Comparative In Vitro Study on Magnetic Iron Oxide Nanoparticles for MRI Tracking of Adipose Tissue-Derived Progenitor Cells

Magnetic resonance imaging (MRI) using measurement of the transverse relaxation time (R2*) is to be considered as a promising approach for cell tracking experiments to evaluate the fate of transplanted progenitor cells and develop successful cell therapies for tissue engineering. While the relationship between core composition of nanoparticles and their MRI properties is well studied, little is known about possible effects on progenitor cells. This in vitro study aims at comparing two magnetic iron oxide nanoparticle types, single vs. multi-core nanoparticles, regarding their physico-chemical characteristics, effects on cellular behavior of adipose tissue-derived stem cells (ASC) like differentiation and proliferation as well as their detection and quantification by means of MRI. Quantification of both nanoparticle types revealed a linear correlation between labeling concentration and R2* values. However, according to core composition, different levels of labeling concentrations were needed to achieve comparable R2* values. Cell viability was not altered for all labeling concentrations, whereas the proliferation rate increased with increasing labeling concentrations. Likewise, deposition of lipid droplets as well as matrix calcification revealed to be highly dose-dependent particularly regarding multi-core nanoparticle-labeled cells. Synthesis of cartilage matrix proteins and mRNA expression of collagen type II was also highly dependent on nanoparticle labeling. In general, the differentiation potential was decreased with increasing labeling concentrations. This in vitro study provides the proof of principle for further in vivo tracking experiments of progenitor cells using nanoparticles with different core compositions but also provides striking evidence that combined testing of biological and MRI properties is advisable as improved MRI properties of multi-core nanoparticles may result in altered cell functions.     

Human ASPM participates in spindle organisation, spindle orientation and cytokinesis

Background

For clinical applications of mesenchymal stem cells (MSCs), labeling and tracking is crucial to evaluate cell distribution and homing. Magnetic resonance imaging (MRI) has been successfully established detecting MSCs labeled with superparamagnetic particles of iron oxide (SPIO). Despite initial reports that labeling of MSCs with SPIO is safe without affecting the MSC's biology, recent studies report on influences of SPIO-labeling on metabolism and function of MSCs. Exposition of cells and tissues to high magnetic fields is the functional principle of MRI. In this study we established innovative labeling protocols for human MSCs using clinically established SPIO in combination with magnetic fields and investigated on functional effects (migration assays, quantification of colony forming units, analyses of gene and protein expression and analyses on the proliferation capacity, the viability and the differentiation potential) of magnetic fields on unlabeled and labeled human MSCs. To evaluate the imaging properties, quantification of the total iron load per cell (TIL), electron microscopy, and MRI at 3.0 T were performed.

Results

Human MSCs labeled with SPIO permanently exposed to magnetic fields arranged and grew according to the magnetic flux lines. Exposure of MSCs to magnetic fields after labeling with SPIO significantly enhanced the TIL compared to SPIO labeled MSCs without exposure to magnetic fields resulting in optimized imaging properties (detection limit: 1,000 MSCs). Concerning the TIL and the imaging properties, immediate exposition to magnetic fields after labeling was superior to exposition after 24 h. On functional level, exposition to magnetic fields inhibited the ability of colony formation of labeled MSCs and led to an enhanced expression of lipoprotein lipase and peroxisome proliferator-activated receptor-γ in labeled MSCs under adipogenic differentiation, and to a reduced expression of alkaline phosphatase in unlabeled MSCs under osteogenic differentiation as detected by qRT-PCR. Moreover, microarray analyses revealed that exposition of labeled MSCs to magnetic fields led to an up regulation of CD93 mRNA and cadherin 7 mRNA and to a down regulation of Zinc finger FYVE domain mRNA. Exposition of unlabeled MSCs to magnetic fields led to an up regulation of CD93 mRNA, lipocalin 6 mRNA, sialic acid acetylesterase mRNA, and olfactory receptor mRNA and to a down regulation of ubiquilin 1 mRNA. No influence of the exposition to magnetic fields could be observed on the migration capacity, the viability, the proliferation rate and the chondrogenic differentiation capacity of labeled or unlabeled MSCs.

Conclusions

In our study an innovative labeling protocol for tracking MSCs by MRI using SPIO in combination with magnetic fields was established. Both, SPIO and the static magnetic field were identified as independent factors which affect the functional biology of human MSCs. Further in vivo investigations are needed to elucidate the molecular mechanisms of the interaction of magnetic fields with stem cell biology.     

Effects of iron oxide incorporation for long term cell tracking on MSC differentiation in vitro and in vivo

Successful cell therapy will depend on the ability to monitor transplanted cells. With cell labeling, it is important to demonstrate efficient long term labeling without deleterious effects on cell phenotype and differentiation capacity. We demonstrate long term (7 weeks) retention of superparamagnetic iron oxide particles (SPIO) by mesenchymal stem cells (MSCs) in vivo, detectable by MRI. In vitro, multilineage differentiation (osteogenic, chondrogenic and adipogenic) was demonstrated by histological evaluation and molecular analysis in SPIO labeled and unlabeled cells. Gene expression levels were comaparable to unlabeled controls in adipogenic and chondrogenic conditions however not in the osteogenic condition. MSCs seeded into a scaffold for 21 days and implanted subcutaneously into nude mice for 4 weeks, showed profoundly altered phenotypes in SPIO labeled samples compared to implanted unlabeled control scaffolds, indicating chondrogenic differentiation. This study demonstrates long term MSC traceability using SPIO and MRI, uninhibited multilineage MSC differentiation following SPIO labeling, though with subtle but significant phenotypical alterations.     

