Supplementary Material for: Roles of Thyroid Hormones, Mast Cells, and Inflammatory Mediators in the Initiation and Progression of Autoimmune Thyroid Diseases

<b><i>Background:</i></b> Interrelation between thyrocytes and immunocytes has been established. However, the roles of mast cells and thyroid hormones in the triggering mechanism of thyroid autoimmune processes have been insufficiently investigated. This study evaluated the role of thyroid hormones and mast cells in the mechanisms of losing tolerance to thyroid autoantigens. <b><i>Materials and Methods:</i></b> Two groups of patients were studied: patients with Graves’ disease and patients with nodular euthyroid goiter. Wistar rats with induced exogenous hypothyroidism, thyrotoxicosis, and thyrotoxicosis in parallel with administration of interleukin-2 were used. The levels of hormones, autoantibodies, and cytokines in the serum and thyroid tissue were analyzed through the enzyme-linked immunosorbent assay. Morphological verification was performed through the immune-histochemical method with antibodies against tryptase and CD86. Transmission electron microscopy and laser confocal microscopy were used. <b><i>Results:</i></b> In both Graves’ disease and induced thyrotoxicosis, we detected a significant increase in the levels of interferon-γ, active interfollicular infiltration and degranulation of mast cells, and the intrathyroid overexpression of CD86. Complex analysis of the rat’s thyroid morphofunctional state and systemic and local levels of cytokines in induced thyrotoxicosis and hypothyroidism demonstrated an increase of intrathyroid degranulation of mast cells and a drastic disruption of IFNγ/IL10 balance. <b><i>Conclusions:</i></b> When exposed to excessive amounts of thyroid hormones, an inflammatory response is triggered in the thyroid gland, and mast cells overexpress costimulating CD86 in the thyroid. This finding confirms their possible involvement in auto­antigen presentation. Significant increase in the levels of interferon-γ shows a determining influence of cytokine on the progression of the pathological process.

**背景：** 甲状腺细胞（thyrocyte）与免疫细胞（immunocyte）之间的相互作用已得到证实。然而，肥大细胞（mast cell）与甲状腺激素（thyroid hormones）在甲状腺自身免疫过程（thyroid autoimmune processes）触发机制中的作用，目前仍未得到充分研究。本研究旨在探讨甲状腺激素与肥大细胞在甲状腺自身抗原耐受丧失（losing tolerance to thyroid autoantigens）机制中的作用。
**材料与方法：** 本研究纳入两组受试者：格雷夫斯病（Graves’ disease）患者与结节性正常甲状腺肿（nodular euthyroid goiter）患者。实验采用经造模的Wistar大鼠（Wistar rats），模型分别为外源性甲状腺功能减退症（exogenous hypothyroidism）、甲状腺毒症（thyrotoxicosis），以及联合给予白细胞介素-2（interleukin-2）的甲状腺毒症模型。采用酶联免疫吸附试验（enzyme-linked immunosorbent assay）检测血清与甲状腺组织中激素、自身抗体及细胞因子水平；采用抗类胰蛋白酶（tryptase）与抗CD86抗体的免疫组织化学法（immune-histochemical method）进行形态学验证。此外，辅以透射电子显微镜（transmission electron microscopy）与激光共聚焦显微镜（laser confocal microscopy）开展相关观测。
**结果：** 在格雷夫斯病患者与造模甲状腺毒症大鼠中，均检测到干扰素-γ（interferon-γ）水平显著升高、活跃的滤泡间浸润（interfollicular infiltration）与肥大细胞脱颗粒（degranulation of mast cells）现象，以及甲状腺内CD86过表达。对造模甲状腺毒症与甲状腺功能减退症大鼠的甲状腺形态功能状态，以及全身与局部细胞因子水平进行综合分析后发现，大鼠甲状腺内肥大细胞脱颗粒程度升高，且IFNγ/IL10平衡出现显著紊乱。
**结论：** 当机体暴露于过量甲状腺激素时，甲状腺内会触发炎症反应，且肥大细胞在甲状腺内过表达共刺激分子CD86。该结果证实肥大细胞可能参与自身抗原呈递（autoantigen presentation）过程。干扰素-γ水平的显著升高，提示该细胞因子对病理进程的发展具有决定性影响。