mRNA-mediated glycoengineering ameliorates deficient homing of human stem cell–derived hematopoietic progenitors
Jungmin Lee, … , Robert Sackstein, Derrick J. Rossi
Jungmin Lee, … , Robert Sackstein, Derrick J. Rossi
Published June 1, 2017; First published May 8, 2017
Citation Information: J Clin Invest. 2017;127(6):2433-2437. https://doi.org/10.1172/JCI92030.
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Brief Report Stem cells Transplantation

mRNA-mediated glycoengineering ameliorates deficient homing of human stem cell–derived hematopoietic progenitors

Abstract

Generation of functional hematopoietic stem and progenitor cells (HSPCs) from human pluripotent stem cells (PSCs) has been a long-sought-after goal for use in hematopoietic cell production, disease modeling, and eventually transplantation medicine. Homing of HSPCs from bloodstream to bone marrow (BM) is an important aspect of HSPC biology that has remained unaddressed in efforts to derive functional HSPCs from human PSCs. We have therefore examined the BM homing properties of human induced pluripotent stem cell–derived HSPCs (hiPS-HSPCs). We found that they express molecular effectors of BM extravasation, such as the chemokine receptor CXCR4 and the integrin dimer VLA-4, but lack expression of E-selectin ligands that program HSPC trafficking to BM. To overcome this deficiency, we expressed human fucosyltransferase 6 using modified mRNA. Expression of fucosyltransferase 6 resulted in marked increases in levels of cell surface E-selectin ligands. The glycoengineered cells exhibited enhanced tethering and rolling interactions on E-selectin–bearing endothelium under flow conditions in vitro as well as increased BM trafficking and extravasation when transplanted into mice. However, glycoengineered hiPS-HSPCs did not engraft long-term, indicating that additional functional deficiencies exist in these cells. Our results suggest that strategies toward increasing E-selectin ligand expression could be applicable as part of a multifaceted approach to optimize the production of HSPCs from human PSCs.

Authors

Jungmin Lee, Brad Dykstra, Joel A. Spencer, Laurie L. Kenney, Dale L. Greiner, Leonard D. Shultz, Michael A. Brehm, Charles P. Lin, Robert Sackstein, Derrick J. Rossi

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Figure 2

FUT6 modified mRNA–mediated glycoengineering enhances tethering and rolling of hiPS-HSPCs on endothelial cells under shear conditions.

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FUT6 modified mRNA–mediated glycoengineering enhances tethering and roll...
(A) Representative histograms showing expression of sLex on hiPS-HSPCs and PBMC control cells measured by HECA452 antibody. (B) Representative histograms showing expression of sLex on hiPS-HSPCs 24 hours after transfection with modified mRNAs encoding FUT3, FUT6, and FUT7. (C) Western blot with an E-selectin–Ig chimera or β-actin (loading control) on lysates of hiPS-HSPCs cells mock transfected or transfected with the indicated modified mRNAs 2 days earlier. (D) Quantitation of E-selectin–mediated rolling of control or FUT6 modified mRNA–transfected hiPS-HSPCs on TNF-α–activated HUVECs under increasing shear stress. E-selectin blocking antibody reduces rolling to baseline, while no interaction was observed on HUVECs not activated with TNF-α. n = 3. (E) FUT6 modified mRNA–transfected hiPS-HSPCs exhibit reduced rolling velocities on TNF-α–activated HUVECs. n = 3. Error bars indicate SEM. *P < 0.05, **P < 0.01 by Student’s t test.