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Blood. 2011 Aug 18;118(7):1774-83. doi: 10.1182/blood-2010-11-320705. Epub 2011 Jun 9.

Analysis of glycoprotein E-selectin ligands on human and mouse marrow cells enriched for hematopoietic stem/progenitor cells.

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1
Department of Dermatology, Brigham and Women's Hospital, Boston, MA, USA.

Abstract

Although well recognized that expression of E-selectin on marrow microvessels mediates osteotropism of hematopoietic stem/progenitor cells (HSPCs), our knowledge regarding the cognate E-selectin ligand(s) on HSPCs is incomplete. Flow cytometry using E-selectin-Ig chimera (E-Ig) shows that human marrow cells enriched for HSPCs (CD34(+) cells) display greater E-selectin binding than those obtained from mouse (lin(-)/Sca-1(+)/c-kit(+) [LSK] cells). To define the relevant glycoprotein E-selectin ligands, lysates from human CD34(+) and KG1a cells and from mouse LSK cells were immunoprecipitated using E-Ig and resolved by Western blot using E-Ig. In both human and mouse cells, E-selectin ligand reactivity was observed at ~ 120- to 130-kDa region, which contained two E-selectin ligands, the P-selectin glycoprotein ligand-1 glycoform "CLA," and CD43. Human, but not mouse, cells displayed a prominent ~ 100-kDa band, exclusively comprising the CD44 glycoform "HCELL." E-Ig reactivity was most prominent on CLA in mouse cells and on HCELL in human cells. To further assess HCELL's contribution to E-selectin adherence, complementary studies were performed to silence (via CD44 siRNA) or enforce its expression (via exoglycosylation). Under physiologic shear conditions, CD44/HCELL-silenced human cells showed striking decreases (> 50%) in E-selectin binding. Conversely, enforced HCELL expression of LSK cells profoundly increased E-selectin adherence, yielding > 3-fold more marrow homing in vivo. These data define the key glycoprotein E-selectin ligands of human and mouse HSPCs, unveiling critical species-intrinsic differences in both the identity and activity of these structures.

PMID:
21659548
PMCID:
PMC3158712
DOI:
10.1182/blood-2010-11-320705
[Indexed for MEDLINE]
Free PMC Article
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