Single-Cell Analysis of Human SVF Reveals Early Transcriptional and Communication Shifts During ASC Culture Expansion

Background

Adipose-derived stem cells (ASCs) are known in regenerative medicine due to their multi-potency, immunomodulatory function, and facile isolation from the stromal vascular fraction (SVF). However, the specific SVF cell types that contribute to therapeutic efficacy and the mechanisms by which either fresh SVF or ASCs promote repair remain poorly defined. Fresh SVF is a heterogeneous mixture of hematopoietic cells, endothelial cells, pericytes, fibroblasts, and stromal progenitors (1). Upon culture expansion, this mixture shifts toward a mesenchymal stromal cell–enriched population dominated by CD73 -+ -CD90 -+ -CD105 -+ - ASCs, accompanied by loss of immune and endothelial lineages (2). Although these phenotypic changes are well recognized, the transcriptional trajectories, lineage relationships, and cell–cell communication events underlying this transition have not been fully resolved at single-cell resolution. To address this gap, we mapped SVF composition through initial culture expansion, defined accompanying gene-expression profiles, and characterized ligand–receptor–mediated communication using a genomics-driven single-cell RNA analysis.

Methods

Stromal vascular fraction (SVF) was obtained from human adipose samples, and ASCs were isolated through standard adherence-based culture techniques. RNA was extracted with a TRIzol kit. Single-cell RNA sequencing libraries were generated, and transcriptomic profiles were analyzed using Loupe Browser and BBrowserX. Cell identities were assigned based on canonical lineage markers and integrated with established reference datasets to annotation of cell.