Diagnosis and discrimination of autoimmune Graves' disease and Hashimoto's disease using thyroid-stimulating hormone receptor-containing recombinant proteoliposomes
Diagnosis and discrimination of autoimmune Graves' disease and Hashimoto's disease using thyroid-stimulating hormone receptor-containing recombinant proteoliposomes
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DOI：
10.1016/j.jbiosc.2009.06.006
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发表时间：
2009-12-01
影响因子：
2.8
通讯作者：
Yoshimura, Tetsuro
中科院分区：
文献类型：
--
作者：
Fukushima, Hidetaka;Matsuo, Hideaki;Yoshimura, Tetsuro
格雷夫斯病（GD）是一种甲状腺自身免疫性疾病，由抗促甲状腺激素受体（TSHR）的自身抗体引起。目前，TSHR自身抗体的诊断测试基于间接竞争性结合测定，该测定测量TSHR自身抗体抑制促甲状腺激素（TSH）与TSHR结合的能力。在这里，我们开发了一种针对 GD 自身抗体的特异性和直接诊断方法，该方法将固定化的含有 TSHR 的重组蛋白脂质体纳入酶联免疫吸附测定 (ELISA) 中。为了减少自身抗体与重组蛋白脂质体的非特异性结合，我们研究了聚乙二醇 (PEG)-脂质对市售抗 TSHR 抗体 (aTSHRAb) 结合的影响。与不含PEG-脂质的脂质体相比，将PEG-脂质掺入脂质体减少了非特异性结合，并且添加封闭试剂进一步减少了非特异性反应性。 aTSHRAb 对 PEG 修饰的 TSHR 重组蛋白脂质体表现出比不含 TSHR 的 PEG 修饰的脂质体（裸脂质体）更高的反应性。重要的是，与甲状腺功能亢进相关的 GD 患者的血清自身抗体表现出与 TSHR 重组蛋白脂质体的显着特异性结合。与甲状腺功能减退相关的桥本氏病 (HD) 患者的血清自身抗体也与脂蛋白体 TSHR 发生特异性反应。这些结果表明固定化 TSHR 重组蛋白脂质体可以作为 GD 和 HD 的直接诊断测试。此外，鉴于目前尚无可用于检测 HD 自身抗体的竞争试验，TSHR 重组蛋白脂质体 ELISA 与间接竞争 TSHR 结合试验的结合可能是区分 GD 和 HD 的有效方法。 (C) 2009 年，日本生物技术协会。版权所有。
Graves' disease (GD) is an autoimmune disease of the thyroid gland caused by autoantibodies against thyroid-stimulating hormone receptor (TSHR). Currently, the diagnostic test for TSHR autoantibodies is based on an indirect competitive binding assay that measures the ability of TSHR autoantibodies to inhibit the binding of thyroid-stimulating hormone (TSH) to TSHR. Here, we have developed a specific and direct diagnostic method for autoantibodies in GD that incorporates immobilized TSHR-containing recombinant proteoliposomes into an enzyme-linked immunosorbent assay (ELISA). To reduce non-specific binding of autoantibodies to recombinant proteoliposomes, we investigated the effect of polyethylene glycol (PEG)-lipid on the binding of commercially available anti-TSHR antibodies (aTSHRAb). The incorporation of PEG-lipids into liposomes decreased non-specific binding, as compared to liposomes that did not contain PEG-lipids, and the addition of blocking reagents further decreased non-specific reactivity. aTSHRAb exhibited higher reactivity towards PEG-modified TSHR recombinant proteoliposomes than PEG-modified liposomes without TSHR (bare liposomes). Importantly, serum autoantibodies from patients with GD, which is associated with hyperthyroidism, exhibited remarkably specific binding to TSHR recombinant proteoliposomes. Serum autoantibodies from patients with Hashimoto's disease (HD), which is associated with hypothyroidism, also reacted specifically with proteoliposomal TSHR. These results suggest that immobilized TSHR recombinant proteoliposomes can serve as a direct diagnostic test for GD and HD. Furthermore, given that there is no competition test currently available for detecting autoantibodies in HD, the combination of TSHR recombinant proteoliposome ELISA and indirect competitive TSHR binding assay might be an effective way to discriminate between GD and HD. (C) 2009, The Society for Biotechnology, Japan. All rights reserved.