摘要
目的分析亚洲带绦虫实验感染乳猪后囊尾蚴寄生处肝脏组织半胱氨酸天冬氨酸蛋白酶3(Cysteine aspartyl proteinase 3,Caspase-3)的表达变化情况,为探讨Caspase-3在亚洲带绦虫感染致乳猪肝损伤中的作用提供基础资料。方法将采自贵州省都匀市良亩乡的亚洲带绦虫孕节解剖后,以生理盐水反复清洗并离心收集虫卵。20d龄约克施格杂交乳猪12头随机分为实验组和对照组各6头,实验组以定量虫卵15万个/头灌胃感染,于感染后第15d和第75d解剖乳猪,取实验组囊尾蚴寄生处肝组织和对照组对应部位肝组织,运用实时荧光定量PCR和免疫印迹方法分别定性定量分析Caspase-3在实验组和对照组中mRNA水平、蛋白水平的表达,并进一步运用免疫组织化学方法对Caspase-3在实验组和对照组中的蛋白表达水平和蛋白表达部位进行分析和定位。结果感染后第15d,实时荧光定量PCR结果显示实验组Caspase-3mRNA水平低于对照组(P=0.011),免疫印迹结果和免疫组化结果均显示实验组Caspase-3蛋白表达量低于对照组(P=0.008,P=0.004);此外,实验组感染后第75d的Caspase-3mRNA水平、蛋白表达量均高于实验组感染后第15d的样本(P=0.018,P=0.003,P=0.002);免疫组织化学结果显示Caspase-3阳性染色主要定位在肝细胞胞质中,呈黄色或棕黄色。感染后第75d实验组和对照组各项结果差异均无统计学意义。结论 Caspase-3在亚洲带绦虫感染早期的乳猪肝组织中低表达,而感染晚期正常表达,提示Caspase-3可能参与了亚洲带绦虫致乳猪早期肝损伤的调节。
In order to provide substantial scientific information for exploring the mechanism of porcine liver injury caused by Taenia asiatica fT. asiatica）, the expression of Cysteine aspartyl proteinase 3 （Caspase-3） from liver tissues of porkets that were experimentally infected by T. asiatica was examined. The T. asiatica adults were collected from the taeniasis patients in Duyun, Guizhou Province and identified biologically. The eggs were harvested from gravid proglottids and prepared by repeated washing and centrifugation. Twelve 20-days old Yorkshire and Seghers hybrid porkets were randomly divided into experimental and control groups as six pigs per group. The experimental group was orally administrated with 1.5 × 10^6 eggs per porket at day 0 post-infection. The porkers of both groups were sacrificed on the day 15 and day 75 post-infection （three pigs per time point） respectively, and liver samples were collected for further experiments. Quantitative real-time polymerase chain reaction method was employed to detect the mRNA levels of Caspase-3, and western blotting and immunohistoehemistry methods were performed to detect the level of Caspase-3 expression in both groups. At the day 15 post-infection, the mRNA level and expression level of Caspase-3 of the experimental group were signifi- cantly decreased, comparison with the control group （P= 0. 011, P=0. 008 and P=0. 004 respectively）. It was posi- tive with Caspase-3 when yellow or brown signal appeared in the cytoplasm of liver cells by immunohistochemistry. How- ever, at the day 75 post-infection, the mRNA level and expression level of Caspase-3 of the experimental group were dramatically similar to the control group. Furthermore, in the experimental group, the mRNA level and expression level of Caspase-3 were significantly increased at day 75 post-infection than day 15 post-infection （P= 0. 018, P= 0. 003 and P= 0. 002 respectively）. These results suggested that Caspase-3 might he involved into the regulation of the damage of porcine liver induced by T. asiatica challenge at the early infection stage and have on effect to the hepatic injury because of the dramatic recovery of Caspase-3 at the consequent infection stage.
出处
《中国人兽共患病学报》
CAS
CSCD
北大核心
2017年第4期326-331,342,共7页
Chinese Journal of Zoonoses
基金
国家自然科学基金项目(No.81160205)
贵州省优秀科技教育人才省长资金项目(黔省专合字[2008]55号)
贵州省留学人员科技创新项目(黔人项目资助合同[2015]1号)联合资助~~
