The intricate cellular ecosystem of human peripheral veins as revealed by single-cell transcriptomic analysis

Abstract

The venous system has been historically understudied despite its critical roles in blood distribution, heart function, and systemic immunity. This study dissects the microanatomy of upper arm veins at the single cell level, and how it relates to wall structure, remodeling processes, and inflammatory responses to injury. We applied single-cell RNA sequencing to 4 non-diseased human veins (3 basilic, 1 cephalic) obtained from organ donors, followed by bioinformatic and histological analyses. Unsupervised clustering of 20,006 cells revealed a complex ecosystem of endothelial cell (EC) types, smooth muscle cell (SMCs) and pericytes, various types of fibroblasts, and immune cell populations. The venous endothelium showed significant upregulation of cell adhesion genes, with arteriovenous zonation EC phenotypes highlighting the heterogeneity of vasa vasorum (VV) microvessels. Venous SMCs had atypical contractile phenotypes and showed widespread localization in the intima and media. MYH11+DESlo SMCs were transcriptionally associated with negative regulation of contraction and pro-inflammatory gene expression. MYH11+DEShi SMCs showed significant upregulation of extracellular matrix genes and pro-migratory mediators. Venous fibroblasts ranging from secretory to myofibroblastic phenotypes were 4X more abundant than SMCs and widely distributed throughout the wall. Fibroblast-derived angiopoietin-like factors were identified as versatile signaling hubs to regulate angiogenesis and SMC proliferation. An abundant monocyte/macrophage population was detected and confirmed by histology, including pro-inflammatory and homeostatic phenotypes, with cell counts positively correlated with age. Ligand-receptor interactome networks identified the venous endothelium in the main lumen and the VV as a niche for monocyte recruitment and infiltration. This study underscores the transcriptional uniqueness of venous cells and their relevance for vascular inflammation and remodeling processes. Findings from this study may be relevant for molecular investigations of upper arm veins used for vascular access creation, where single-cell analyses of cell composition and phenotypes are currently lacking.

Single-Cell Analyses Offer Insights into the Different Remodeling Programs of Arteries and Veins

“…In addition to the above differences in EC abundance between arteries and veins, the identification of EC phenotypes by subclustering analysis revealed a striking contrast in vessel-specific subpopulations (Figure 2A). Six main EC phenotypes were uncovered by this analysis: (1) ACKR1+, previously identified in the main lumen and venules of veins [29]; (2) SEMA3G+, of arteriolar or capillary characteristics; (3) ITLN1+, here identified as the predominant phenotype in brachial arteries; (4) EFEMP1+, valvularlike ECs; (5) PROX1+, lymphatic ECs; and (6) AIF1+, an inflammatory subtype also detected in this study. ITLN1+ cells represent more than half of arterial ECs and are barely detected in veins (Figure 2A, Table 2).…”

“…Vascular ECs comprise a wide diversity of transcriptional phenotypes depending on their physiological states and macro-or microvascular locations [29]. Combined with hemodynamic conditions, this phenotypic variability influences vascular function and remodeling processes.…”

“…The increased diversity of EC subtypes in veins and the overall higher proportion of ECs compared with arteries can be explained by the higher counts of vasa vasorum in the venous wall (Figure 2E,F). This may also be related to the higher proportion of pericytes in veins interacting with microvessels and elevated counts of infiltrated monocyte/macrophages [29] (Table 1). Table 3 presents a summary of markers for the main EC subtypes in arteries and veins.…”