Identification and Mapping of Human Lymph Node Stromal Cell Subsets by Combining Single-Cell RNA Sequencing with Spatial Transcriptomics

Lymph node stromal cells (LNSCs) have a crucial immunomodulatory function, but their heterogeneity in humans is incompletely understood. Here, we report the single-cell RNA sequencing (scRNA-seq) of 9267 LNSCs isolated from a human lymph node (LN). This study comprehensively defines the gene signatures of 10 fibroblast subtypes: CCL21⁺ SC, CCL19⁺ SC, CD34⁺CXCL14⁺ SC, pericytes, DES⁺ SC, LAMP5⁺ SC, NR4A1⁺BCAM⁺ SC, HLA-DR⁺ SC, SEPT4⁺ SC and GLDN⁺ SC. The existence of these subtypes was validated across 13 LN donors using 2 publicly available datasets and our dataset. To explore the heterogeneous stromal compartment within the complex LN tissue architecture, we integrated the scRNA-seq profiles of the identified LNSC subsets with a publicly available human spatial transcriptomic LN dataset and predicted their location within the complex LN tissue architecture. Each LNSC subtype was spatially restricted to specific LN regions, indicating different LNSC–lymphocyte interactions, which were further investigated using NicheNet. The positioning of distinct LNSC subtypes within different LN regions sets the stage for future research on the relationship between LNSC-specific niches and immunomodulatory function during health and disease.