Haizao-Gancao herb pair ameliorates propylthiouracil-induced goiter by regulating the Beclin1-mediated autophagy

Haizao, Sargassum, is widely used to treat goiter. Gancao, Glycyrrhizae radix et rhizome, is renowned for reducing toxicity or increasing effects in traditional Chinese medicine. As a classic herb pair, Haizao-Gancao (HG) is a commonly used and effective combined therapy for goiter. The underlying biological mechanisms of HG on goiter is still unclear.

To explore the effect of HG on goiter, employing molecular docking combined with experimental validation to elucidate the molecular mechanism.

The rat goiter model was created by gavageing propylthiouracil (PTU) intragastrically for a duration of 14 days. The rats had been separated into six groups: control, model, euthyrox, HG high-dose (HG-H), medium-dose (HG-M) and low-dose (HG-L) group. Based on general observations (such as the rats' living status, body weight, and rectal temperature), the relative weight of the thyroid, thyroid function, hematoxylin-eosin (HE) staining, and transmission electron microscopy (TEM) to view the pathological variations of the thyroid glands, the effect of HG was evaluated. Discovered the chemical composition of HG by UPLC-MS/MS and the possible targets were predicted adopting several databases. Next, we explored their pharmacological mechanisms using molecular docking and validated key targets using western blotting (WB) and co-immunoprecipitation (Co-IP).

HG significantly increased the levels of triiodothyronine(T3), free triiodothyronine (FT3), free thyroxine (FT4), gained body weight and reduced tumescent thyroid glands in PTU-induced rats. The model group pathological changes such as uneven size, irregular shape and disordered arrangement of follicular epithelial cells occurred. However, HG groups thyroid follicles and epithelial cells appeared apparently normal. A variety of characteristic changes of autophagic vesicles appeared in the HG groups as opposed to the model group. In conclusion, the HG-L showed the best therapeutic effect. By UPLC-MS/MS, the major chemical components of HG were identified. The result revealed that HG contained flavonoids, alkaloids, organic acids, phenolic acidsand and terpenoids, etc. The molecular docking results of formononetin and naringenin and Beclin1 protein showed a high interaction of -5.38 kcal/mol and -5.25 kcal/mol. This implies that formononetin and naringenin may have a therapeutic effect in goiter rats by controlling the Beclin1-mediated autophagy. Western Blot (WB) and co-immunoprecipitation (Co-IP) results showed that HG can disrupt Beclin1/class III phosphatidylinositol 3-kinase(PI3KC3) binding and promote Beclin1/B-cell leukemia/lymphoma-2(Bcl-2) complex formation. Taken together, results demonstrate that autophagy inhibition via reducing Beclin1/PI3KC3 formation and increasing Beclin1/Bcl-2 binding activity.

HG ameliorates propylthiouracil-induced goiter by regulating the Beclin1-mediated autophagy, thus promoting the autophagy of thyroid cells.