Biochemical and Biophysical Research Communications
Effects of iron oxide incorporation for long term cell tracking on MSC differentiation in vitro and in vivo
Section snippets
Experimental protocol
Cell culture. Human bone marrow stromal cells (BMSCs) were isolated from femoral shaft biopsies of a patient undergoing total hip replacement after written consent (MEC-2004-142). BMSCs were isolated from the aspirated marrow according to previously described procedures [14]. Cells from the fourth passage were used for the experiments. Mesenchymal stem cells purchased from Cambrex (Verviers, Belgium) were used in the in vivo study.
Cell labeling. Cells were labeled with Endorem (Guerbet S.A.,
Multilineage differentiation of adult human MSCs
To determine the multilineage potential of labeled cells, SPIO labeled and unlabeled MSCs were cultured either in control medium or medium supplemented with adipogenic, chondrogenic or osteogenic factors for 13 days. Lineage specific histological staining was then performed. For both labeled and unlabeled MSCs, 13 days culture in appropriate differentiation medium led to positive staining for adipogenic (Fig. 1C and D), chondrogenic (Fig. 1H and I) and osteogenic (Fig. 1M and N), demonstrated
Adipogenesis
Following culture with adipogenic factors, significantly elevated levels of fatty acid binding protein-4 (FABP4), also known as aP2, were observed compared to unlabeled, untreated control cultures for both labeled and unlabeled cells (Fig. 2A; P = 0.0001, Two way ANOVA with repeated measures, n = 3). SPIO labeling had no effect on this expression. The same effect was observed with peroxisome proliferator-acvitated receptor γ (PPARγ), another marker of adipogenesis. Fig. 4B shows a significant
Discussion
Several groups have demonstrated the differentiation potential of stem cells or progenitor cells following SPIO labeling using different transfection methods [19], [20], [21], [22]. However, these papers have not investigated chondrogenic differentiation, nor have they looked beyond basic histological/immunohistological analyses or at multilineage MSC differentiation. A recent paper by Heymer et al. performed gene expression analyses but did not examine the effects of in vivo implantation on
Acknowledgments
We thank Nicole Kops for her assistance with the histological staining of implanted scaffolds, Corina de Ridder for her assistance with the implantation of the collagen-GAG scaffolds and Gavin Houston, Applied Science Laboratory, GE Healthcare for critical review of the manuscript and for helping with the optimization of data acquisition. We also acknowledge financial support from the Programme for Research in Third Level Institutions, The Science Foundation Ireland’s President of Ireland Young
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Both authors have contributed equally.