Identification of autoantigens in Vogt-Koyanagi-Harada disease using recombinant retrovirus encoding melanocyte-specitic antigen genes

Vogt-Koyanagi-Harada病（VKH）似乎是一种针对黑色素细胞的自身免疫性疾病。葡萄膜炎的炎症性质以及与HLA-DRB1*0405的显著关联表明，CD4+ T细胞对HLA-DRB1*0405呈递的黑色素细胞特异性蛋白的反应参与了VKH的发生。我们已经发现了许多黑色素瘤抗原被肿瘤浸润的T淋巴细胞识别，包括黑素体抗原（酪氨酸酶、TRP1、TRP2、gp100和MART1）。在黑色素瘤患者的免疫治疗中，有时会发生皮肤色素沉着，并与黑色素瘤消退相关，这表明这些抗原也参与了对黑色素细胞的自身免疫反应。为了鉴定VKH特异性自身抗原，我们分析了T细胞对VKH黑色素细胞的反应。脉络丛中受黑素细胞刺激的T细胞可能在脑脊液中富集。我们最初试图从VKH患者脑脊液中的淋巴细胞中产生一些T细胞系和克隆。用极限稀释法从3例VKH患者中分离出61个T细胞克隆。患者的24个克隆以HLA- DR限制性方式识别了培养黑色素瘤细胞系（Skmel23）的裂解物。观察它们对HLA-DRB1*0405和各种黑色素细胞特异性抗原cdna共转染的293IMDR1或293CIITA的反应。KU-MEL-1被1个T细胞系和1个克隆以HLA-DRB1*0405限制性方式识别。此外，26例VKH患者中有15例（58%）血清中检测到KU-MEL-1 IgG抗体。因此KU-MEL-1可能作为CD4+ T细胞识别的自身抗原之一参与了VKH的发生。鉴定T细胞识别的自身抗原可能有助于了解VKH的病理过程，并为VKH患者开发新的诊断和治疗方法。

Vogt-Koyanagi-Harada disease (VKH) appears to be an autoimmune disease against melanocytes. Inflammatory nature of the uveitis and a significant association with HLA-DRB1*0405 suggest that CD4+ T cells reactive to melanocyte-specific proteins presented by HLA-DRB1*0405 are involved in the development of VKH. We have been identified many melanoma antigens recognized by tumor infiltrating T lymphocytes, including melanosomal antigen (tyrosinase, TRP1, TRP2, gp100, and MART1). In the immunotherapy for patients with melanoma, skin depigmentation sometimes occurs and correlates with melanoma regression, suggesting that these antigens are also involved in autoimmune responses to melanocytes. To identify VKH-specific autoantigens, we have analyzed T cell responses against melanocytes in VKH. T cells stimulated with melanocytes in choroid plexus may be enriched in cerebrospinal fluids. We initially attempted to generate some of T cell lines and clones from lymphocytes in cerebrospinal fluids of VKH patients. Total of 61 T cell clones were isolated from 3 VKH patients by the limiting dilution method. 24 clones from a patient recognized lysate from cultured melanoma cell line (Skmel23) in an HLA- DR restricted manner. Their responses against 293IMDR1 or 293CIITA co-transfected with HLA-DRB1*0405 and various melanocyte-specific antigen cDNAs were evaluated. KU-MEL-1 was recognized by one T cell line and one clone in HLA-DRB1*0405 restricted manner. Furthermore, IgG antibodies against KU-MEL-1 were detected in the sera from 15 of 26 (58%) VKH patients. Therefore KU-MEL-1 may be involved in development of VKH as one of the auto antigens recognized by CD4+ T cells. Identification of auto antigens recognized by T cells may lead to understanding of the pathological process involved in VKH as well as to development of new diagnostic and therapeutic methods for VKH patients.