氟他胺诱导干眼模型大鼠角膜和泪腺组织病理变化及TNF-α、IL-1α的表达-《眼科新进展》

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Title:: Pathological changes of cornea and lacrimal glands in dry eye rat model induced by flutamide and expression of TNF-α and IL-1α

作者:: 彭玲; 蔡伟浩; 陆晓和; 钟彦彦; 易锐文; 510282　广东省广州市，南方医科大学珠江医院眼科

Author(s):: PENG Ling ; CAI Wei-Hao; LU Xiao-He; ZHONG Yan-Yan ; YI Rui-Wen; Department of Ophthalmology, Zhujiang Hospital of Southern Medical Univerisity , Guangzhou 510282 , Guangdong Province . China

关键词:: 干眼动物模型; 雄激素受体拮抗剂; 氟他胺; 肿瘤坏死因子-α; 白细胞介素-1α

Keywords:: dry eye arumal model; androgen receptor antagonist; flutamide ; TNF-α ; IL-lα

DOI:: 10.13389/j.cnki.rao.2016.0135

文献标志码:: A

摘要:: 目的　通过雄激素受体拮抗剂氟他胺溶液灌胃建立大鼠干眼模型，观察干眼大鼠角膜及泪腺组织病理变化及ＴＮＦ-α、ＩＬ-１α的表达。方法　取３２只ＳＰＦ级ＳＤ雌性大鼠随机分为４组，氟他胺高、中、低浓度组和对照组，每组８只，每天分别用浓度为１６ｍｇ·ｋｇ－１、１３ｍｇ·ｋｇ－１、１０ｍｇ·ｋｇ－１氟他胺溶液和１ｍＬ生理盐水灌胃。给药后０周、２周、４周、８周、１６周、２０周分别检测各组基础泪液分泌试验（ＳｃｈｉｒｍｅｒⅠｔｅｓｔ，ＳⅠｔ）值、泪膜破裂时间（ｂｒｅａｋ-ｕｐｔｉｍｅ，ＢＵＴ）及角膜荧光素钠染色情况。给药后０周、１０周、２０周采用放射免疫法检测各组血清睾酮浓度。给药后２０周处死大鼠取各组角膜与泪腺组织行病理学观察，免疫组织化学方法检测各组ＴＮＦ-α、ＩＬ-１α的表达情况。结果　给药后４周时高、中浓度组ＳⅠｔ值开始减小，与对照组比较差异均有统计学意义（均为Ｐ＜０．０５）。８周时高、中、低浓度组的ＳⅠｔ值均小于对照组（均为Ｐ＜０．０５），２０周时高（３．２３±０．３１）ｍｍ、中（３．９８±０．３６）ｍｍ、低（４．７３±０．１８）ｍｍ浓度组的ＳⅠｔ值进一步减小，且均较对照组（９．６０±０．６０）ｍｍ下降大于５０％（均为Ｐ＜０．０５），高、中、低浓度组两两比较差异均有统计学意义（均为Ｐ＜０．０５）。给药后４周时高浓度组大鼠即出现ＢＵＴ缩短，８周时，高、中、低浓度组的ＢＵＴ均短于对照组（均为Ｐ＜０．０５），２０周时高（２．４０±０．６１）ｓ、中（４．３２±０．６８）ｓ、低（６．１０±０．８１）ｓ浓度组的ＢＵＴ较对照组（９．１２±０．７３）ｓ进一步缩短，各组之间两两比较差异均有统计学意义（均为Ｐ＜０．０５）。给药后４周时高、中浓度组开始出现角膜上皮点状荧光素钠着色，１６周时中、低浓度组出现小片状着色，高浓度组可见角膜上皮弥漫性大片状着色，２０周时各组染色评分两两比较差异均有统计学意义（均为Ｐ＜０．０５）。给药后２０周时睾酮浓度高浓度组（５．０８±０．２１）ｐｇ·ｍＬ－１、中浓度组（４．２３±０．１６）ｐｇ·ｍＬ－１、低浓度组（３．５８±０．２４）ｐｇ·ｍＬ－１较对照组（２．２４±０．１５）ｐｇ·ｍＬ－１不同程度升高，与对照组比较差异均有统计学意义（均为Ｐ＜０．０５），高、中、低浓度组两两比较差异均有统计学意义（均为Ｐ＜０．０５）。ＨＥ染色显示高、中、低浓度组角膜出现不同程度鳞状上皮层次增加，基底细胞排列紊乱，基质层水肿；高、中、低浓度组泪腺上皮细胞小而密集，腺泡萎缩融合，结构紊乱，高浓度组改变最为显著。免疫组织化学结果显示高、中、低浓度组角膜与泪腺上皮细胞中ＴＮＦ-α、ＩＬ-１α阳性表达细胞数均高于对照组（均为Ｐ＜０．０５）。结论　雄激素受体拮抗剂氟他胺溶液灌胃可以成功诱导混合型干眼模型，干眼大鼠角膜和泪腺上皮细胞发生病理学改变，炎性因子ＴＮＦ-α、ＩＬ-１α的表达明显增高。

