Mechanisms of Sini Powder intervention in Graves′ disease based on Th1/Th2/Th17 cells immune imbalance

To investigate the effects of Sini Powder on Graves′ disease (GD) model mice and to explore its mechanism in regulating Th1/Th2/Th17 cells immune imbalance at three levels: immune center (spleen), humoral circulation (serum), and target organ (thyroid).

Twenty-four female BALB/c mice were randomly divided into normal group (n = 6) and modeling group (n = 18)using a randomized block design. GD model was established through intramuscular injection of AdTSHR289 adenovirus. After successful modeling, GD mice were randomly assigned to the model, methimazole (4.55×10^-3 g/kg), and Sini Powder (5.46 g/kg) groups using random number table, with corresponding interventions administered daily for 6 weeks. HE staining was used to assess pathological changes in thyroid follicular epithelial cells; serum levels of thyrotropin receptor antibody (TRAb) and thyroxine (T₄) were measured using radioimmunoassay. The proportions of Th1, Th2, and Th17 cells in spleen tissues were analyzed by flow cytometry; mRNA and protein expressions of Th1-derived interferon-γ (IFN-γ), Th2-derived interleukin-4 (IL-4), and Th17-derived interleukin-17A (IL-17A) in spleen were detected by quantitative reverse transcription PCR and Western blotting. Serum levels of IFN-γ, IL-4, IL-10, IL-17A, as well as regulated on activation, normal T-cell expressed and secreted (RANTES), macrophage inflammatory protein-1β (MIP-1β), and monocyte chemoattractant protein-1 (MCP-1) were measured using Luminex protein biochip assay. Immunohistochemistry was performed to localize and assess the expression of IFN-γ, IL-4, and IL-17A in thyroid tissues.

Sini Powder reduced thyroid volume of model mice, as well as decreased serum TRAb and T₄ levels (P< 0.01), ameliorated pathological damage to thyroid follicular structures, and alleviated splenomegaly. Mechanistic studies revealed that compared with the normal group, the model group exhibited increased trends of the proportions of Th1 and Th17 cells, along with an elevated Th1/Th2 ratio, whereas Th2 cell proportion decreased. Expressions of IFN-γ and IL-17A mRNA, as well as IFN-γ protein expression in spleen increased(P<0.01), with an upward trend in IL-17A protein expression, while IL-4 mRNA expression was downregulated (P<0.01). The serum levels of IFN-γ, IL-10, and RANTES elevated (P<0.01), with upward trends in IL-17A, MIP-1β, and MCP-1, and a downward trend in IL-4 level. The expressions of IFN-γ, IL-4, and IL-17A in thyroid tissues increased. These findings suggest the presence of Th1/Th2/Th17 cells immune imbalance in GD model mice. The methimazole and Sini Powder modulated expressions of IFN-γ, IL-4, and IL-17A in spleen, thyroid, and serum, as well as regulated the expression of chemokines including RANTES, MIP-1β, and MCP-1 of the GD model mice.

Sini Powder may exert intervention effect on goiter induced by GD via regulating Th1/Th2/Th17 cells immune imbalance and modulating chemokines such as RANTES, MIP-1β, and MCP-1.