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Molecular biological significance of asialoglycoprotein receptor in autoimmune hepatitis type I

自身免疫性I型肝炎脱唾液酸糖蛋白受体的分子生物学意义

基本信息

批准号：
06454261
负责人：
KIYOSAWA Kendo
金额：
$ 4.61万
依托单位：
Shinshu University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1996
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06454261/
关键词：
autoimmune hepatitis asialoglycoprotein receptor autoantibody molecular biology

项目摘要

Reverse transcription and polymerase chain reaction was applied to isolate cDNAs encoding asialoglycoprotein receptor (ASGPR) H1 and L-H2 from human liver mRNAs. Using these cDNAs, recombinant ASGPR protein was expressed in E.coli. Rabbit antisera specific to human ASGPR H1 or L-H2 was obtained by immunizing these recombinant proteins. Enzyme-linked immunosorbent assay (ELISA) using bacterial recombinant antigen was developed to measure serum antibodies to ASGPR.Sera from autoimmune hepatitis, primary biliary cirrhosis, alcoholic liver disease, chronic hepatitis C,chronic hepatitis B,systemic lupus erythematosus, and healthy subjects were tested. High OD value was obtained almost exclusively in autoimmune hepatitis and chronic hepatitis C.The OD value of anti-ASGPR antibodies detected by ELISA using recombinant antigen was correlated with anti-ASGPR antibody titers that were determined by ELISA using human liver antigen. By using extracellular domain of ASGPR protein in ELISA,similar sensitivity to pick-up anti-ASGPR antibodies was demonstrated in autoimmune hepatitis and less reactivity was detected in chronic hepatitis C.It was suggested that main epitope which was reactive with antibodies in autoimmune hepatitis could be located on the extracellular domain of the ASGPR protein. Lymphocytes from peripheral blood mononuclear cells in a patient with autoimmune hepatitis showed a stimulation index as 4 in lymphocyte stimulation test with recombinant ASGPR antigen. There could exist a certain class of lymphocytes reactive with human ASGPR in autoimmune hepatitis patients.

应用逆转录-聚合酶链反应技术从人肝组织中分离编码去唾液酸糖蛋白受体（ASGPR）H1和L-H2的cDNA。利用这些cDNA，重组ASGPR蛋白在大肠杆菌中表达。用重组蛋白免疫兔获得抗人ASGPR H1和L-H2的兔抗血清。应用细菌重组抗原建立酶联免疫吸附试验（ELISA），检测自身免疫性肝炎、原发性胆汁性肝硬化、酒精性肝病、慢性丙型肝炎、慢性B型肝炎、系统性红斑狼疮和健康人血清中抗ASGPR抗体。高OD值几乎完全在自身免疫性肝炎和慢性丙型肝炎中获得。使用重组抗原通过ELISA检测的抗ASGPR抗体的OD值与使用人肝抗原通过ELISA测定的抗ASGPR抗体滴度相关。用ASGPR蛋白胞外区进行ELISA检测，发现自身免疫性肝炎对抗ASGPR抗体的敏感性相似，而慢性丙型肝炎对抗ASGPR抗体的敏感性较低，提示自身免疫性肝炎与抗体反应的主要抗原表位可能位于ASGPR蛋白胞外区。1例自身免疫性肝炎患者外周血单个核细胞经重组ASGPR抗原刺激试验，刺激指数为4。自身免疫性肝炎患者体内可能存在一定种类的ASGPR反应性淋巴细胞。