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Ex vivo glycan engineering of CD44 programs human multipotent mesenchymal stromal cell trafficking to bone

Nature Medicine volume 14, pages 181–187 (2008)Cite this article

Abstract

The capacity to direct migration ('homing') of blood-borne cells to a predetermined anatomic compartment is vital to stem cell–based tissue engineering and other adoptive cellular therapies. Although multipotent mesenchymal stromal cells (MSCs, also termed 'mesenchymal stem cells') hold the potential for curing generalized skeletal diseases, their clinical effectiveness is constrained by the poor osteotropism of infused MSCs (refs. 1–3). Cellular recruitment to bone occurs within specialized marrow vessels that constitutively express vascular E-selectin4,5, a lectin that recognizes sialofucosylated determinants on its various ligands. We show here that human MSCs do not express E-selectin ligands, but express a CD44 glycoform bearing α-2,3-sialyl modifications. Using an α-1,3-fucosyltransferase preparation and enzymatic conditions specifically designed for treating live cells, we converted the native CD44 glycoform on MSCs into hematopoietic cell E-selectin/L-selectin ligand (HCELL)6, which conferred potent E-selectin binding without effects on cell viability or multipotency. Real-time intravital microscopy in immunocompromised (NOD/SCID) mice showed that intravenously infused HCELL+ MSCs infiltrated marrow within hours of infusion, with ensuing rare foci of endosteally localized cells and human osteoid generation. These findings establish that the HCELL glycoform of CD44 confers tropism to bone and unveil a readily translatable roadmap for programming cellular trafficking by chemical engineering of glycans on a distinct membrane glycoprotein.

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Figure 1: MSCs lack E-selectin ligands but express CD44 reactive with SACK-1 mAb, which recognizes a sialic acid–dependent epitope displayed on a CD44-specific N-glycan substitution.
Figure 2: FTVI treatment of MSCs elaborates sialofucosylations on N-linked glycans of CD44, resulting in HCELL expression.
Figure 3: HCELL+ MSCs have markedly enhanced shear-resistant adhesive interactions with endothelial E-selectin under defined shear stress conditions.
Figure 4: Human MSC homing to mouse bone marrow.

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Acknowledgements

We thank C.A. Knoblauch, L. Liu and J.Y. Lee for assistance in manuscript preparation and for skilled technical support, as well as P.V. Hauschka for helpful discussion of the data. We are grateful to the staff of the Cell Processing Laboratory of the Bone Marrow Transplantation Unit at the Massachusetts General Hospital and the Cell Manipulation Core Facility of Dana Farber Cancer Center for their assistance in procuring the bone marrow harvest filter sets. This effort was supported by National Institutes of Health grants RO1 HL73714 (R.S.), RO1 HL60528 (R.S.) and Massachusetts General Hospital Wellman Center Advanced Microscopy startup fund (C.P.L.). This report is dedicated to the memory of Dr. Harvey R. Colten.

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Authors and Affiliations

  1. Departments of Dermatology and Medicine, Brigham and Women's Hospital and Harvard Skin Disease Research Center, Harvard Medical School, Boston, 02115, Massachusetts, USA

    Robert Sackstein, Jasmeen S Merzaban, Derek W Cain & Nilesh M Dagia

  2. Department of Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, 02115, Massachusetts, USA

    Robert Sackstein

  3. Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, 02115, Massachusetts, USA

    Joel A Spencer & Charles P Lin

  4. Research Specialties, Sigma-Aldrich, Buchs, CH-9470, Switzerland

    Roland Wohlgemuth

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  1. Robert Sackstein
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  7. Roland Wohlgemuth
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Contributions

R.S. conceived the study and reagents, created hybridomas, developed the SACK-1 mAb and the conditions for surface fucosylation of live cells, performed experiments and supervised all research, wrote the manuscript and funded the research; J.S.M., D.W.C. and N.M.D. performed cell culture, biochemical studies and adhesion assays; J.A.S. and C.P.L. performed intravital microscopy and C.P.L. partially funded the research; R.W. synthesized fucosyltransferase.

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Correspondence to Robert Sackstein.

Supplementary information

Supplementary Text and Figures

Supplementary Figs. 1–6 and Supplementary Methods (PDF 5151 kb)

Supplementary Movie 1

This video segment shows a marrow sinusoidal endothelial bed within the first minute after injection of FTVI-treated, HCELL-expressing MSCs (bright cells). Note evident rolling interactions and firm adherence of MSCs onto marrow sinusoidal endothelium. (MP4 3000 kb)

Supplementary Movie 2

This video segment shows a marrow sinusoidal endothelial bed withing the first minute after injection of FTVI-Sialidase MSCs. Compared to HCELL+ MSCs shown in Video 1, FTVI-Sialidase MSCs show minimal binding interactions with marrow sinusoidal endothelium. (MP4 3100 kb)

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Sackstein, R., Merzaban, J., Cain, D. et al. Ex vivo glycan engineering of CD44 programs human multipotent mesenchymal stromal cell trafficking to bone. Nat Med 14, 181–187 (2008). https://doi.org/10.1038/nm1703

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