Abstract
Identification and isolation of hematopoietic stem cells (HSCs) in mice is most commonly based on the expression of surface molecules Kit and Sca-1 and the absence of markers of mature lineages. However, Sca-1 is absent or weakly expressed in hematopoietic progenitors in many strains, including nonobese diabetic (NOD), BALB/c, C3H, and CBA mice. In addition, both Kit and Sca-1 levels are modulated following bone marrow injury. In these cases, other markers and dye exclusion methods have been employed to identify HSCs, yet there is no antibody-based stain that enables identification of HSCs and early progenitors when Kit and Sca-1 are inadequate. CD201 is a marker that is highly restricted to HSCs and progenitors, and CD27 is expressed at moderate-to-high levels on HSCs. We show here that combining CD201 and CD27 enables highly efficient isolation of long-term HSCs in NOD mice as well as in other strains, including SJL, FVB, AKR, BALB/c, C3H, and CBA. We also find that HSCs appear to maintain expression of CD201 and CD27 after hematopoietic injury when Kit expression is downregulated. These results suggest a widely applicable yet simple alternative for HSC isolation in settings where Kit and Sca-1 expression are insufficient.
Copyright © 2015 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Antigens, Ly / genetics
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Antigens, Ly / physiology
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Autoimmunity
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Blood Cells / chemistry*
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Blood Cells / cytology
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Bone Marrow / radiation effects
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Cell Lineage
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Cell Separation / methods*
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Diabetes Mellitus, Type 1 / blood
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Diabetes Mellitus, Type 1 / genetics
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Diabetes Mellitus, Type 1 / pathology
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Endothelial Protein C Receptor
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Flow Cytometry / methods*
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Gene Expression
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Hematopoietic Stem Cells / chemistry*
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Hematopoietic Stem Cells / cytology
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Membrane Proteins / deficiency
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Membrane Proteins / genetics
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Membrane Proteins / physiology
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Mice
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Mice, Inbred NOD
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Mice, Inbred Strains / blood*
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Mice, Inbred Strains / genetics
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Mice, Transgenic
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Proto-Oncogene Proteins c-kit / deficiency
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Proto-Oncogene Proteins c-kit / genetics
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Proto-Oncogene Proteins c-kit / physiology
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Radiation Chimera
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Radiation Injuries, Experimental / blood
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Radiation Injuries, Experimental / pathology
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Receptors, Cell Surface / biosynthesis
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Receptors, Cell Surface / blood*
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Receptors, Cell Surface / genetics
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Tumor Necrosis Factor Receptor Superfamily, Member 7 / biosynthesis
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Tumor Necrosis Factor Receptor Superfamily, Member 7 / blood*
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Tumor Necrosis Factor Receptor Superfamily, Member 7 / genetics
Substances
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Antigens, Ly
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Endothelial Protein C Receptor
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Ly6a protein, mouse
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Membrane Proteins
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Procr protein, mouse
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Receptors, Cell Surface
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Tumor Necrosis Factor Receptor Superfamily, Member 7
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Proto-Oncogene Proteins c-kit