Development and Characterization of Desmoglein-3 Specific T cells from Japanese Patients with Pemphigus Vulgaris

本研究的目的是调查日本患者对T淋巴细胞的反应，对白内障（PV）的反应，对Desmoglein 3、PV的自动抗原，并对Dsg 3-特定T细胞的特性进行表征。我们在14个日本PV患者中使用Amagai博士使用PCR-RFLP （聚合酶链反应-限制片段长度多态性）方法进行了高分辨率HLA II型。与健康个人相比，DRB 1*14 （14/14：100%）、DRB 1 *0503 （12/14：85%）或DRB 1 *0301（9/12：64%）都有重要的关联。2. 11种可重组Dsg 3融合蛋白（Dsg 3）1-11）与麦芽糖结合蛋白（MBP）准备好每个40个氨基酸被15个驻地标记，从Dsg 33的EC 1开始。对14名PV患者和20个健康控制的PBMC与11个重组Dsg 3融合蛋白和[YYYD 13 YD 1H]-胸腺嘧啶的克隆的自主T细胞反应。体外T细胞的初级反应（SI=1.9-2.7）对Dsg 3.9 （201-204）或Dsg 3.10 （226-265）有观察到的11/14 PV患者和7/14健康个人对与PV相关的HLA-DRB 1 *14和DQB 1 *05的观察。在相反的情况下，来自6种正常控制的PBMC携带HLA II类其他的DRB 1 *14和DQB 1 *05没有被重组Dsg 3融合蛋白刺激。这些观测表明，可以在PV患者中检测到Dsg 3的T细胞反应，并在健康的捐赠者携带主要的历史兼容性复合体类II警报中确定或类似于那些在PV中具有高度优先地位的人。并建议Dsg 3分子EC2-3区域可能包含由Dsg 3特定T细胞识别的表位。本研究将允许我们进一步发展和表征日本PV患者的Dsg 3特定T细胞克隆体。

The purpose of this study is to investigate the response of T lymphocytes from Japanese patients with pemphigus vulgaris (PV) to Desmoglein 3, the autoantigen of PV, and to characterize the properties of Dsg 3-specific T cells.1. We performed high resolution HLA class II typing in 14 Japanese PV patients presented from Dr Amagai using the PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) method. There was a significant association of either DRB 1*14 (14/14:100%), DQB1*0503 (12/14:85%), or DQB1*0301(9/12:64%), when compared with healthy individuals. 2. The 11 recombinant Dsg 3 fusion proteins (Dsg 3. 1-11) with Maltose Binding Protein (MBP) were prepared to encompass each 40 amino acids staggered by 15 residues, starting from the EC 1 of Dsg 33. Autoreactive T cell responses to Dsg 3 were investigated in 14 PV patients and 20 healthy controls by coculture of PBMC with the 11 recombinant Dsg 3 fusion proteins and the incorporation of [ィイD13ィエD1H]-thymidine. Primary in vitro T cell responses (SI=1.9-2.7) to Dsg 3.9 (residues 201-204) or Dsg 3.10 (residues 226-265) were observed in 11/14 PV patients and 7/14 healthy individuals expressing the PV-associated HLA-DRB1*14 and DQB1*05. In contrast, PBMC from 6 normal controls carrying HLA class II alleles other than DRB1*14 and DQB1*05 were not stimulated by recombinant Dsg 3 fusion proteins. These observations demonstrate that T cell response to Dsg 3 can be detected in PV patients and in healthy donors carrying major histocompatibility complex class II alleles identical or similar to those highly prevalent in PV. And that suggests EC2-3 region of the Dsg 3 molecule may contain epitopes that are recognized by Dsg 3-specific T cell. This study will allow us to further develop and characterize the Dsg 3-specific T cell clone in Japanese PV patients.