Single-cell RNA sequencing on human bone marrow-derived mesenchymal stem cells

To investigate the transcriptional diversity of BM-MSCs, we performed single-cell RNA sequencing (scRNA-seq) on freshly isolated CD271+ BM-derived mononuclear cells (BM-MNCs) from two human subjects. We successfully identified LEPRhiCD45low BM-MSCs in the CD271+ BM-MNC population, and further classified the BM-MSCs into distinct subpopulations corresponding to the osteogenic, chondrogenic, and adipogenic differentiation trajectories, as well as terminal-stage quiescent cells. This study provides a systematic dissection of human BM-MSCs at the single-cell resolution, offering novel insights into the extent of their cellular heterogeneity and importance for bone homeostasis. Overall design: We conducted scRNA-seq on freshly isolated CD271+ BM-MNCs from the femur head of two human subjects (one with osteoporosis and one with osteoarthritis). Cells were isolated by positive selection of CD271 using magnetic microbeads, and mRNA libraries were prepared and sequenced using the 10x Genomics Chromium system. To investigate the cellular heterogeneity within BM-MSCs, we extracted LEPR+CD45- cells (clusters C and D) from the original dataset for further analysis.