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Quantitative imaging of haematopoietic stem and progenitor cell localization and hypoxic statusin the bone marrow microenvironment
Csar Nombela-Arrieta1,5, Gregory Pivarnik1, Beatrice Winkel1, Kimberly J. Canty1, Brendan Harley2,
John E. Mahoney3, Shin-Young Park1, Jiayun Lu1, Alexei Protopopov3,4 and Leslie E. Silberstein1,5
The existence of a haematopoietic stem cell niche as a spatially conned regulatory entity relies on the notion that haematopoietic stem and progenitor cells (HSPCs) are strategically positioned in unique bone marrow microenvironments with dened anatomical and functional features. Here, we employ a powerful imaging cytometry platform to perform a comprehensive quantitative analysis of HSPC distribution in bone marrow cavities of femoral bones. We nd that HSPCs preferentially localize in endosteal zones, where most closely interact with sinusoidal and non-sinusoidal bone marrow microvessels, which form a distinctive circulatory system. In situ tissue analysis reveals that HSPCs exhibit a hypoxic prole, dened by strong retention of pimonidazole and expression of HIF-1 , regardless of localization throughout the bone marrow, adjacency to vascular structures or cell-cycle status. These studies argue that the characteristic hypoxic state of HSPCs is not solely the result of a minimally oxygenated niche but may be partially regulated by cell-specic mechanisms.
The bone marrow cavities of long bones are the principal sites of postnatal haematopoiesis, which is sustained by a rare population of HSPCs (ref. 1). As for other well-defined adult stem cell types, HSPCs have been proposed to reside in defined anatomical locations, where they receive and integrate regulatory cues from neighbouring cells, extracellular matrix components and/or soluble factors24. The precise definition of the physical localization and physiological features of HSPC niches has been greatly hampered by the technical difficulties associated with imaging long bones, the need for complex cell-surface
1Division of Transfusion Medicine, Department of Laboratory Medicine, Childrens Hospital Boston, 300 Longwood Avenue, Boston, Massachusetts 02115, USA.
2Department for Chemical and Biomolecular Engineering, University of Illinois, Urbana Champaign, Illinois 61801, USA. 3Division of Hematologic Malignancies, Dana-Farber Cancer Institute, Boston, Massachusetts 02115, USA. 4Present address: Institute for Applied Cancer Science, The University of Texas MD Anderson Cancer Center, Texas 77030, USA.
5Correspondence should be addressed to C.N-A. or L.E.S. (e-mail: mailto:[email protected]
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