Abstract
Imaging technologies that allow the non-invasive monitoring of stem cells in vivo play a vital role in cell-based regenerative therapies. Recently, much interest has been generated in reporter genes that enable simultaneous monitoring of the anatomical location and viability of cells using magnetic resonance imaging (MRI). Here, we investigate the efficacy of ferritin heavy chain-1 (Fth1) and transferrin receptor-1 (TfR1) as reporters for tracking mesenchymal stem cells. The overexpression of TfR1 was well tolerated by the cells but Fth1 was found to affect the cell’s iron homeostasis, leading to phenotypic changes in the absence of iron supplementation and an upregulation in transcript and protein levels of the cell’s endogenous transferrin receptor. Neither the sole overexpression of Fth1 nor TfR1 resulted in significant increases in intracellular iron content, although significant differences were seen when the two reporter genes were used in combination, in the presence of high concentrations of iron. The supplementation of the culture medium with iron sources was a more efficient means to obtain contrast than the use of reporter genes, where high levels of intracellular iron were reflected in transverse (T2) relaxation. The feasibility of imaging iron-supplemented cells by MRI is shown using a 3R-compliant chick embryo model.
Highlights
The development of imaging techniques that allow the monitoring of a cell’s anatomical location in vivo has seen great progress in recent years
The actual iron internalised by cells when supplemented for 24 h with ferric citrate is displayed in Figure 2B; by supplementing the culture medium with ferric citrate the intracellular iron content can be increased from 0.03 to 0.15 pg/cell
We have investigated the effects of co-supplementing the medium with ascorbic acid, holo-transferrin or a combination of those, each of which allowed a stepwise increase in the intracellular iron content of
Summary
The development of imaging techniques that allow the monitoring of a cell’s anatomical location in vivo has seen great progress in recent years. The heavy subunit incorporates iron several fold faster than the light subunit [12,13] These two genes (TfR1 and Fth1) have been previously explored as reporter genes for the MRI of cancer cells [14,15,16,17] and stem cells [18,19,20,21]. Iron accumulation in the target cells is usually achieved by overexpression of these proteins via chromosomal insertion of the respective genes under the control of a constitutive promoter Some of these studies have displayed limited success in animal models, no reports exist so far on the potential of using a combination of these two genes in mesenchymal stem cells (MSCs). We explore the interplay between exogenous and endogenous iron regulatory proteins, and determine the extent to which MRI reporter genes can improve contrast in MSCs
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