A Promising Mouse Model of Graves’ Orbitopathy Induced by Adenovirus Expressing Thyrotropin Receptor A Subunit.

Graves’ orbitopathy (GO) is the most common and serious manifestation of Graves’ disease (GD). It is characterized by orbital inflammation and tissue remodeling. Although several GO models have been reported, most lack a full assessment or mechanistic evaluation. Here, we established a promising mouse model mimicking many aspects of human GO with a frequency of 70%, and characterized the key role of T-cells in the progression of GO.
An adenovirus expressing the human thyroid-stimulating hormone (TSH) receptor A subunit (Ad-TSHRA) was injected in the muscles of female BALB/C mice 9 times to induce GO. At predetermined time points, histological examinations of retrobulbar tissues and thyroid glands were performed to dynamically monitor changes; serum autoantibodies (TRAbs) and total thyroxine (TT4) levels were examined to evaluate thyroid function. Flow cytometry of CD4+ T-cell subgroups and RNA sequencing (RNA-seq) of splenocytes were also performed to explore the underlying mechanism.
After 9 injections, seven of ten mice challenged with Ad-TSHRA developed the orbital changes associated with GO. Seven mice manifested retrobulbar fibrosis, and four showed adipogenesis. Exophthalmia, conjunctival redness and orbital lymphocyte infiltration were also observed in a subset of mice. The orbitopathy was first detected after 7 injections and followed the hyperplastic change observed in thyroids after 4 injections. Flow cytometry revealed increased proportions of Th1 cells, decreased proportions of Th2 cells and regulatory T (Treg) cells in the splenocytes of GO mice. This change in CD4+ T-cell subgroups was confirmed by orbital immunohistochemical staining. Genes involved in T-cell receptor signaling, proliferation, adhesion, inflammation and cytotoxicity were upregulated in GO mice according to the RNA-seq; a trend of upregulation of these GO-specific genes was observed in mice with hyperthyroidism without orbitopathy after 4 injections.
A GO mouse model was successfully established by administering 9 injections of Ad-TSHRA. The model was achieved with a frequency of 70% and revealed the importance of T-cell immunity. A potential time window from Graves’ hyperthyroidism to GO was presented for the first time. Therefore, this model could be used to study the pathogenesis and novel treatments for GO.