Emodin-induced ERα degradation via SYVN1 alleviates vascular calcification by preventing HIF-1α deacetylation in chronic kidney disease

Background

Vascular calcification is a major cause of death in chronic kidney disease (CKD), with osteoblastic transdifferentiation of vascular smooth muscle cells (VSMCs) considered to be the crucial pathologic process. However, there remains a significant deficiency in effective prevention and treatment strategies.

Purpose

In an in vitro calcification screening model, we observed an inhibitory activity of Emodin on osteoblastic transdifferentiation of VSMCs. In the present study, we therefore aimed to evaluate its efficacy in CKD-induced medial vascular calcification in vitro and in vivo, and to further explore the underlying mechanisms.

Study design

A7R5 and MOVAS cells were treated with sodium phosphate to induce osteogenic transdifferentiation as in vitro models, while mice were fed with adenine and a high-phosphorus diet, and additionally received an intraperitoneal injection of 10,000 IU Vitamin D3 to induce chronic kidney disease (CKD) as an in vivo model. Vitamin K2 served as a positive control.

Methods

Alizarin Red S staining and VON-KOSSA staining were used to evaluate the effects of Emodin on osteogenic transdifferentiation. Western blotting, RT-qPCR and Von-Kossa staining were used to detect the effects of Emodin on aortic calcification in CKD mice in vivo. Chemical biology techniques including ITC, fluorescence titration, dual fluorescein reporter genes, CETSA, and DARTS, were used to detect the binding activity of Emodin to ERα and SYVN1. Immunoprecipitation, immunostaining, etc. were used to explore the mechanisms, and small molecule inhibitors and small RNA interference were used to verify the target of Emodin.

Results

Emodin could effectively inhibit the osteogenic transdifferentiation of A7R5 and MOVAS cells in vitro, and alleviate aortic calcification in CKD mice in vivo. Mechanism study revealed that Emodin could act as a molecular glue that binds directly to ERα and SYVN1 and enhances their interaction, thereby accelerating the ubiquitination degradation process of ERα. The decrease in ERα diminished the inhibition of ERα on the deacetylation of HIF-1α by SIRT6, thereby inhibiting VSMC osteogenic transdifferentiation and relieving the vascular cells calcification.

Conclusion

Our study demonstrates that ERα has a non-genomic effect to inhibit the deacetylation of HIF-1α by SIRT6, which can be abrogated by Emodin through SYVN1-mediated ERα degradation. These results provide evidences for Emodin to serve as a candidate drug for controlling clinical vascular calcification in CKD.