Background

[PubMed]

Magnetic iron microbeads coupled with HEA-125 monoclonal antibody against the epithelial cell adhesion molecule (EpCAM), abbreviated as EpCAM microbeads, have been developed primarily for the positive selection or depletion of EpCAM-positive cells (1, 2). McClelland et al. have demonstrated the feasibility of cell tracking with magnetic resonance imaging (MRI) after the hepatic progenitor cells are labeled with EpCAM microbeads (1).

Superparamagnetic iron oxide (SPIO)-based agents have been intensively tested for use in cell tracking with MRI in both preclinical and clinical situations (3, 4). SPIO particles provide a strong change in signal per unit of metal in T2-weighted images without a significant effect on labeled cells and host (3, 5). For efficient cell labeling, SPIO particles are generally coated with low-molecular-weight polymers or dendrimers leading to clusters of electron-dense crystal cores covered with the polymer or dendrimer. Surface coating increases the stability of SPIO particles and allows further chemical modification of the particles with targeting ligands. Cell tracking studies have shown that the migration and homing capabilities of SPIO-labeled cells can be monitored in vivo over days to months (3, 4, 6).

Nevertheless, there are still many challenges to overcome before MRI cell tracking can be considered a robust technique in preclinical settings or in clinical applications (3, 7). MRI detects the presence of SPIO contrast agents, regardless of whether SPIO particles remain in the relevant cells, are lost to the extracellular matrix, or are transferred to other cells. It is still not possible to use MRI to discriminate live cells from dead cells or relevant cells from phagocytes. Detection sensitivity also becomes an issue when cells actively divide and migrate, in which case the SPIO labels are quickly divided among daughter cells to levels that are undetectable with MRI. In vivo quantification of the cell number is more challenging because of the contrast agent dilution during cell division, contrast agent transfer to others cells, other sources of iron in tissue, and technical limitations (4, 7).

McClelland et al. addressed the problem of contrast agent dilution in MRI cell tracking by using EpCAM microbeads as the label (1). The human EpCAM, also known as CD326 or epithelial-specific antigen, is a cell-surface antigen and is found on hepatic progenitor cells, including human hepatic stem cells (hHpSCs) and hepatoblasts (hHBs), on liver cancer stem cells, and on proliferating epithelial cells in other tissues (8-10). The human EpCAM is not expressed in animal cells. McClelland et al. studied the labeling of hHpSCs with EpCAM microbeads in vitro, ex vivo, and in vivo and imaged the labeled cells with MRI (1). The investigators demonstrated that the hHpSCs could be labeled with EpCAM microbeads before or after transplantation, and the transplanted hHpSCs could be monitored and counted repeatedly in the same host by injection of the label just prior to MRI (1).

Synthesis

[PubMed]

EpCAM microbeads (brand name, CD326 MicroBeads) are commercially available and have an overall diameter of 50–100 nm (1). The magnetic iron microbeads were coupled with HEA-125 monoclonal antibodies against the human EpCAM. The amount of monoclonal antibodies per microbead was not reported.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

Cell labeling with EpCAM microbeads was analyzed after the hHpSCs, prepared from human fetal livers, were exposed to the beads at a concentration of 10 µl EpCAM microbeads per 10⁷ cells in 100 µl buffer for 40 min at 4°C (1). After labeling, excess microbeads were washed, and the cells were cultured for 24 more hours. Transmission electron microscopy, scanning electron microscopy, and dispersive x-ray imaging confirmed the cell internalization and cell surface attachment of the EpCAM microbeads. The labeling efficiency. i.e., the amount of Fe or beads per cell, and the effect of labeling on cell function were not reported.

Detection feasibility with MRI was tested with labeled and unlabeled cell aggregates embedded in 1% agarose on 35-mm Petri dishes (1). In contrast to the controls without cells and with unlabeled cells, gel that contained labeled cells showed conspicuously large clusters of signal voids of varying sizes, which were attributed to the cell aggregates of varying sizes. Detection sensitivity was not reported.

