Fluorophore-conjugated iron oxide nanoparticle labeling and analysis of engrafting human hematopoietic stem cells.
The use of nanometer-sized iron oxide particles combined with molecular imaging techniques enables dynamic studies of homing and trafficking of human hematopoietic stem cells (HSC). Identifying clinically applicable strategies for loading nanoparticles into primitive HSC requires strictly defined culture conditions to maintain viability without inducing terminal differentiation. In the current study, fluorescent molecules were covalently linked to dextran-coated iron oxide nanoparticles (Feridex) to characterize human HSC labeling to monitor the engraftment process. Conjugating fluorophores to the dextran coat for fluorescence-activated cell sorting purification eliminated spurious signals from nonsequestered nanoparticle contaminants. A short-term defined incubation strategy was developed that allowed efficient labeling of both quiescent and cycling HSC, with no discernable toxicity in vitro or in vivo. Transplantation of purified primary human cord blood lineage-depleted and CD34(+) cells into immunodeficient mice allowed detection of labeled human HSC in the recipient bones. Flow cytometry was used to precisely quantitate the cell populations that had sequestered the nanoparticles and to follow their fate post-transplantation. Flow cytometry endpoint analysis confirmed the presence of nanoparticle-labeled human stem cells in the marrow. The use of fluorophore-labeled iron oxide nanoparticles for fluorescence imaging in combination with flow cytometry allows evaluation of labeling efficiencies and homing capabilities of defined human HSC subsets.
Molecular Imaging Center, Mallinckrodt Institute of Radiology, Department of Molecular Biology and Pharmacology, Washington University School of Medicine, St. Louis, Missouri, USA.
Dustin J
DJ
Maxwell
Jesper
J
Bonde
David A
DA
Hess
Sarah A
SA
Hohm
Ryan
R
Lahey
Ping
P
Zhou
Michael H
MH
Creer
David
D
Piwnica-Worms
Jan A
JA
Nolta
D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D002453:Cell Cycle; D002470:Cell Survival; D003114:Colony-Forming Units Assay; D003911:Dextrans; D052203:Ferrosoferric Oxide; D005456:Fluorescent Dyes; D006085:Graft Survival; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000145:classification / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D007501:Iron / Q000493:pharmacokinetics; D058185:Magnetite Nanoparticles; D053768:Metal Nanoparticles; D051379:Mice; D016688:Mice, Inbred NOD; D018345:Mice, Knockout; D016513:Mice, SCID; D010087:Oxides / Q000493:pharmacokinetics; D014183:Transplantation, Heterologous
D018952:Antigens, CD34; D003911:Dextrans; D005456:Fluorescent Dyes; D058185:Magnetite Nanoparticles; D010087:Oxides; D007501:Iron; C065507:ferumoxides; D052203:Ferrosoferric Oxide