. The treated effect of Ac-DEVD-CHO was in a dose dependent manner. Conclusions Caspase 3 plays an important role in retinal apoptosis of chick myopia. Ac-DEVD-CHO can effectively ameliorate retinal apoptosis by blocked the effect of Caspase 3.