Single-nucleus RNA-sequencing of venous endothelial cells from patients with OSA revealed 204 differentially expressed genes and reduced cell junction assembly compared to healthy controls.
Observational (n=27)
Single-nucleus RNA sequencing of venous endothelial cells reveals that obstructive sleep apnea is associated with downregulation of pathways critical for endothelial function and vascular health, providing mechanistic insights into OSA-related cardiovascular risk.
Abstract Rationale Obstructive sleep apnea (OSA) triples cardiovascular risk; however, the underlying mechanisms remain unclear. With the recent discovery of the crucial role of venous ECs in human atherosclerotic plaque progression, we hypothesized that unbiased profiling of venous ECs from patients with OSA and OSA-free controls will provide novel insights into mechanisms of increased cardiovascular risk in OSA. Methods The ECs were harvested from human brachial vein and processed for single-nucleus RNA-sequencing (snRNA-seq) following the protocol from the manufacturer (CG000124). Harvested ECs were incubated with nuclei isolation buffer for 45 seconds on ice after RBC removal. Nuclei were washed and filtered through a 40 μm cell strainer (H13680-0040) before processing using the 10X Genomics 3’v4 reagent kits. Libraries were sequenced using NovaSeq6000 and the FASTQ files were processed using CellRanger-9.0.1 and aligned to GRCh38-2024-A transcriptome with Ensembl110 annotations. Individual samples were QC’ed before integrating with Harmony. Empty droplets and debris were removed with DIEM, multiplets were removed with DoubletFinder, and ambient RNAs were removed with SoupX. Log2fold change ≥0.58 and Bonferroni corrected P 0.05 were used to define differentially expressed genes. Gene set enrichment analysis was conducted using clusterProfiler. Results The EC snRNA-seq analysis showed 204 differentially expressed genes in otherwise healthy patients with OSA (n = 14, 6F; mean±SD age 44±9 y, BMI 35±7 kg/m2, AHI 33 ±18/h) vs. healthy controls (n = 13, 6F; mean±SD age 41±16 y, BMI 28±4 kg/m2, AHI 1.8 ±1.3/h). Gene set enrichment analysis showed that ECs from patients with OSA have reduced cell junction assembly (NES=-1.6, Padj=3.5E-5) and organization (NES=-1.44, Padj=6.9E-5), negative regulation of cell population proliferation (NES=-1.43, Padj=0.0001), blood vessel morphogenesis (NES=-1.40, Padj=0.001) and development (NES=-1.38, Padj=0.001), vasculogenesis (NES=-1.75, Padj=0.002), EC differentiation (NES=-1.58, Padj=0.013) and nitric oxide metabolic process (NES=-1.68, Padj=0.013) pathways. To provide insights into the role of venous ECs in OSA-associated cardiovascular disease risk, we analyzed publicly available single-cell RNA sequencing datasets from human atherosclerotic plaques. Venous-specific genes (FLRT2, ARHGAP6) that were significantly downregulated in patients with OSA compared with controls were present in datasets from human atherosclerotic plaques. Conclusions Unbiased profiling of accessible venous ECs by snRNA-seq analysis from patients with OSA and OSA-free controls may provide novel insights into pathways activated by IH in inaccessible venous ECs located within atherosclerotic plaque that play role in plaque progression and increased cardiovascular risk in OSA. This abstract is funded by: NIH
Hsu et al. (Fri,) conducted a observational in Obstructive sleep apnea (n=27). Obstructive sleep apnea vs. Healthy controls was evaluated on Differentially expressed genes in venous endothelial cells. Single-nucleus RNA-sequencing of venous endothelial cells from patients with OSA revealed 204 differentially expressed genes and reduced cell junction assembly compared to healthy controls.