Single-cell analysis of cultured bone marrow stromal cells reveals high similarity to fibroblasts in situ.

• Cultured BM stromal cells are distinct from in vivo mesenchymal stromal cells. • Cultured BM stromal cells are transcriptionally similar to in vivo fibroblasts. • Cultured BM stromal cells retain plasticity ex vivo. • Cultured BM stromal cells have hematopoiesis-support capacity, increased through TGFb inhibition. Within the heterogenous pool of bone marrow stromal cells, mesenchymal stromal cells (MSCs) are of particular interest because of their hematopoiesis-supporting capacities, contribution to disease progression, therapy resistance, and leukemic initiation. Cultured bone marrow-derived stromal cells (cBMSCs) are used for in vitro modeling of hematopoiesis–stroma interactions, validation of disease mechanisms, and screening for therapeutic targets. Here, we place cBMSCs (mouse and human) in a bone marrow tissue context by systematically comparing the transcriptome of plastic-adherent cells on a single-cell level with in vivo counterparts. Cultured BMSCs encompass a rather homogenous cell population, independent of the isolation method used and, although still possessing hematopoiesis-supporting capacity, are distinct from freshly isolated MSCs and more akin to in vivo fibroblast populations. Informed by combined cell trajectories and pathway analyses, we illustrate that TGFb inhibition in vitro can preserve a more "MSC"-like phenotype [ABSTRACT FROM AUTHOR]