去势雄兔干眼模型泪腺上皮细胞凋亡和相关基因表达及局部环孢霉素A的治疗效果

Lacrimal epithelial cell apoptosis and related gene expression in castrated male rabbits with experimental xerophthalmia and the effect of dropping cyclosporine

目的观察去势雄兔泪液分泌、泪膜稳定性变化、泪腺细胞凋亡及相关基因表达和局部应用环孢霉素A的治疗效果。方法健康成年雄兔18只,随机分为正常组、干眼组和治疗组,每组6只。后2组行双侧睾丸及附睾切除制作干眼模型。治疗组去势后4周给予1%环孢霉素A点双眼,分别于去势前和去势后1、2、3、4、6、8、12周检测各组Shirmer试验、泪膜破裂时间、虎红染色;同时应用TUNEL及免疫组织化学法研究泪腺细胞凋亡及相关基因Fas、FasL的表达。结果实验6周后治疗组泪液分泌增加,泪膜破裂时间延长,与干眼组相比差异有非常显著性(t=9.08,P<0.01;t=6.93,P<0.01)。12周后,治疗组Fas、FasL阳性目标平均光密度值较干眼组减少,差异具有显著性(P<0.05);泪腺腺泡和导管上皮细胞中Fas、FasL蛋白阳性平均光密度值与细胞凋亡数都成正相关(r=0.737,P<0.01;r=0.667,P<0.01)。结论去势后雄兔睾酮水平明显低下是造成干眼的原因之一,细胞凋亡可能是发病机制之一;泪腺上皮细胞Fas、FasL的增加与促进细胞凋亡有关;局部环孢霉素A治疗对该型干眼有效。

Objective To evaluate the effect of dropping cyclosporine（CsA） on tear secretion , tear film stability, the apoptosis of lacrimal cells and related gene expression in castrated male rabbits. Methods Eighteen adult male rabbits were divided randomly into normal group, dry eye group and treatment group. Their testicel and parastata cirsoides were exseeted to make dry eye model for the last two groups. The rabbits in treatment group were given 1% eyclosporine（CsA） in both eyes after four weeks. Sehirmer＇ test, tear film break-up time（BUT）, rose bangle staining were performed in the eyes of every rabbit before and after experiment at 1,2,3,4,6,8, 12 weeks. Meanwhile, the lacrimal epithelial cell apoptosis was detected by TUNEL and related gene Fas/FasL expression was measured by immunohistochemical staining. Results After 6 weeks of the experiment, the tear secretion increased and the BUT extended in treatment group. There was a significant difference between dryeye group and treatment group （ t = 9.08, P 〈 0. 01 ; t = 6.93, P 〈 0. 01）. The counts of apoptotic epithelial cells in lacrimal gland of xerophthalmia model were significantly higher than those in dry eye group. After 12 weeks, as compared with that in dry eye group, the Fas/FasL positive objective average optical density significantly decreased in treatment group （ P 〈 0. 05）. There was a significant positive correlation between the expression of Fas/FasL and the apoptotic cells numbers（ r = 0. 737, P 〈 0. 01; r =0. 667, P 〈 0.01 ）. Conclusion The low eststemneon level of castrated male rabbits is one of the main reasons of dry eye. The apoptosis may be one of the mechanisms. Increased expression of Fas and FasL in lacrimal gland could promote apoptosis. Cyclosporine administrated locally has an effect on this kind of dry eye.