Astragaloside IV Binds with RhoA, Inhibits EndMT and Ameliorates Myocardial Fibrosis in Mice.

Astragaloside IV (ASIV), the main active component of the traditional Chinese medicine HuangQi, exhibits ameliorating effects on myocardial fibrosis through unclear mechanisms. To investigate the effects of ASIV on Endothelial-to-mesenchymal transition (EndMT) in myocardial fibrosis, 10 ng/mL TGF-β1 was used to induce EndMT in human umbilical vein endothelial cells (HUVECs) in vitro, and a 5 mg/kg/d subcutaneous injection of Isoproterenol (ISO) was used to induce myocardial fibrosis in mice in vivo. The drug affinity-responsive target stability (DARTS) was used to identify the target proteins of ASIV in endothelial cells. The results showed that ASIV could significantly inhibit the TGF-β1-induced EndMT, which includes changes in cytoskeletal structure, the expression of EndMT markers, cell migration potency, and cell glycolysis rate. ASIV significantly ameliorated ISO-induced myocardial fibrosis in mice and inhibited EndMT in heart tissues. The Ras homolog gene family member A (RhoA) protein was found to be a possible direct binding target of ASIV in endothelial cells. The binding affinity between ASIV and RhoA was confirmed by molecular docking and the cellular thermal shift assay (CETSA). ASIV inhibited the RhoA-related pathway in the heart tissues of myocardial fibrosis mice. In addition, siRNA knockdown of RhoA expression or treatment with RhoA agonists was found to significantly affect the inhibition of EndMT by ASIV. The results suggested that ASIV could significantly inhibit the EndMT by binding with RhoA, and that the inhibition of EndMT by ASIV contributed to its amelioratory effects on myocardial fibrosis. This discovery provided a theoretical basis for the application of ASIV and HuangQi in the treatment of myocardial fibrosis.