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Updated: Jul 26, 2026
Isolation of Valvular Endothelial Cells
Published on: December 29, 2010
Side- and Disease-Dependent Changes in Human Aortic Valve Cell Population and Transcriptomic Heterogeneity Determined
Nicolas Villa-Roel1, Christian Park1, Aitor Andueza1
1Wallace H. Coulter Department of Biomedical Engineering, Emory University and Georgia Institute of Technology, 1760 Haygood Drive, Health Sciences Research Bldg E170, Atlanta, GA 30322, USA.
Insights
Calcific aortic valve disease (CAVD) affects the fibrosa side more due to distinct cell differences. Targeting these unique valvular endothelial cells (VECs) in the fibrosa may offer new therapeutic strategies for CAVD.
Area of Science:
- Cardiovascular Biology
- Molecular Medicine
- Genomics
Background:
- Calcific aortic valve disease (CAVD) is common in the elderly, lacking effective drug therapies.
- CAVD preferentially affects the fibrosa side of aortic valves, with unknown reasons.
- Hypothesis: Side-dependent transcriptomic and cellular differences predispose the fibrosa to CAVD.
Purpose of the Study:
- Investigate side-dependent transcriptomic and cellular phenotypes in human aortic valves.
- Identify mechanisms underlying CAVD development in the fibrosa layer.
- Discover potential therapeutic targets for CAVD.
Main Methods:
- Single-cell RNA sequencing of endothelial-enriched samples from fibrosa and ventricularis sides.
- Analysis of human aortic valve leaflets from five donors across disease spectrum.
- Utilized a novel method for side-specific sample collection.
Main Results:
- Identified 27 cell clusters, including valvular endothelial cells (VECs), valvular interstitial cells (VICs), and immune cells.
- Discovered side-dependent VEC subtypes with distinct gene expression.
- Observed increased inflammatory VICs, macrophages, and T-cells with CAVD progression.
- Found upregulation of AP-1 transcription factors and EGR1 in fibrosa of diseased leaflets.
- Identified CAVD-associated VEC clusters linked to inflammation, EMT, apoptosis, and fibrosis.
Conclusions:
- Valvular endothelial cells (VECs) exhibit significant heterogeneity dependent on tissue side and CAVD status.
- Unique VEC clusters and pathways in the fibrosa of diseased valves suggest novel pathogenic mechanisms.
- Differentially regulated genes and pathways in fibrosa VECs represent potential therapeutic targets for CAVD.
Background:
Calcific aortic valve disease (CAVD) is a highly prevalent disease, especially in the elderly population, but there are no effective drug therapies other than aortic valve repair or replacement. CAVD develops preferentially on the fibrosa side, while the ventricularis side remains relatively spared through unknown mechanisms. We hypothesized that the fibrosa is prone to the disease due to side-dependent differences in transcriptomic patterns and cell phenotypes.
Methods:
To test this hypothesis, we performed single-cell RNA sequencing using a new method to collect endothelial-enriched samples independently from the fibrosa and ventricularis sides of freshly obtained human aortic valve leaflets from five donors, ranging from non-diseased to fibrocalcific stages.
Results:
From the 82,356 aortic valve cells analyzed, we found 27 cell clusters, including seven valvular endothelial cell (VEC), nine valvular interstitial cell (VIC), and seven immune, three transitional, and one stromal cell population. We identified several side-dependent VEC subtypes with unique gene expression patterns. Homeostatic VIC clusters were abundant in non-diseased tissues, while VICs enriched with fibrocalcific genes and pathways were more prevalent in diseased leaflets. Furthermore, homeostatic macrophage (MΦ) clusters decreased while inflammatory MΦ and T-cell clusters increased with disease progression. A foamy MΦ cluster was increased in the fibrosa of mildly diseased tissues. Some side-dependent VEC clusters represented non-diseased, protective phenotypes, while others were CAVD-associated and were characterized by genes enriched in pathways of inflammation, endothelial-mesenchymal transition, apoptosis, proliferation, and fibrosis. Interestingly, we found several activator protein-1 (AP-1)-related transcription factors (FOSB, FOS, JUN, JUNB) and EGR1 to be upregulated in the fibrosa and diseased aortic valve leaflets.
Conclusions:
Our results showed that VECs are highly heterogeneous in a side- and CAVD-dependent manner. Unique VEC clusters and their differentially regulated genes and pathways found in the fibrosa of diseased tissues may represent novel pathogenic mechanisms and potential therapeutic targets.
