Abstract Body (Do not enter title and authors here): Background:
Fragility of the ascending aortic wall has been observed in patients with aortic regurgitation (AR), possibly due to turbulent retrograde flow. However, the underlying cellular and structural mechanisms remain unclear. To investigate the effects of AR-induced retrograde flow on endothelial cells (ECs) and smooth muscle cells (SMCs) in the ascending aorta, we developed a catheter-based AR model in rats and performed histological and single-cell analyses.
Methods:
AR was induced in 10-week-old Sprague–Dawley rats by echocardiography-guided perforation of the aortic valve via the right common carotid artery. Sham-operated controls underwent the same procedure without valve injury. Ascending aortic tissues were collected at 1, 2, and 4 weeks for histological analysis. EC polarity was assessed by whole-mount immunohistochemistry at one week. Single-cell RNA sequencing (scRNA-seq) was performed on aortic tissues, with differential gene expression and pathway enrichment analyzed using Seurat and clusterProfiler.
Results:
Confocal imaging revealed that ECs in AR rats showed disrupted polarity and a more rounded morphology compared to controls. Quantitative analysis demonstrated significantly reduced aspect ratio and cell orientation angle in the AR group. Histological analysis showed progressive medial degeneration by 4 weeks, including elastic fiber fragmentation, mucopolysaccharide accumulation, and fibrosis. Picro-Sirius Red staining demonstrated a significant increase in fibrotic area at 4 weeks. Furthermore, serial echocardiography revealed time-dependent dilation of the ascending aorta in AR rats.
ScRNA-seq revealed that ECs downregulated shear stress–responsive and anti-inflammatory genes. SMCs showed decreased expression of contractile markers and extracellular matrix (ECM) remodeling genes. Gene Ontology analysis revealed upregulation of inflammatory signaling and suppression of shear stress–responsive pathways in ECs. ECs also exhibited increased Wnt and TGF-β signaling—pathways known to drive SMC phenotypic switching—which was accompanied by phenotypic modulation and impaired ECM remodeling in SMCs.
Conclusion:
AR-induced retrograde flow leads to early EC polarity disruption and progressive medial degeneration in the ascending aorta. These structural changes are accompanied by shear stress–dependent transcriptional suppression in ECs and phenotypic modulation in SMCs, potentially contributing to aortic wall fragility.