Abstract:: Objective To establish the dry eye rat model induced by flutanude, observe the pathological changes of cornea and lacrimal glands and the expression of tumor necrosis factor-e/ ( TNF-e/ ) , interleukin-le/ ( IL-Ie/ ) . Methods Thirty-two specific pathogen free SD rats were randomly divided into four groups ( 8 cases in each group) : high , medium .low concentration group and control group , were respectively irrigated stomach with flutamide solution on concentration of 16 mg ’ kg", 13 mg ’ kg ’l , 10 mg . kg -l and I mL saline solution every day. Each group on week 0 .2 .4 ,8 . 16 . 20 were evaluated the Schirmer I test ( SIt) .tear break-up time ( BUT) and corneal fluorescence staining. Each group on week 0 , 10 ,20 were measured serum testosterone levels by radioimmunoassay. The animals in each group were sacrificed on week 20 . the cornea and lacrimal gland were taken to perform the histopathological observation , and the immunohistochemistry method was used to detect the expression of TNF-e/ and IL-ld?? Results On week 4 . SIt values of high , medium concentration groups began to decrease, there were statistical difference compared with control group ( all P < 0. 05 ) . On week 8 , SIt values of high , medium .low concentration groups were all lower than that of control group ( all P < 0. 05 ) ; On week 20 . SIt values of high , medium .low concentration group were (3. 23 +0. 31 ) mm . ( 3. 98 + 0. 36) mm , ( 4. 73 + 0. 18 ) mm , respectively, which were further shortened, compared with control group ( 9. 60 + 0. 60 ) mm all decreased more than 50% . and there were significant differences among high , medium , low concentration groups ( all P < 0. 05 ) . On week 4 . BUT of high concentration group began to shorten; On week 8 . BUT of high, medium . low concentration groups were shorter than control group ( all P < 0. 05 ) ; On week 20 , BUT of high , medium ,low concentration groups were ( 2. 40 + 0. 61 ) seconds, ( 4. 32 + 0. 68 ) seconds , ( 6. 10 + 0. 81 ) seconds , which were all lower than control group ( 9. 12 + 0. 73 ) seconds . there were statistical differences among each group ( all P < 0. 05 ) . On week 4 . the corneal epithelial punctate fluorescein staining of high, medium concentration groups were positive ; On week 16 . the diffuse large sheet high staining appeared in concentration group , and the small pieces of corneal epithelial stairung was rn medium . low concentration groups ; On week 20 . the stairung scores of each group were significant different ( all P < 0. 05 ) . On week 20 , the serum testosterone level in high,medium ,low concentration groups were (5. 08 + 0. 21 ) pg . mL -l , ( 4. 32 + 0. 16) pg . mL ’l . ( 3. 58 + 0. 24 ) pg . mL -l , which were higher than that in control group ( 2. 24 + 0. 15 ) pg . mL -l ( all P < 0. 05 ) , and there were statistical differences among high , medium , low concentration groups ( all P < 0. 05 ) . HE staining showed that the corneal squamous epithelial cell layers of high ,medium , low concentration group were increased.the basal cells were disorder . and the stromal layer was edema and thickening , the lacrimal gland epithelial cells were small and dense with structural disorder, acinar atrophy was appeared, the high concentration group got the most significant changes. Immunohistochemistry showed the expression of TNF-e/ . IL-I eL in lacrimal gland and corneal epithelial cells of high,medium , low concentration group were significantly higher than that of control group ( all P < 0. 05 ) . Conclusion Irrigated stomach with androgen receptor antagonist flutamide solution can induce the dry eye model. and the pathological changes are seen rn cornea and lacrimal glands of rat . the expression of TNF-e/ , IL-I eL are obvious increased.

《眼科新进展》[ISSN:1003-5141/CN:41-1105/R]

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