摘要
目的建立SD大鼠急性内毒素性葡萄膜炎(endotoxin induced uveitis,EIU)模型,研究益母草碱(leonurine,SCM-198)是否对脂多糖(lipopolysaccharide,LPS)所致眼部结构、功能损伤具有保护作用,并探讨其作用机制。方法体重180~200 g正常雄性SD大鼠36只,随机分为3组。实验组:又分为4个亚组,分别以每日每千克体重10、20、40、80 mg的SCM-198生理盐水溶液灌胃,共21天;阴性对照组:每日以0.9%Na Cl溶液10 m L/kg体重灌胃;阳性对照组:以每日每千克体重0.5 mg的地塞米松(dexamethasone,DEX)磷酸钠生理盐水溶液灌胃。组间饲养无差异,每7天测体重。第18天灌胃后,全部大鼠全麻并充分暗适应后行全视网膜电图(electroretinogram,ERG)检测;第21天灌胃后以1 g/L的伤寒沙门氏菌(S.typhi)LPS生理盐水溶液0.1 m L于全部大鼠左足底注射造模,造模后24 h行全麻下ERG检测,裂隙灯下行双眼炎症评分,取左眼房水-80℃冻存后处死动物,取右侧眼球固定后石蜡包埋切片,HE染色,光学显微镜下观察,免疫组织化学(immunohistochemistry,IHC)染色检测肿瘤坏死因子-α(tumour necrosis factor-α,TNF-α)、细胞间黏附分子-1(intercellular cell adhesion molecule-1,ICAM-1)表达水平;对留取的房水样本用BCA法测定蛋白浓度,Western blot检测房水TNF-α、白细胞介素-1β(interleukin-1β,IL-1β)、IL-6、ICAM-1水平;以SPSS 19.0软件处理所得数据,综合分析SCM-198对EIU模型大鼠的保护作用。结果实验组、阴性对照组第21天时体重均显著高于阳性对照组(P<0.05)。实验组裂隙灯下炎症评分低于阴性对照组、高于阳性对照组;实验组病理切片炎症程度介于阴性对照组和阳性对照组之间;SCM-198 20 mgkg-1d-1组造模后0.01 cd下ERG a波波幅显著低于造模前(P<0.05),亦低于其他组造模后;SCM-198 10 mgkg-1d-1组房水TNF-α、IL-6和IL-1β表达低于其他各组。结论 SCM-198灌胃对EIU模型大鼠具有抗炎保护作用,且常规剂量下未见明显全身不良反应;可在一定程度上保护视网膜功能,减轻炎症反应及葡萄膜结构损伤;NF-κB在其抗炎效应中地位重要。SCM-198对炎症相关性眼病治疗具有潜在价值,但最佳给药方式、剂量等仍需进一步探索。
Objective To investigate whether the protective effects of leonurine（SCM-198） against endotoxin induced uveitis（EIU） of SD rats caused by lipopolysaccharide（LPS） was existing,and discuss the underlying mechanisms. Methods Thirty-six normal healthy male SD rats were divided into 3 groups randomly with the same baseline bodyweight and feeding conditions. All rats received intragastric administration every day. The experimental group was devided into 4 subgroups,rats in these subgroups received SCM-198 intragastric administration by as the dose of 10,20,40 and 80 mg/kg bodyweight per day,rats in the negative control group received intragastric administration of normal saline 10 m L/kg per day,rats in the positive control group received intragastric administration of dexamethasone（DEX） 0. 5 mg/kg bodyweight per day. All rats received a 21-day-intragastric administration. The body weight of all rats was monitored every 7 days. The electroretinogram（ERG） examination was taken in the 18 th day. All rats received a 100 mg S. typhi LPS intraperitoneal injection after the 21 st intragastric administration. Twenty-four hours later,following anaesthesia,all rats received another ERG examination,and inflammation was scoring under microscope by 2 experienced ophthalmologists,after that the aqueous humor of all rats was collected from the left eye. The aqueous humor was kept in-80 ℃ immediately. Then the rats were sacrificed and the right eyes were immediately enucleated to finish the HE staining and immunohistochemistry（IHC） staining examination of tumor necrosis factor-α（TNF-α） and intercellular cell adhesion molecule-1（ICAM-1）. The total amount of protein in aqueous humor was detected by BCA test. Western blot was used to examine the expression of TNF-α,interleukin-1β（IL-1β）,IL-6 and ICAM-1. All data was analyzed by SPSS 19. 0,and differences were considered significant at P〈 0. 05. Results The body weight of the rats in positive control group was significantly lower（P 〈0. 05） than the experimental group and the negative control group after the 21-day-intragastric administration. The inflammatory score of experimental group was lower than that of the negative control group,but higher than the score of positive control group. The HE staining sections showed the similar results. The a wave of ERG in 0. 01 cd of rats received 20 mg/kg SCM-198 daily intragastric administration after LPS injection was significantly lower than that before the LPS injection（P 〈0. 05）,also lower than other groups after LPS injection. The expression of TNF-α,IL-6 and IL-1β in the aqueous humor of the rats in the subgroup of SCM-198 10 mg/kg daily intragastric administration was lower than other groups. Conclusions Intragastric administration of SCM-198 has protective effect against endotoxin induced uveitis in SD rats without obvious adverse reaction,which could alleviate the imflammatory reaction and the damage to the uvea construction. NF-κB plays an important role in the reaction. Thus,SCM-198 is a candidate potent compound with potential therapeutic applications in inflammation associated eye diseases. While the best mode and dose of administration should be further investigated.
作者
熊佳伟
顾纪锋
石碗如
吴甦潜
麦尔哈巴.肖开提
朱依谆
莫晓芬
XIONG Jia-wei;GU Ji-feng;SHI Wan-ru;WU Su-qian;Marhaba Shewket;ZHU Yi-zhun;MO Xiao-fen(Department of Ophthalmology;Department of Pharmacy, Eye and ENT Hospital , Hehai University, Shanghai 200031 , China;Department of Pharmacy, Macao University of Science and Technology, Macao 999078, China;Department of Pharmacology and Biochemisty, School of Pharmacy, Fuclan University, Shanghai 201203, China)
出处
《复旦学报（医学版）》
CAS
CSCD
北大核心
2018年第3期291-296,329,共7页
Hehai University Journal of Medical Sciences
基金
上海市科委生物医药处中医引导科技项目(13401906900)
国家自然科学基金面上项目(81570842)~~
