Study on B cell subsets producing ongan-specific autoantibodies

B细胞亚群中存在CD5^+B细胞和CD5^-B细胞。我们先前发现CD5~+B细胞在自身免疫性甲状腺疾病和其他自身免疫性疾病中增加。Graves病患者甲状腺功能亢进状态下CD5~+B细胞数量明显高于正常状态，且与血清甲状腺激素或甲状腺自身抗体水平相关。因此，我们检测了哪些B细胞亚群产生甲状腺自身抗体，以及甲状腺激素是否会增加CD5^+B细胞。我们使用细胞分选仪(FACStar)从自身免疫性甲状腺疾病患者的外周血单个核细胞中分离CD5^+B细胞和CD5^-B细胞。每个B细胞亚群在B细胞和T细胞有丝分裂原的刺激下，与照射后的自体淋巴细胞饲养细胞培养7天。然后，用ELISPOT方法明确了CD5-B细胞产生Ig G类的甲状腺过氧化物酶抗体和甲状腺球蛋白抗体。然而，我们没有发现产生TSH受体抗体的B细胞亚群。有趣的是，CD5^+B和CD5^-B细胞都产生了IgM类甲状腺自身抗体。另一方面，我们给C57BL/6J小鼠注射甲状腺素或甲状旁腺素1-6个月，以制造甲状腺功能亢进或甲状腺功能低下的小鼠。用双色流式细胞仪检测脾淋巴细胞亚群。甲亢小鼠NK细胞和T细胞增加，CD5β-B细胞增加。CD5+B细胞不随血清甲状腺激素水平的变化而变化。此外，随着桥本病的加重，破坏性甲状腺功能亢进症患者血清甲状腺激素水平升高，但CD5+B细胞减少，提示CD5+B细胞可能影响CD5+-B细胞产生甲状腺抗体，CD5+B细胞的增加可能是Graves病发病的基础。

CD5^+B cells and CD5^-B cells are present in a B cell subpopulation. We previously found that CD5^+B cells are increased in autoimmune thyroid disease as well as in other autoimmune diseases. The number of CD5^+B cells is increased more in the hyperthyroid state than in the euthyroid state in Graves' disease, and is correlated with the serum levels of thyroid hormones or thyroid autoantibodies. Therefore, we examined which subset of B cells produces thyroid autoantibodies, and whether thyroid hormones may increase CD5^+B cells.We separated CD5^+B cells and CD5^-B cells from peripheral blood mononuclear cells from patients with autoimmune thyroid disease by using a cell sorter (FACStar). Each subset of B cells was cultured for 7 days with feeder cells of irradiated autologous lymphocytes in stimulation with B and T cell mitogens. Then, we clarified that CD5-B cells produce IgG-class of thyroid-peroxidase antibodies and of thyroglobulin antibodies by using ELISPOT assay. However, we did not identify a B cell subset producing TSH-receptor antibodies. Interestingly, IgM-class of thyroid autoantibodies was produced by both CD5^+B and CD5^-B cells.On the other hand, we administrated thyroxine or PTU to C57BL/6J mice for 1-6 months to make hyper- or hypothyroid mice. Splenic lymphocyte subsets were examined by two-clour flow cytomotry. NK cells and T cells were increased in hyperthyroid mice, and CD5^-B cells were increased in hypothyroid mice. CD5^+B cells were did not change with the serum levels of thyroid hormone. Furthermore, CD5^+B cells decreased in patients with destructive thyrotoxicosis due to the aggravation of Hashimoto's disease, even though the serum levels of thyroid hormones increased.These data suggest that CD5^+B cells may affect the production of thyroid-antibodies by CD5^-B cells, and that the increase of CD5^+B cells might be fundamental in the pathogenosis of Graves' disease.