Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
J Invest Dermatol. 2010 May;130(5):1391-9. doi: 10.1038/jid.2010.16. Epub 2010 Feb 11.
Scurfy mice have a deletion in the Foxp3 gene, resulting in a failure to generate Foxp3(+) regulatory T cells, and they subsequently develop severe CD4(+) T-cell-mediated autoimmune inflammation. Multiple organs are involved, but the skin is one of the main organs affected. During the course of disease, Scurfy mice develop autoantibodies; however, the targeted antigens are unknown. In this study, we show that Scurfy mice develop autoantibodies directed against skin antigens. Using western blot analysis, we found that Scurfy serum reacted with proteins in total skin lysate, as well as in a keratinocyte lysate. Most of the Scurfy sera tested identified a major band at 50 kDa. Transfer of Scurfy CD4(+) T cells into nu/nu mice yielded autoantibodies with similar reactivity. Further analysis using 2D western blots, followed by peptide mass fingerprinting, identified several keratins as targets. To confirm this observation, we chose one of the identified targets, keratin 14, and prepared recombinant proteins encompassing the N-terminal, middle, and C-terminal portions of the keratin 14 protein. Scurfy serum predominantly recognized the C-terminal fragment. Sera from patients with immunodysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome, the human disease resulting from FOXP3 mutations, also recognized skin antigens, including keratin 14. Thus, the results of our study indicate that autoantibodies in Scurfy mice and patients with IPEX target keratins.
患皮疹的小鼠 Foxp3 基因缺失，导致无法生成 Foxp3(+)调节性 T 细胞，随后它们会发展出严重的 CD4(+)T 细胞介导的自身免疫炎症。多个器官都会受到影响，但皮肤是主要的受影响器官之一。在疾病过程中，皮疹小鼠会产生自身抗体；然而，针对的抗原尚不清楚。在这项研究中，我们表明皮疹小鼠会产生针对皮肤抗原的自身抗体。通过 Western blot 分析，我们发现皮疹血清与总皮肤裂解物以及角质形成细胞裂解物中的蛋白发生反应。大多数测试的皮疹血清都在 50 kDa 处识别出一个主要条带。将皮疹 CD4(+)T 细胞转移到 nu/nu 小鼠中会产生具有相似反应性的自身抗体。使用 2D Western blot 进一步分析，然后进行肽质量指纹图谱分析，鉴定出几种角蛋白作为靶标。为了证实这一观察结果，我们选择了鉴定出的靶标之一，角蛋白 14，并制备了包含角蛋白 14 蛋白 N 端、中间和 C 端部分的重组蛋白。皮疹血清主要识别 C 端片段。免疫失调、多内分泌腺病、肠病、X 连锁（IPEX）综合征患者的血清，即源自 FOXP3 突变的人类疾病患者的血清，也识别皮肤抗原，包括角蛋白 14。因此，我们的研究结果表明，皮疹小鼠和 IPEX 患者的自身抗体针对角蛋白。
