PTH‐induced EndMT via miR‐29a‐5p/GSAP/Notch1 pathway contributed to valvular calcification in rats with CKD

Background Endothelial‐to‐mesenchymal transition (EndMT) is a common pathophysiology in valvular calcification (VC) among non‐chronic kidney disease (CKD) patients. However, few studies were investigated in CKD‐induced VC. Parathyroid hormone (PTH) was considered to be an important component of EndMT in CKD‐induced cardiovascular diseases. Therefore, determining whether PTH could induce valvular EndMT and elucidating corresponding mechanism involved further study. Methods Performing a 5/6 nephrectomy with a high phosphorus diet was done to construct VC models in rats with CKD. miRNA sequencing was used to ascertain changes in microRNA in human umbilical vein endothelial cells (HUVECs) intervened by PTH. VC was observed by Von Kossa staining and scanning electron microscope. Results PTH induced valvular EndMT in VC. Global microRNA expression profiling of HUVECs was examined in PTH versus the control in vitro, in which miR‐29a‐5p was most notably decreased and was resumed by PTHrP(7‐34) (PTH‐receptor1 inhibitor). Overexpression of miR‐29a‐5p could inhibit PTH‐induced EndMT in vitro and valvular EndMT in vivo. The dual‐luciferase assay verified that γ‐secretase‐activating protein (GASP) served as the target of miR‐29a‐5p. miR‐29a‐5p‐mimics, si‐GSAP and DAPT (γ‐secretase inhibitor) inhibited PTH‐induced γ‐secretase activation, thus blocking Notch1 pathway activation to inhibit EndMT in vitro. Moreover, Notch1 pathway activation was observed in VC. Blocking Notch1 pathway activation via AAV‐miR‐29a and DAPT inhibited valvular EndMT. In addition, blocking Notch1 pathway activation was also shown to alleviate VC. Conclusion PTH activates valvular EndMT via miR‐29a‐5p/GSAP/Notch1 pathway, which can contribute to VC in CKD rats. A model depicting how miR‐29a‐5p regulating VECs EndMT. PTH could trigger a decrease in miR‐29a through PTHR1 of VECs, leading to an increase in GSAP at the protein level. GSAP further activated γ‐secretase to increase the level of NICD in the cytoplasm, which promoted the increase of HES1 and Snail expression in the nucleus to mediate the activation of Notch1 signal and endothelial‐to‐mesenchymal transition (EndMT) which cause VECs transferring to valvular interstitial cells (VICs).