Piezo1 mediates oscillatory shear stress-induced endothelial-to-mesenchymal transition through the AP-1 and KLF2 regulatory pathways in mitral regurgitation

Oscillatory shear stress (OSS) has been implicated in various cardiovascular pathologies, particularly in endothelial-to-mesenchymal transition (EndoMT). In the mitral valve regurgitation, OSS predisposes the endothelium to dysfunction, inflammation, and pathological remodeling. Piezo1, a mechanosensitive channel, plays a critical role in sensing fluid shear stress, but its downstream transcriptional effectors and their roles in valvular pathophysiology are poorly defined. We employed a transverse aortic constriction (TAC) mouse model to simulate pathological shear stress and induce mitral valve regurgitation. Immunofluorescence staining, calcium ion imaging, and Western blotting were used to assess Piezo1 activity and KLF2 expression. To identify key downstream mediators, a transcription factor-based screening approach was implemented. The role of Piezo1 was further investigated through siRNA-mediated knockdown and pharmacological inhibition with GsMTx4. Human umbilical vein endothelial cells (HUVECs) and porcine heart valve endothelial cells (PPHVECs) were subjected to different shear stress models to investigate changes in gene expression and cellular behavior. OSS significantly upregulated Piezo1 expression and activity. Our screening and validation efforts identified the transcription factor AP-1 as a pivotal downstream effector of Piezo1. Piezo1 activation under OSS promoted calcium influx, leading to increased AP-1 expression and subsequent EndoMT progression. Conversely, Piezo1 inhibition attenuated calcium influx, suppressed AP-1, and reversed the EndoMT phenotype. Furthermore, we discovered that the AP-1/KLF2 axis constitutes a major regulatory pathway, with Piezo1/AP-1 activation repressing the atheroprotective factor KLF2. Additionally, OSS modulated the FOXP1-Notch1 signaling pathway in a Piezo1-dependent manner. Critically, the pharmacological inhibitor GsMTx4 effectively blunted the pro-EndoMT effects of OSS. Our findings identify AP-1 as a key transcription factor linking Piezo1 activation to EndoMT under OSS. Piezo1 drives endothelial remodeling via coordinated regulation of the AP-1/KLF2 axis. Pharmacological targeting of Piezo1 represents a promising strategy for treating shear stress-induced valvular disorders.