人源细粒棘球蚴囊壁与囊液中宿主蛋白的鉴定与分析  

Identification and analysis of host proteins in the cyst wall and cyst fluid of Echinococcus granulosus in human infection

作　　者：李涛[1] 李子华[2,3,4] 张翠影 赵巍 LI Tao;LI Zihua;ZHANG Cuiying;ZHAO Wei(General Hospital of Ningxia Medical University,Yinchuan 750004,China;School of Basic Medicine of Ningxia Medical University,Yinchuan 750004,China;Science and Technology Research Center of Ningxia Medical University,Yinchuan 750004,China;Ningxia Key Laboratory of Common Infectious Diseases,Yinchuan 750004,China)

机构地区：[1]宁夏医科大学总医院,银川750004 [2]宁夏医科大学基础医学院,银川750004 [3]宁夏医科大学科学技术研究中心,银川750004 [4]宁夏常见传染病重点实验室,银川750004

出　　处：《中国寄生虫学与寄生虫病杂志》2023年第6期677-682,共6页Chinese Journal of Parasitology and Parasitic Diseases

基　　金：银川市科技支撑项目(2023SF10);宁夏医科大学2023年校级科研项目重点项目(XZ2023027)。

摘　　要：目的通过对细粒棘球蚴囊壁及囊液中的宿主蛋白成分进行鉴定,进一步阐明细粒棘球绦虫与宿主之间相互作用的现象。方法收集2022年宁夏医科大学总医院肝胆外科10例细粒棘球蚴病患者手术摘除的具有活性的完整棘球蚴包囊,同时取患者血样。无菌抽取子囊中无色透明的囊液,离心收集上清液;子囊内壁反复冲洗后剪碎,裂解匀浆,收集上清液。用二喹啉甲酸(BCA)蛋白定量试剂盒提取囊液和囊壁全蛋白,经过十二烷基磺酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)电泳后,切取囊壁全蛋白泳道显著的蛋白条带,进行质谱检测,进行蛋白质免疫印迹(Western blotting)分析。对患者血清和囊液的生化成分浓度进行定量检测。结果10例细粒棘球蚴病患者的肝脏功能为Child-Pugh A或B级。病灶直径为2.9~16.3 cm,为WHO-IWGE分型的CE2(多子囊型)或CE3(内囊塌陷型)。SDS-PAGE电泳分析结果显示,棘球蚴囊壁蛋白在相对分子质量(Mr)为70000、50000、25000处显示出清晰的蛋白条带,囊液蛋白在Mr 70000显示出清晰的蛋白条带。蛋白质谱与Western blotting结果均显示,相对分子质量在约Mr 70000的蛋白中包含人血白蛋白,约Mr 50000的目标蛋白为人IgG重链,约Mr 25000的目标蛋白为人IgG轻链。各生化指标中,囊液和血清中的中K+浓度分别为(5.53±0.86)mmol/L和(3.92±0.33)mmol/L,白蛋白浓度分别为(0.57±0.46)g/L和(39.42±2.77)g/L,胆固醇浓度分别为(0.03±0.02)mmol/L和(3.79±1.53)mmol/L,三者在囊液和血清中的浓度差异有统计学意义(t=5.56、43.71、7.74,P<0.05)。结论宿主来源的血清白蛋白和抗体是细粒棘球蚴囊壁和囊液中含量较高的宿主蛋白。Objective To further elucidate the interaction between Echinococcus granulosus and the host by identifying the host protein components in the cyst wall and cyst fluid of E.granulosus.Methods The active and intact echinococcal cysts removed surgically from 10 patients with E.granulosus were collected from the Department of Hepatobiliary Surgery,Ningxia Medical University General Hospital in 2022.Blood samples were also taken from the same patients.The transparent cyst fluid in the ascus was extracted aseptically,and centrifuged to collect the supernatant;the inner wall of the ascus was repeatedly washed,cut into pieces,lysed and homogenised to collect the supernatant.The bicinchoninic acid(BCA)protein quantification kit was used to extract the cyst fluid and cyst wall total protein.The cyst wall total protein lane was cut out after sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE)electrophoresis.The significant protein bands were analysed by mass spectrometry and Western blotting analysis.Quantitative analysis was performed to determine the biochemical component concentrations in the patient serum and cyst fluid.Results The liver function of 10 patients with echinococcosis granulosus was Child-Pugh A or B.The diameter of the lesion ranged from 2.9 to 16.3 cm,which were classified as CE2(multiple subcysts)or CE3(collapsed internal capsule)according to the WHO-IWGE classification.SDS-PAGE electrophoresis analysis results suggested that the echinococcal cyst wall protein showed clear protein bands at the relative molecular mass(Mr)of 70000,50000,and 25000,and the cyst fluid protein showed clear bands at Mr 70000.Protein bands.The protein spectrum and Western blotting results showed that the protein with a relative molecular mass of approximately Mr 70000 contained human albumin,the target protein with a relative molecular mass of approximately Mr 50000 was a human IgG heavy chain,and the target protein with a relative molecular mass of approximately Mr 25000 was a human IgG light chain.Among the var

关 键 词：细粒棘球绦虫 白蛋白 抗体 囊壁 囊液 

分 类 号：R532.32[医药卫生—内科学]