摘要
目的 探讨可诱导共刺激分子（ICOS）/可诱导共刺激分子配体（ICOSL）信号通路对日本血吸虫病所致肝纤维化的影响.方法 建立ICOSL基因敲除（ICOSL-KO）日本血吸虫病小鼠模型78只和野生型C57BL/6J日本血吸虫病小鼠模型77只,分别于感染前（0周）及感染后4、7、12、16和20周采集小鼠血清,应用ELISA法检测血清中透明质酸（HA）和羟脯氨酸（HYP）的含量.应用免疫组织化学法检测小鼠肝脏中TGF-β1、α-平滑肌肌动蛋白（a-SMA）和Ⅰ型胶原蛋白（Collagen-Ⅰ）水平.分别应用HE染色法和胶原纤维Masson染色法动态观察感染小鼠肝脏内虫卵肉芽肿病变及纤维化程度.组间计量资料比较采用t检验,组间率的比较采用x2检验.结果 小鼠感染日本血吸虫后,随着时间的延长,其血清中HA、HYP水平逐渐升高.在感染后7、12、16和20周,ICOSL-KO小鼠血清HA水平均低于野生型小鼠[（161.32±15.44）比（186.01±21.24） ng/mL,t=2.528 2,P＜0.05;（166.73±18.18）比（231.39±20.12） ng/mL,t=4.342 4,P＜0.05;（193.58±21.06）比（252.51±25.29） ng/mL,t=4.003 9,P＜0.05;（253.98±24.53）比（310.88±23.86） ng/mL,t=3.718 0,P＜0.05].而ICOSL-KO小鼠的血清HYP水平则在感染后12、16和20周均低于野生型小鼠,两组比较差异均有统计学意义（均P＜0.05）.免疫组织化学检查结果显示,ICOSL-KO小鼠于感染后7～20周肝脏TGF-β1、α-SMA和Collagen-Ⅰ蛋白表达水平皆显著低于野生型小鼠（均P＜0.05）.肝组织HE染色结果显示,ICOSL-KO小鼠的肝脏虫卵肉芽肿体积小于野生型C57BL/6J小鼠（均P＜0.01）.Masson染色结果显示,ICOSL-KO小鼠肝纤维化程度显著低于野生型小鼠,两组纤维化程度评分的差异有统计学意义（均P＜0.05）.野生型C57BL/6J小鼠死亡率高于ICOSL-KO小鼠,在感染后20周,两组病死率的差异有统计学意义（55.84％比37.18％,x2=5.427,P＜0.05）.结论 感染日本血吸虫的ICOSL-KO小鼠肝纤维化程度和相关指标均显著降低,表明ICOS/ICOSL信号通路与感染日本血吸虫导致的肝纤维化密切相关.
Objective To analyze the effect of inducible costmulator （ICOS）/inducible costmulator ligand （ICOSL） signaling pathway on hepatic fibrosis in mice infected with Schistosoma japonicum.Methods Seventy-eight ICOSL knockout （ICOSL-KO） mice and 77 wild type C57BL/6J mice were used as experimental schistosomiasis model infected with Schistosoma japonicum.The sera of mice were collected on the day before infection （0 week）,and at 4,7,12,16 and 20 weeks post infection.Then,the concentrations of hyaluronic acid （HA） and hydroxyproline （HYP） in mice sera were measured by sandwich enzyme linked immunosorbent assay （ELISA） kits.The expressions of transforming growth factor β1 （TGF-β1）,α-smooth muscle actin （a-SMA） and Collagen-Ⅰ in livers from ICOSL-KO/wild type mice were assessed by immunohistochemical staining.The granulomatous pathology and fibrosis level in mice liver were dynamically observed by hematoxylin and eosin （HE） staining and Masson＆#39;s staining,respectively.The difference between groups was detected by t test or x2 test when appropriate.Results After infection with Schistosoma japonicum,the levels of HA and HYP were gradually increased.In ICOSL-KO mice,the levels of HA at 7,12,16 and 20 weeks post infection were all significantly lower than those in wild type mice [（161.32±15.44） vs （186.01±21.24） ng/mL,t=2.528 2,P〈0.05; （166.73±18.18） vs （231.39±20.12） ng/mL,t=4.342 4,P〈0.05; （193.58±21.06） vs （252.51±25.29） ng/mL,t=4.003 9,P〈0.05; （253.98±24.53） vs （310.88±23.86） ng/mL,t=3.718 0,P〈0.05].Similarly,HYP levels in ICOSL-KO mice at 12,16 and 20 weeks post infection were all significantly lower than those in wild type mice （all P〈0.05）.Immunohistochemical staining showed that TGF-β1,α-SMA and Collagen-Ⅰ expressions in liver of ICOSL-KO mice from 7 to 20 weeks post infection were all significantly lower than those of wild type mice （all P〈0.05）.HE staining showed,the volume of liver egg granulomas of ICOSL-KO mice was significantly smaller than that of wild type C57BL/6J mice （P〈0.01）.Furthermore,Masson＆#39;s staining showed that the level of hepatic fibrosis in ICOSL-KO mice was lower than that in wild type mice and the fibrosis scores were statistically different between two groups （all P〈0.05）.The mortality rate of the wilde type C57BL/6J mice was higher than that of ICOSL-KO mice.After 20 weeks of infection,the difference was statistically significant （55.84 % vs 37.18 %,x2 =5.427,P〈0.05）.Conclusions The degree of hepatic fibrosis and related indicators are obviously down-regulated in ICOSL-KO mice infected with Schistosoma japonicum.These findings suggest that ICOS/ICOSL signaling pathway has an important impact on the process of hepatic fibrosis caused by Schistosoma japonicum.
出处
《中华传染病杂志》
CAS
CSCD
北大核心
2015年第2期96-101,共6页
Chinese Journal of Infectious Diseases
基金
国家自然科学基金资助项目（81171603）
安徽理工大学博士基金资助项目（11046）
安徽理工大学中青年学术骨干培养工程资助项目（2013026）
