A Cellular Taxonomy of the Bone Marrow Stroma in Homeostasis and Leukemia
Authors: Konstantinos D Kokkaliaris
Monika S Kowalczyk
Ani Papazian
Francois Mercier
David T Scadden
Matan Hofree
Dongjun Lee
Marcin Tabaka
Karin Gustafsson
Dariusz Przybylski
Danielle Dionne
Youmna Kfoury
Nicolas Severe
Orit Rozenblatt-Rosen
Ayshwarya Subramanian
Aviv Regev
Eeshit Dhaval Vaishnav
Ninib Baryawno
Orr Ashenberg
Journal: Cell (2019)
Discipline: MOLECULAR BIOLOGY & GENETICS
Keywords: bone marrow niche; hematopoiesis; leukemia; single-cell RNA-sequencing; stem cell
Affiliations: Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA 02114, USA; Harvard Stem Cell Institute, Cambridge, MA 02138, USA; Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA; Childhood Cancer Research Unit, Dep. of Children's and Women's Health, Karolinska Institutet, 171 77 Stockholm, Sweden., Klarman Cell Observatory, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA., Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA 02114, USA; Harvard Stem Cell Institute, Cambridge, MA 02138, USA; Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA., Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA 02114, USA; Harvard Stem Cell Institute, Cambridge, MA 02138, USA; Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA; Department of Convergence Medical Science, Pusan National University School of Medicine, Yangsan 50612, Republic of Korea., Klarman Cell Observatory, Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA; Howard Hughes Medical Institute, Koch Institute of Integrative Cancer Research, Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA. Electronic address: ., Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA 02114, USA; Harvard Stem Cell Institute, Cambridge, MA 02138, USA; Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA. Electronic address: .

Abstract:  Stroma is a poorly defined non-parenchymal component of virtually every organ with key roles in organ development, homeostasis, and repair. Studies of the bone marrow stroma have defined individual populations in the stem cell niche regulating hematopoietic regeneration and capable of initiating leukemia. Here, we use single-cell RNA sequencing (scRNA-seq) to define a cellular taxonomy of the mouse bone marrow stroma and its perturbation by malignancy. We identified seventeen stromal subsets expressing distinct hematopoietic regulatory genes spanning new fibroblastic and osteoblastic subpopulations including distinct osteoblast differentiation trajectories. Emerging acute myeloid leukemia impaired mesenchymal osteogenic differentiation and reduced regulatory molecules necessary for normal hematopoiesis. These data suggest that tissue stroma responds to malignant cells by disadvantaging normal parenchymal cells. Our taxonomy of the stromal compartment provides a comprehensive bone marrow cell census and experimental support for cancer cell crosstalk with specific stromal elements to impair normal tissue function and thereby enable emergent cancer. 

Retracted: No

PubPeer comments:
 Regarding sorting strategy Fig S1A mentions viability marker 7AAD as a singlet gate. Was FSC-H vs FSC-W -A also performed? Also the sorting gate for CD71 cuts right through the population. What was the recovery of cells during the process? How did you control whether you re not biased in enriching or losing certain cells along the process? Also, no comment on how this is basically the end of Nestin as a surrogate stromal marker? Not only is it not expressed on LepR-MSCs but as shown with reporter mice already, there s substantial expression in endothelial cells. , 
 #1 Dear Lisianthius Thank you for your comments. Below are the responses to your comments. Regarding the gating strategy for 7-AAD cell viability, we did not perform FSC-H vs FSC-W -scatter. Since we also use Calcein AM as a positive selection for live cells we are certain that we capture mostly live cells. Any cells that we capture by 10X we filter out in silico. The CD71 was run with FMO to make sure we know where to draw the gates. Any selection strategy will introduce bias. Ours is the most common one in the field: (i) sorting cells that are negative for heme markers should include all stroma cell populations; (ii) tolerance of microfluidic streaming that has been shown to reduce adipocyte, megakaryocyte and neutrophil populations. Much of the published work has been based on a Nestin-GFP indicator mouse where there is well-documented discrepancy between GFP and endogenous Nestin expression. Interpreting the data needs to be done with that in mind. ,