Objective: To investigate the protective effect and its immunoregulatory mechanism of Total Glucosides of Paeony (TGP) against Graves' Disease (GD) model on BALB/c mice. Methods: Fifty female (6 weeks old, weighing 16-18 g) BALB/c mice of specific pathogen free were divided into control group according to random number table method, model group, early low-dose TGP intervention group (250 mg·kg-1·d-1), early high-dose TGP intervention group (500 mg·kg-1·d-1), and late TGP intervention group, with 10 mice in each group. Except the control group, the other 4 groups were immunized 3 times (0, 3rd, and 6th week) with recombinant adenovirus expressing the thyroid stimulating hormone receptor (TSHR) A subunit to establish the GD model. The early low-dose and high-dose intervention group were given diets containing different doses of TGP throughout the whole process, and the late intervention group was given diets containing low doses of TGP from the 1st week after the 2nd immunization (week 4). The levels of thyrotropin receptor antibody (TRAb) and total thyroxine (TT4) were detected in the tail venous blood of mice at the 4th week. At the 10th week, the serum TRAb and TT4 levels and the ratio of regulatory T cells (Treg) in each group were detected, and the pathological changes of thyroid tissue were observed. Serum helper T cell 1(Th1) and Th2 cell-related factors interleukin-2 (IL-2), IL-4, IL-5, IL-10, IL-12p70, granulocyte-macrophage colony-stimulating factor (GM-CSF), interferon-γ (IFN-γ) and tumor necrosis factors-α (TNF-α) were detected to investigate the protective effect of TGP on GD model in BALB/c mice and its mechanism. Results: At the 4th week, The level of TT4 [(55.07±12.89) μg/L] in early high-dose intervention group was lower than that in model group [(74.33±8.63) μg/L] (all P<0.05). The level of TT4 in early low-dose intervention group and late intervention group and model group had no statistical significance (all P>0.05). TRAb level of mice between early low-dose, early high-dose, late intervention groups and model group was no significant difference (all P>0.05). At the 10th week, TRAb [(90.00±26.89) U/L] and TT4[(32.66±8.11) μg/L] levels in the early high-dose intervention group were lower than those in the model group [(396.97±95.35) U/L, (73.70±16.33) μg/L] (all P<0.05). The TRAb and TT4 levels in the early low-dose intervention group and late intervention group were not significantly different from those in the model group (all P>0.05). The thyroid tissue of hyperthyroidism mice in the early high dose intervention group showed focal hypertrophic changes, while the thyroid tissue of other hyperthyroidism mice showed diffuse hypertrophic changes. The CD4+CD25+/CD4+Treg ratio in early high-dose intervention group was higher than that in model group at the 10th week (4 weeks after three recombinant adenovirus immunization) (P<0.05). Compared with the model group at the 10th week, the levels of IL-2, IL-12p70 and IFN-γ in the early high-dose intervention group were all decreased (all P<0.05), and the levels of IL-10 were increased (P<0.05). Conclusion: Early high-dose (500 mg·kg-1·d-1) TGP intervention group displays a protective effect against GD mice, the mechanism of which may be related to regulatory T cell function changes and Th1/Th2 cytokine balance restoration.
目的: 探讨白芍总苷对BALB/c小鼠Graves病(GD)模型的保护作用及其机制。 方法: 将50只(6周龄,体重16~18 g)无特定病原体级雌性BALB/c小鼠按随机数字表法分为5组,分别设为对照组、模型组、早期低剂量白芍总苷干预组(250 mg·kg-1·d-1)、早期高剂量白芍总苷干预组(500 mg·kg-1·d-1)和晚期白芍总苷干预组,每组各10只。除对照组外,其余4组均注射表达促甲状腺激素受体(TSHR)A亚单位的重组腺病毒进行3次免疫(第0、3和6周),完成GD建模;早期低剂量和高剂量干预组全程予以含不同剂量白芍总苷的饲料,晚期干预组自第2次免疫后1周(第4周)起加用含低剂量白芍总苷的饲料。于第4周取小鼠尾静脉血检测促甲状腺素受体抗体(TRAb)、总甲状腺素(TT4)水平;第10周检测小鼠血清TRAb及TT4水平及各组调节性T细胞(Treg)比率,并观察甲状腺组织病理变化。检测各组血清辅助性T细胞1(Th1)及Th2细胞相关因子白细胞介素-2(IL-2)、IL-4、IL-5、IL-10、IL-12p70、粒细胞-巨噬细胞集落刺激因子(GM-CSF)、干扰素-γ(IFN-γ)和肿瘤坏死因子-α(TNF-α)等水平,探讨白芍总苷对BALB/c小鼠GD模型的保护作用及其机制。 结果: 第4周时,早期高剂量干预组TT4[(55.07±12.89)μg/L]水平低于模型组[(74.33±8.63)μg/L](P<0.05),早期低剂量干预组和晚期干预组TT4水平和模型组差异无统计学意义(均P>0.05);早期低剂量干预组、早期高剂量干预组和晚期干预组小鼠TRAb水平和模型组差异均无统计学意义(均P>0.05)。第10周时,早期高剂量干预组TRAb[(90.00±26.89)U/L]、TT4[(32.66±8.11)μg/L]水平均低于模型组[(396.97±95.35)U/L,(73.70±16.33)μg/L](均P<0.05),而早期低剂量干预组和晚期干预组小鼠TRAb、TT4水平与模型组差异均无统计学意义(均P>0.05)。早期高剂量干预组甲亢小鼠甲状腺组织呈局灶性增生性改变,而其他组甲亢小鼠甲状腺组织呈弥漫性增生性改变。第10周(3次重组腺病毒免疫后4周),早期高剂量干预组小鼠CD4+CD25+/CD4+Treg比值高于模型组(P<0.05)。第10周小鼠血清细胞因子和模型组相比,早期高剂量干预组IL-2、IL-12p70和IFN-γ水平均降低(均P<0.05),IL-10水平升高(P<0.05)。 结论: 早期高剂量(500 mg·kg-1·d-1)白芍总苷对小鼠GD模型具有保护性作用,其作用机制可能和调节性T细胞功能改变及Th1/Th2细胞因子平衡恢复有关。.