Costagliola S, Many M C, Stalmans-Falys M, Tonacchera M, Vassart G, Ludgate M
Institut de Recherches Interdisciplinaires en Biologie Humaine et Nucleaire, Universite Libre de Bruxelles, Belgium.
Endocrinology. 1994 Nov;135(5):2150-9. doi: 10.1210/endo.135.5.7956939.
In a preliminary study, we observed the production of TSH binding-inhibiting (TBII) and thyroid-blocking (TBAb) antibodies accompanied by lymphocytic infiltration of the thyroid in a pool of male BALB/c mice immunized with the extracellular domain (ECD) of the human TSH receptor (TSHR) expressed as a maltose-binding protein (MBP) fusion in bacteria. In the present study we evaluated the humoral response to the same antigenic preparation in a new series of individual male and female BALB/c mice immunized ip on day 0 with 100 micrograms MBP-ECD and days 25, 39, and 53 with 50 micrograms MBP-ECD in an adjuvant composed of aluminum oxide, magnesium hydroxide, and Bordetella pertussis vaccine. Mice immunized with MBP served as control. Individual sera and immunoglobulins were tested for TBII, TBAb, and thyroid-stimulating antibodies (TSAb) on days 0, 32, 46, and 60, and total circulating T4 levels were measured by RIA. Animals were killed on day 120, their thyroids were examined histologically, the infiltrates were characterized using monoclonal antibodies specific for T-cells (total, activated, helper, and suppressor), B-cells, and macrophages. Sera and immunoglobulins G of the MBP-treated control group were all negative for TSAb, TBAb, and TBII activity. The receptor-immunized mice, despite having high titers of antibodies to the immunogen in an enzyme-linked immunosorbent assay, displayed a heterogeneous response in terms of biological activity, with 3 of 7 female and 4 of 8 male mice having TBAb/TBII activities that persisted and whose activity increased throughout the experiment. No significant TSAb antibody activity was observed. Total T4 levels were also heterogeneous even before immunization, but 9 of 15 MBP-ECD-treated mice had levels below the normal range after immunization, and 7 of these also had TBII/TBAb activities. At the end of the experiment, only 4 of the MBP-ECD-treated female mice survived, but all of them had a severe lymphocytic infiltration of their thyroid, composed mostly of activated T-cells, although B-cells and macrophages were also present. A similar infiltrate was seen in 4 of 8 male MBP-ECD-treated mice. No infiltrate was observed in male or female MBP-treated mice. The model described demonstrates the feasibility of using the TSHR as an immunogen to overcome tolerance and mimics some characteristics of human autoimmune disease of the thyroid.
在一项初步研究中，我们观察到，在用在细菌中表达为麦芽糖结合蛋白（MBP）融合体的人促甲状腺激素受体（TSHR）的细胞外结构域（ECD）免疫的一群雄性BALB/c小鼠中，产生了促甲状腺激素结合抑制（TBII）和甲状腺阻断（TBAb）抗体，同时伴有甲状腺的淋巴细胞浸润。在本研究中，我们评估了新的一组雄性和雌性BALB/c小鼠对相同抗原制剂的体液反应，这些小鼠于第0天腹腔注射100微克MBP-ECD，并于第25、39和53天腹腔注射50微克MBP-ECD，佐剂由氧化铝、氢氧化镁和百日咳博德特氏菌疫苗组成。用MBP免疫的小鼠作为对照。在第0、32、46和60天检测个体血清和免疫球蛋白的TBII、TBAb和促甲状腺激素刺激抗体（TSAb），并通过放射免疫分析法测量总循环T4水平。在第120天处死动物，对其甲状腺进行组织学检查，使用针对T细胞（总T细胞、活化T细胞、辅助性T细胞和抑制性T细胞）、B细胞和巨噬细胞的单克隆抗体对浸润细胞进行表征。MBP处理的对照组的血清和免疫球蛋白G的TSAb、TBAb和TBII活性均为阴性。尽管在酶联免疫吸附测定中受体免疫的小鼠对免疫原具有高滴度抗体，但在生物学活性方面表现出异质性反应，7只雌性小鼠中有3只和8只雄性小鼠中有4只具有持续存在且在整个实验过程中活性增加的TBAb/TBII活性。未观察到明显的TSAb抗体活性。即使在免疫前，总T4水平也存在异质性，但15只接受MBP-ECD处理的小鼠中有9只在免疫后水平低于正常范围，其中7只也具有TBII/TBAb活性。在实验结束时，接受MBP-ECD处理的雌性小鼠中只有4只存活，但它们的甲状腺均有严重的淋巴细胞浸润，主要由活化的T细胞组成，尽管也存在B细胞和巨噬细胞。在8只接受MBP-ECD处理的雄性小鼠中有4只出现了类似的浸润。在接受MBP处理的雄性或雌性小鼠中未观察到浸润。所描述的模型证明了使用TSHR作为免疫原克服耐受性并模拟人类甲状腺自身免疫疾病某些特征的可行性。