McClelland et al. established an MRI method for both in vitro and in vivo quantification of the cell number (1). Three-dimensional aggregates of the hHpSCs were generated by placing cells in culture dishes. The radius, diameter, and area of the cell aggregates were measured under microscopy as well as with MRI, assuming that a single hHpSC has a radius of ~8 μm and that the descendents of hHpSCs (the hHBs) have a single-cell radius of ~11 μm. A strong correlation (r² = 0.99) was obtained between the cell aggregate radii obtained under microscopy and those measured with MRI. The cell aggregates appeared larger (64% larger in this case) in MRI than in actuality because the influence of the magnetic beads that give rise to the MRI contrast usually extends beyond their physical boundary. On the basis of the cell number and cell aggregate radius measured under microscopy, equations for the cell count (N) in terms of MRI-observed aggregate radius (r_MRI) were then established as N ≈ 104 + 4.2E – 4r³_MRI for spherical aggregates and N ≈ 104 + 1.0E – 4V_MRI for non-spherical aggregates, where the E represents the power of 10 and the V_MRI is the volume of the MRI region of interest. Cell numbers could be computed directly from the in vivo MRI images with the use of these equations (1).

Animal Studies

Human Studies

[PubMed]

No references are currently available.

References

1.

McClelland, R., E. Wauthier, T. Tallheden, L.M. Reid, and E. Hsu, In Situ Labeling and Magnetic Resonance Imaging of Transplanted Human Hepatic Stem Cells. Mol Imaging Biol, 2010. [PMC free article: PMC3727160] [PubMed: 20890665]

2.

Bulte J.W., Douglas T., Witwer B., Zhang S.C., Strable E., Lewis B.K., Zywicke H., Miller B., van Gelderen P., Moskowitz B.M., Duncan I.D., Frank J.A. Magnetodendrimers allow endosomal magnetic labeling and in vivo tracking of stem cells. Nat Biotechnol. 2001;19(12):1141–7. [PubMed: 11731783]

3.

Arbab A.S., Liu W., Frank J.A. Cellular magnetic resonance imaging: current status and future prospects. Expert Rev Med Devices. 2006;3(4):427–39. [PubMed: 16866640]

4.

Bulte J.W. In vivo MRI cell tracking: clinical studies. AJR Am J Roentgenol. 2009;193(2):314–25. [PMC free article: PMC2857985] [PubMed: 19620426]

5.

Himmelreich U., Dresselaers T. Cell labeling and tracking for experimental models using magnetic resonance imaging. Methods. 2009;48(2):112–24. [PubMed: 19362150]

6.

Kiessling, F., Noninvasive cell tracking. Handb Exp Pharmacol, 2008(185 Pt 2): p. 305-21. [PubMed: 18626608]

7.

Srinivas M., Aarntzen E.H., Bulte J.W., Oyen W.J., Heerschap A., de Vries I.J., Figdor C.G. Imaging of cellular therapies. Adv Drug Deliv Rev. 2010;62(11):1080–93. [PubMed: 20800081]

8.

van der Gun B.T., Melchers L.J., Ruiters M.H., de Leij L.F., McLaughlin P.M., Rots M.G. EpCAM in carcinogenesis: the good, the bad or the ugly. Carcinogenesis. 2010;31(11):1913–21. [PubMed: 20837599]

9.

Trzpis M., McLaughlin P.M., de Leij L.M., Harmsen M.C. Epithelial cell adhesion molecule: more than a carcinoma marker and adhesion molecule. Am J Pathol. 2007;171(2):386–95. [PMC free article: PMC1934518] [PubMed: 17600130]

10.

Munz M., Baeuerle P.A., Gires O. The emerging role of EpCAM in cancer and stem cell signaling. Cancer Res. 2009;69(14):5627–9. [PubMed: 19584271]