[Study on protective effects and mechanisms of total glucosides of Paeony on Graves disease in BALB/c mice]

Abstract

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, 3^rd, and 6^th 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 1^st week after the 2^nd 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 4^th week. At the 10^th 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 4^th 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 10^th 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 10^th week (4 weeks after three recombinant adenovirus immunization) (P<0.05). Compared with the model group at the 10^th 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.