Transferrin receptor upregulation: in vitro labeling of rat mesenchymal stem cells with superparamagnetic iron oxide.
OBJECTIVE
To prospectively evaluate the influence of superparamagnetic iron oxide (SPIO) or ultrasmall SPIO (USPIO) particles on the surface epitope pattern of adult mesenchymal stem cells (MSCs) by regulating the expression of transferrin receptor and to prospectively evaluate the influence of transfection agents (TAs) on the uptake of SPIO or USPIO particles in MSCs.


METHODS
The study was approved by the institutional animal care committee of the University of Tübingen. MSCs were isolated from the bone marrow of four rats. To obtain highly homogeneous MSC populations, MSCs from one rat were single-cell cloned. One MSC clone was characterized and selected for the labeling experiments. The MSCs, which were characterized with flow cytometry and in vitro differentiation, were labeled with 200 microg/mL SPIO or USPIO or with 60 microg/mL SPIO or USPIO in combination with TAs. Aggregations of labeled cells were accommodated inside a defined volume in an agar gel matrix. Magnetic resonance (MR) imaging was performed to measure SPIO- or USPIO-induced signal voids. Quantification of cellular total iron load (TIL) (intracellular iron plus iron coating the cellular surface), determination of cellular viability, and electron microscopy were also performed.


RESULTS
Labeling of MSCs with SPIO or USPIO was feasible without affecting cell viability (91.1%-94.7%) or differentiation potential. For MR imaging, SPIO plus a TA was most effective, depicting 5000 cells with an average TIL of 76.5 pg per cell. SPIO or USPIO particles in combination with TAs coated the cellular surface but were not incorporated into cells. In nontransfected cells, SPIO or USPIO was taken up. MSCs labeled with SPIO or USPIO but without a TA showed enhanced expression of transferrin receptor, in contrary to both MSCs labeled with SPIO or USPIO and a TA and control cells.


CONCLUSIONS
SPIO or USPIO labeling without TAs has an influence on gene expression of MSCs upregulating transferrin receptor. Furthermore, SPIO labeling with a TA will coat the cellular surface.
Schäfer
Richard
R
Institute of Clinical and Experimental Transfusion Medicine, University Medical Center Tübingen, Hoppe-Seyler-Str 3, D-72076 Tübingen, Germany.
Kehlbach
Rainer
R
Wiskirchen
Jakub
J
Bantleon
Rüdiger
R
Pintaske
Jörg
J
Brehm
Bernhard R
BR
Gerber
Annika
A
Wolburg
Hartwig
H
Claussen
Claus D
CD
Northoff
Hinnak
H
D000818:Animals; D003287:Contrast Media / Q000493:pharmacokinetics*; D003911:Dextrans; D005240:Feasibility Studies; D052203:Ferrosoferric Oxide; D005434:Flow Cytometry; D015870:Gene Expression / Q000187:drug effects; D007501:Iron / Q000493:pharmacokinetics*; D008279:Magnetic Resonance Imaging / Q000379:methods*; D058185:Magnetite Nanoparticles; D008297:Male; D059630:Mesenchymal Stem Cells; D010087:Oxides / Q000493:pharmacokinetics*; D010316:Particle Size; D011446:Prospective Studies; D051381:Rats; D017207:Rats, Sprague-Dawley; D011990:Receptors, Transferrin / Q000378:metabolism*; D014162:Transfection; D015854:Up-Regulation
D003287:Contrast Media; D003911:Dextrans; D058185:Magnetite Nanoparticles; D010087:Oxides; D011990:Receptors, Transferrin; D007501:Iron; C065507:ferumoxides; D052203:Ferrosoferric Oxide