目的检测HBV preS1蛋白及ASGPR的亚细胞定位及在肝组织中的分布情况,为研究二者的生物学功能提供依据。方法激光共聚焦显微镜检测HBV preS1蛋白及ASGPR在HepG2细胞中的亚细胞定位情况以及二者是否存在共定位;免疫组织化学技术检测HBV preS1蛋白和ASGPR在肝组织中的表达情况。结果激光共聚焦显微镜下观察,在HepG2细胞的胞膜、胞质及胞核中均可见明亮的绿色荧光,表明HBV preS1蛋白弥散分布于肝细胞的胞膜、胞质及胞核中;在胞膜上可见明亮的红色荧光,而胞质中的红色荧光较弱,表明ASGPR主要表达于肝细胞膜上,在肝细胞质中呈弱表达。提示在HepG2细胞的细胞膜上HBV preS1蛋白与ASGPR具有共定位。在乙肝肝硬化患者的肝组织中,HBV preS1蛋白在肝细胞的胞膜、胞质及胞核中均有阳性表达;ASGPR不仅在肝细胞膜上有阳性表达,在胞质中的表达率也较高。此外,ASGPR在肝组织中的分布形式与HBV preS1蛋白无明显差异(P〉0.05),并且二者的表达水平呈显著正相关(Pearson相关系数0.776,P〈0.000 1)。结论 ASGPR除介导HBV入侵外,可能还参与了病毒的分泌及胞内运输等其他生命活动。
Objective To verify the intracellular localization and tissue distribution of HBV preS1 protein and ASGPR so as to provide the basis for further study on their biological functions.Methods Intracellular localization of HBV preS1 protein and ASGPR was analyzed by confocal microscopy,respectively.Tissue specificity of HBV preS1 protein and ASGPR was analyzed by immunohistochemistry,respectively.Results Microscopy revealed that HBV preS1 protein was located on the membranes and in the cytoplasms and nuclei,while ASGPR was found on the cell membranes.We observed by double-channel confocal microscopy that HBV preS1 protein and ASGPR were co-localized on membranes of HepG2 cells;the localization pattern of ASGPR was similar to that of HBV preS1 protein in the liver tissues of HBsAg-positive hepatic cirrhosis patients(P〉0.05).In addition,the expressions of ASGPR and HBV preS1 protein were correlated significantly with each other(Pearson corrlection0.776,P〈0.0001).Conclusion Besides mediating HBV entry,ASGPR may also play an important role in other life activities such as viral secretion and intracellular transport.