Revealing the Mechanism of Buzhong Yiqi Tang in Ameliorating Autoimmune Thyroiditis via the Toll-like Receptor Pathway.

Objective: Autoimmune thyroiditis (AIT) is one of the most common autoimmune diseases and often causes hypothyroidism in patients. As a traditional formulation in my country, Buzhong Yiqi decoction (BZYQD) has significant effects in improving clinical symptoms of AIT and reducing autoantibody titers, but its specific mechanism of action needs to be further explored. The purpose of this study was to explore the effective targets and related mechanisms of Buzhong Yiqi decoction in AIT mice based on transcriptome sequencing technology.

Methods: Forty NOD.H-2h4 mice were selected and 0.05% NaI was drinking ad libitum for 8 weeks to establish AIT mice, and drug intervention was performed according to groups for 8 weeks. The groups were as follows: control group, Model Group, Buzhong Yiqi Decoction group (9.56 g·kg-1) and Positive control group (Se yeast tablets, 3.033×10-5 g·kg-1), of which Buzhong Yiqi Decoction was the clinical equivalent dose. Thyroid tissues of the Model Group, blank group and Buzhong Yiqi Decoction group were subjected to transcriptome sequencing to analyze the expression of differential genes, and enrichment analysis was carried out. Hematoxylin and eosin staining (HE staining) was used to detect the pathological changes in thyroid tissues, and enzymelinked immunosorbent assay (ELISA) was used to detect the content of serum thyroglobulin antibody (TGAb) to determine the intervention effect of Buzhong Yiqi Decoction; Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) was performed based on the transcriptome sequencing results to detect the expression of TLR8, JUN, TICAM2, TIRAP, and IL-1β mRNA in thyroid tissue.

Results: According to the transcriptome results, compared with the blank group, there were 327 significantly up-regulated genes and 440 significantly down-regulated genes in the Model Group; compared with the Model Group, there were 502 significantly up-regulated genes and 380 significantly down-regulated genes in the Buzhong Yiqi Decoction group, mainly enriched in immune inflammation and other related pathways including Toll-like receptors. Animal experiments showed that compared with the control group, the model group had obvious lymphocyte infiltration in thyroid tissue under light microscope, a significant increase in inflammatory cells, a significant increase in TGAb content in serum, and a significant increase in TLR8, JUN, TICAM2, TIRAP, IL-1β mRNA expression was observed (P<0.05 or P<0.01). Compared with the Model Group, Buzhong Yiqi Decoction could significantly improve the inflammatory damage of thyroid tissue in AIT mice, reduce the content of TGAb in serum, and down-regulate the expression of TLR8, JUN, TICAM2, TIRAP, IL-1β mRNA (P<0.05 or P<0.01).

Conclusion: Buzhong Yiqi Decoction can effectively improve the inflammatory damage of AIT, and inhibiting the abnormal activation of the Toll-like receptor pathway may be one of its intervention mechanisms.