Astragaloside IV Improves Pulmonary Vascular Endothelial Dysfunction in Pulmonary Arterial Hypertension by Inhibiting Ferroptosis through Modulation of the Calpain-1/TGF-β/TFRC Pathway

Pulmonary vascular endothelial dysfunction (PVED) is a significant contributor to pulmonary arterial hypertension (PAH). Research indicates that astragaloside IV (AS-IV) has a therapeutic effect on PAH, but the potential mechanism by which it improves the PVED in PAH remains unclear. This study primarily investigated the protective effect of AS-IV on PVED in PAH. Network pharmacological analysis revealed that AS-IV potentially exerts therapeutic effects on PVED in PAH patients mainly by affecting endothelial cell migration and ferroptosis-related biological processes. Elevated pulmonary arterial pressure, PVED, ferroptosis, and increased protein expression of calpain-1, TGF-β, TAZ, and TFRC have been found in hypoxia-induced PAH mice. However, AS-IV therapy reversed hypoxia-induced pulmonary arterial pressure elevation, endothelium-dependent diastolic impairment, elevated NO levels, iron accumulation, lipid peroxidation, and mitochondrial dysfunction. Moreover, AS-IV inhibited the calpain-1/TGF-β/TFRC signaling pathway. After calpain-1 gene knockout, Fer-1, MDL-28170, and SB-431542 induced effects similar to those of AS-IV. Furthermore, the overexpression of calpain-1 in HPAECs through viral transfection further decreased NO levels and aggravated ferroptosis induced by hypoxia. This also led to hypoxia-induced upregulation of the protein expression of calpain-1, TGF-β, TAZ, and TFRC. Moreover, the overexpression of calpain-1 reversed the ameliorative effect of AS-IV on hypoxia-induced dysfunction and ferroptosis in HPAECs. In conclusion, AS-IV can reduce ferroptosis-related iron accumulation, lipid peroxidation, and mitochondrial dysfunction by blocking the calpain-1/TGF-β/TFRC signaling pathway, ultimately improving PVED in hypoxia-induced PAH.