Roxadustat ameliorates vascular calcification in CKD rats by regulating HIF‐2α/HIF‐1α

Abstract Vascular calcification (VC) is a common complication of chronic kidney disease (CKD). VC is a gene‐regulated process similar to osteogenic differentiation. There are still no convincing schemes to prevent and reduce the development of VC. It has been reported that hypoxia‐inducing factor 1α (HIF‐1α) and endothelin‐1(ET‐1) are related to VC. In this study, we found that the expression of ET‐1 and HIF‐1α was enhanced after VC, the interaction between HIF‐1α and ET‐1 was confirmed by CO‐IP and luciferase experiments. We found that ET‐1 was an upregulated differential gene of calcified vascular smooth muscle cells (VSMCs) through gene sequencing. However, hypoxia‐inducing factor 2α (HIF‐2α) and HIF‐1α have antagonistic effects on each other. HIF‐1α is a pro‐inflammatory cytokine, and HIF‐2α can improve inflammation and fibrosis. Roxadustat, as a selective PHD3 inhibitor, preferentially activates HIF‐2α. It is still unclear whether roxadustat improves VC in CKD by regulating the expression of HIF‐2α/HIF‐1α. Alizarin red staining and western blot as well as immunohistochemical results showed that roxadustat could significantly reduce the degree of vascular and VSMCs calcification in CKD rats. Serum HIF‐1α and ET‐1 were significantly decreased after roxadustat treatment. In addition, western blot results showed that roxadustat could decrease the expression of HIF‐1α and ET‐1 in vascular tissues and calcified VSMC, but HIF‐2α expression significantly increased. Interestingly, our study confirmed that activation of HIF‐1α or inhibition of HIF‐2α reversed the ameliorating effect of roxadustat on VC, proving that the effect mediated by roxadustat is HIF‐2α/HIF‐1α dependent. We have demonstrated for the first time that roxadustat improves VC in CKD rats by regulating HIF‐2α/HIF‐1α, thus providing a new idea for the application of roxadustat in VC of CKD.