Abstract：
Background: Graves\' disease (GD), one of the most common forms of autoimmune thyroid disorders, is characterized by hyperthyroidism caused by antibodies (Abs) against the extracellular A-subunit of the thyrotropin receptor (TSHR). Various approaches have been used to create mouse models of GD, including transfected fibroblasts and immunization with plasmids or adenoviruses expressing human TSHR A-subunit (hTSHR A-subunit). These models, however, require repeated immunization and produce inconsistent results. In this study, we established a novel Cre-loxP system-based mouse model that is able to generate the hTSHR A-subunit, mimicking human GD, and characterized the histological changes in Graves\' orbitopathy (GO) progression after a single injection. Materials and Methods: A Cre-loxP system-based mouse model was constructed by inserting the CAG-loxP-STOP-loxP-hTSHR A-subunit cassette into the Rosa26 locus of the mouse genome. Conditional expression of the hTSHR A-subunit was successfully achieved by intramuscular injection of the transactivator of transcription-Cre recombinase (GD mice). Blood tests for anti-TSHR Abs and the total thyroxine (T4) level were performed. Magnetic resonance imaging (MRI) was used to monitor morphological changes in the eyes. A histological examination of the thyroid gland and retrobulbar tissues was performed to observe pathological changes. Results: Twenty-four (8 control and 16 GD) mice were investigated. All GD mice exhibited higher levels of TSHR Abs compared with the control group. Moreover, more than 80% of the mouse models showed elevated T4 levels accompanied by thyroid goiter. MRI analysis revealed an increased volume of retrobulbar tissue, while immunohistochemical staining of orbital tissues exhibited macrophage infiltration and muscle fibrosis in the GD mice, contrasting with the control group. Conclusions: Our novel mouse model for GD, which showed the histological features of GO, was successfully established using the Cre-loxP system. This animal model offers improved insights and contributes to advancing methodological developments for GD and GO.
摘要：
背景：格雷夫斯病（GD），一种最常见的自身免疫性甲状腺疾病,其特征在于由针对促甲状腺激素受体(TSHR)的细胞外A亚基的抗体(Ab)引起的甲状腺功能亢进。各种方法已被用于创建GD的小鼠模型，包括转染的成纤维细胞和用表达人TSHRA亚基(hTSHRA亚基)的质粒或腺病毒免疫。这些模型，然而,需要重复的免疫接种和产生不一致的结果。这里,我们建立了一种新的基于Cre-loxP系统的小鼠模型，该模型能够产生hTSHRA亚基，模仿人类GD，并表征了单次注射后GO进展的组织学变化。
方法：通过将CAG-loxP-STOP-loxP-hTSHRA亚基盒插入小鼠基因组的Rosa26基因座，构建了基于Cre-loxP系统的小鼠模型。通过肌内注射转录反式激活因子（TAT）-Cre重组酶（GD小鼠），成功实现了hTSHRA亚基的条件表达。进行了抗TSHR抗体和总T4水平的血液测试。磁共振成像用于监测眼睛的形态变化。对甲状腺和球后组织进行组织学检查以观察病理变化。
结果：二十四（8对照，研究了16只GD)小鼠。与对照组相比,所有GD小鼠表现出更高水平的TSHR抗体。此外，超过80％的小鼠模型显示T4水平升高，伴有甲状腺甲状腺肿。MRI分析显示球后组织体积增加,而眼眶组织的免疫组织化学染色在GD小鼠中表现出巨噬细胞浸润和肌肉纤维化，与对照组相比。
结论：使用Cre-loxP系统成功建立了显示GO组织学特征的新型GD小鼠模型。这种动物模型提供了改进的见解，并有助于推进GD和GO的方法学发展。
方法：通过将CAG-loxP-STOP-loxP-hTSHRA亚基盒插入小鼠基因组的Rosa26基因座，构建了基于Cre-loxP系统的小鼠模型。通过肌内注射转录反式激活因子（TAT）-Cre重组酶（GD小鼠），成功实现了hTSHRA亚基的条件表达。进行了抗TSHR抗体和总T4水平的血液测试。磁共振成像用于监测眼睛的形态变化。对甲状腺和球后组织进行组织学检查以观察病理变化。
结果：二十四（8对照，研究了16只GD)小鼠。与对照组相比,所有GD小鼠表现出更高水平的TSHR抗体。此外，超过80％的小鼠模型显示T4水平升高，伴有甲状腺甲状腺肿。MRI分析显示球后组织体积增加,而眼眶组织的免疫组织化学染色在GD小鼠中表现出巨噬细胞浸润和肌肉纤维化，与对照组相比。
结论：使用Cre-loxP系统成功建立了显示GO组织学特征的新型GD小鼠模型。这种动物模型提供了改进的见解，并有助于推进GD和GO的方法学发展。
