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Preparation of a Single-Cell Suspension from Mouse Carotid Arteries for Single-Cell Sequencing

Carotid arteries are major blood vessels in the neck that supply blood and oxygen to the brain, but carotid stenosis occurs when carotid arteries are ...

Soochow University

Disturbed blood flow (d-flow) has been known to induce changes of the cells in the arterial wall, increasing the risk of atherosclerosis. However, the heterogeneity of the vascular cell populations under d-flow remains less understood. To generate d-flow in vivo, partial carotid artery ligation (PCL) was performed. Seven days after ligation, single-cell RNA sequencing of nine left carotid arteries (LCA) from the PCL group (10,262 cells) or control group (14,580 cells) was applied and a single-cell atlas of gene expression was constructed. The integrated analysis identified 15 distinct carotid cell clusters, including 10 d-flow-relevant subpopulations. Among endothelial cells, at least four subpopulations were identified, including Klk8 ECs, Lrp1 ECs, Dkk2 ECs, and Cd36 ECs. Analysis of GSVA and single-cell trajectories indicated that the previously undescribed Dkk2 ECs subpopulation was mechanosensitive and potentially transformed from Klk8 ECs under d-flow. D-flow-induced Spp1 VSMCs subpopulation that appeared to be endowed with osteoblast differentiation, suggesting a role in arterial stiffness. Among the infiltrating cell subpopulations, Trem2 Mφ, Birc5 Mφ, DCs, CD4 T cells, CXCR6 T cells, NK cells, and granulocytes were identified under d-flow. Of note, the novel Birc5 Mφ was identified as a potential contributor to the accumulation of macrophages in atherosclerosis. Finally, Dkk2 ECs, and Cd36 ECs were also found in the proatherosclerotic area of the aorta where the d-flow occurs. In conclusion, we presented a comprehensive single-cell atlas of all cells in the carotid artery under d-flow, identified previously unrecognized cell subpopulations and their gene expression signatures, and suggested their specialized functions.

Single-cell RNA-seq reveals cellular heterogeneity of mouse carotid artery under disturbed flow.

The differences in plaque histology between symptomatic and asymptomatic patients have been widely accepted. Whether there is a heterogeneity of cells between symptomatic and asymptomatic plaques remains largely unclear. To reveal the potential heterogeneity within different plaques, which may contribute to different stroke incidences, we obtained the scRNA-seq data from symptomatic and asymptomatic patients and identified eight cell types present in plaques. Further analysis of endothelial cells (ECs) revealed three distinct EC subpopulations appeared to be endowed with specific biological functions such as antigen processing and presentation, cell adhesion, and smooth muscle cell proliferation. Of note, the differentially expressed genes of the EC 2 subpopulation showed that the genes involved in cell adhesion were up-regulated in asymptomatic plaques compared to symptomatic plaques. Integrating the data of intraplaque haemorrhage and plaque stability, the 5th top-enriched biological process was cell adhesion in the stable or non-haemorrhaged plaques compared to unstable plaques or haemorrhaged plaques. Among these cell adhesion-related genes, the intersection gene AOC3 may play a vital role in plaque haemorrhage and plaque stability. Targeting cell adhesion and the specialized genes may provide potential new therapeutic directions to prevent asymptomatic patients from stroke.

Single-cell transcriptional profiling of human carotid plaques reveals a subpopulation of endothelial cells associated with stroke incidences.

Insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3) plays a key role in tumorigenesis and tumor progression. Lung cancer is the leading cause of cancer-related death in men and women all over the world. However, the relationship between IGF2BP3 and small-cell lung cancer (SCLC) has not been reported yet.

Yun Du

Cyrus Tang Hematology Center,
Cyrus Tang Medical Institute,
Cyrus Tang Hematology Center, Cyrus Tang Medical Institute

Soochow University

Preparation of a Single-Cell Suspension from Mouse Carotid Arteries for Single-Cell Sequencing

Yun Du

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Soochow University

Preparation of a Single-Cell Suspension from Mouse Carotid Arteries for Single-Cell Sequencing

Cyrus Tang Hematology Center,
Cyrus Tang Medical Institute,
Cyrus Tang Hematology Center, Cyrus Tang Medical Institute