摘要
背景:膜联蛋白V是近年来被作为肝细胞膜上存在的与乙型肝炎病毒感染相关受体研究的热点之一,并在胎盘组织中也有表达。目的:探讨膜联蛋白V在HBV宫内感染中的作用;揭示HBV进入胎盘组织细胞的方式,为HBV的宫内感染的机理提供理论依据。设计:随机对照实验。单位:泰山医学院。对象:选择2003-01/2004-12济南市妇幼保健院、泰安市中心医院、泰安市妇幼保健院收集的30例HBsAg阳性孕妇足月分娩的胎盘组织,同时收集母血分离血清。实验经过孕妇知情同意及医院伦理委员会批准。兔抗人膜联蛋白V亲和纯化抗体(第一抗体)、鼠抗人HBs mAb(第一抗体)、生物素化羊抗小鼠IgG(第二抗体)均购自武汉博士德生物工程有限公司。方法:用即用型SABC免疫组织化学染色试剂,检测30例HBsAg阳性的足月胎盘组织中,有18例检测到HBsAg,随机选取10例进行荧光双标染色,其染色主要过程为将待测石蜡切片标本常规脱蜡至水;抗原微波热修复;加入第一种一抗(兔抗人AnV亲和纯化抗体1:60,为单克隆抗体)置湿盒中,4℃冰箱过夜;第二种一抗(鼠抗人HBs mAb1:50),置湿盒中,4℃冰箱过夜;加入与第二种一抗相应的二抗(如生物素化的羊抗鼠IgG1:100):加入以PBS作适当稀释的第一种荧光抗体(如FITC-羊抗兔IgG1:50);加入第二种荧光抗体(Avidin-Cy3);缓冲甘油封固;激光共聚焦显微镜下观察并进行图像分析。用IPP4.5图像分析法,先进入IPP4.5系统的Image Analysis程序,然后调入分析文件进行分析。主要观察指标:乙型肝炎病毒感染的胎盘组织中HBsAg/膜联蛋白V的存在与分布情况。结果:选取10例进行荧光双标染色结合激光共聚焦显微镜检测胎盘组织上滋养层细胞乙型肝炎病毒/膜联蛋白V的存在与分布模式。胎盘组织中存在着丰富的膜联蛋白V,位于绒毛合体滋养层细胞,在基质细胞和血管内皮细胞都有膜联蛋白V的表达。并发�
BACKGROUND： Recently, one focus of research has been Annexin V （ANY） existing on hepatic cells membranes as a fundamental receptor related to hepatitis B virus （HBV） infection. Also its expression in placental tissues has been a matter of debate. The study of the relationships between placental cells infected with HBV and their AnV expression will be of great value in future prevention strategies and treatments.
OBJECTIVE： To investigate the presence of AnV in HBV infected human＇s placental cells and its potential role in HBV intrauterine transmission.
DESIGN： Randomized controlled experiment.
SETTING： Taishan Medical College.
MATERIALS ： Placental tissue was collected from HBsAg positive full term pregnant women （30 cases） admitted to Jinan Institute for Maternal and Child Health, Taian Central Hospital and Taian Institute for Maternal and Child Health from January 2003 to December 2004. Maternal serum was also obtained. Informed consents for participating in this study were obtained from all the involved pregnant women and this experiment was approved by the Hospital Ethics Committee. Rabbit-anti-human AnV purified affinity antibody （first antibody）, rat-anti-human HBs mAb （first antibody）, and biotinylated goat-anti-mouse IgG （secondary antibody） were supplied by Wuhan Boster Bioengineering Company.
METHODS： Using SABC immunohistochemical staining reagent, 18 HBsAg positive placentas were obtained from 30 HBsAg infected patients in full term pregnancy. These were considered as the positive group and the other 12 were used as negative controls. The staining process included dewaxing, dehydration of embedded slides and microwave antigen restoration. In the wet box, rabbit-anti-human AnV purified antibody （first antibody, 1：60, monoclonal antibody） was added on the slides and kept at 4 ℃ overnight. Rat-anti-human antibody HBs mAb（secondary antibody, 1：50） was added and kept at 4℃ overnight, after this procedure, biotinylated goat-anti-mouse IgG （1 �
出处
《中国组织工程研究与临床康复》
CAS
CSCD
北大核心
2007年第50期10213-10216,共4页
Journal of Clinical Rehabilitative Tissue Engineering Research
基金
山东省教育厅资助项目(JOIK15)~~