Efficient In Vitro Labeling Rabbit Bone Marrow-Derived Mesenchymal Stem Cells with SPIO and Differentiating into Neural-Like Cells

Mesenchymal stem cells (MSCs) can differentiate into neural cells to treat nervous system diseases. Magnetic resonance is an ideal means for cell tracking through labeling cells with superparamagnetic iron oxide (SPIO). However, no studies have described the neural differentiation ability of SPIO-labeled MSCs, which is the foundation for cell therapy and cell tracking in vivo. Our results showed that bone marrow-derived mesenchymal stem cells (BM-MSCs) labeled in vitro with SPIO can be induced into neural-like cells without affecting the viability and labeling efficiency. The cellular uptake of SPIO was maintained after labeled BM-MSCs differentiated into neural-like cells, which were the basis for transplanted cells that can be dynamically and non-invasively tracked in vivo by MRI. Moreover, the SPIO-labeled induced neural-like cells showed neural cell morphology and expressed related markers such as NSE, MAP-2. Furthermore, whole-cell patch clamp recording demonstrated that these neural-like cells exhibited electrophysiological properties of neurons. More importantly, there was no significant difference in the cellular viability and [Ca²⁺]i between the induced labeled and unlabeled neural-like cells. In this study, we show for the first time that SPIO-labeled MSCs retained their differentiation capacity and could differentiate into neural-like cells with high cell viability and a good cellular state in vitro.     

Optimized labeling of bone marrow mesenchymal cells with superparamagnetic iron oxide nanoparticles and <Emphasis Type="Italic">in vivo</Emphasis> visualization by magnetic resonance imaging

Background  

Stem cell therapy has emerged as a promising addition to traditional treatments for a number of diseases. However, harnessing the therapeutic potential of stem cells requires an understanding of their fate in vivo. Non-invasive cell tracking can provide knowledge about mechanisms responsible for functional improvement of host tissue. Superparamagnetic iron oxide nanoparticles (SPIONs) have been used to label and visualize various cell types with magnetic resonance imaging (MRI). In this study we performed experiments designed to investigate the biological properties, including proliferation, viability and differentiation capacity of mesenchymal cells (MSCs) labeled with clinically approved SPIONs.     

Effects of iron oxide nanoparticles on cardiac differentiation of embryonic stem cells

The therapeutic potential of transplantation of embryonic stem cells (ESCs) in animal model of myocardial infarction has been consistently demonstrated. The development of superparamagnetic iron oxide (SPIO) nanoparticles labeling and cardiac magnetic resonance imaging (MRI) have been increasingly used to track the migration of transplanted cells in vivo allowing cell fate determination. However, the impact of SPIO- labeling on cell phenotype and cardiac differentiation capacity of ESCs remains unclear. In this study, we demonstrated that ESCs labeled with SPIO compared to their unlabeled counterparts had similar cardiogenic capacity, and SPIO-labeling did not affect calcium-handling property of ESC-derived cardiomyocytes. Moreover, transplantation of SPIO-labeled ESCs via direct intra-myocardial injection to infarct myocardium resulted in significant improvement in heart function. These findings demonstrated the feasibility of in vivo ESC tracking using SPIO-labeling and cardiac MRI without affecting the cardiac differentiation potential and functional properties of ESCs.     

Core–shell silver nanoparticles for optical labeling of cells

Silver nanoparticles have been modified with self-assembled monolayers of hydroxyl-terminated long chain thiols and encapsulated with a silica shell. The resulting core–shell nanoparticles were used as optical labels for cell analysis using flow cytometry and microscopy. The excitation of plasmon resonances in nanoparticles results in strong depolarized scattering of visible light, permitting detection at the single nanoparticle level. The nanoparticles were modified with neutravidin via epoxide–azide coupling chemistry, to which biotinylated antibodies targeting cell surface receptors were bound. The nanoparticle labels exhibited long-term stability in solutions with high salt concentrations without aggregation or silver etching. Labeled cells exhibited two orders of magnitude enhancement of the scattering intensity compared with unlabeled cells.     

Toxic Effect of Silica Nanoparticles on Endothelial Cells through DNA Damage Response via Chk1-Dependent G2/M Checkpoint

Silica nanoparticles have become promising carriers for drug delivery or gene therapy. Endothelial cells could be directly exposed to silica nanoparticles by intravenous administration. However, the underlying toxic effect mechanisms of silica nanoparticles on endothelial cells are still poorly understood. In order to clarify the cytotoxicity of endothelial cells induced by silica nanoparticles and its mechanisms, cellular morphology, cell viability and lactate dehydrogenase (LDH) release were observed in human umbilical vein endothelial cells (HUVECs) as assessing cytotoxicity, resulted in a dose- and time- dependent manner. Silica nanoparticles-induced reactive oxygen species (ROS) generation caused oxidative damage followed by the production of malondialdehyde (MDA) as well as the inhibition of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px). Both necrosis and apoptosis were increased significantly after 24 h exposure. The mitochondrial membrane potential (MMP) decreased obviously in a dose-dependent manner. The degree of DNA damage including the percentage of tail DNA, tail length and Olive tail moment (OTM) were markedly aggravated. Silica nanoparticles also induced G2/M arrest through the upregulation of Chk1 and the downregulation of Cdc25C, cyclin B1/Cdc2. In summary, our data indicated that the toxic effect mechanisms of silica nanoparticles on endothelial cells was through DNA damage response (DDR) via Chk1-dependent G2/M checkpoint signaling pathway, suggesting that exposure to silica nanoparticles could be a potential hazards for the development of cardiovascular diseases.     

Duration of in vitro storage affects the key stem cell features of human bone marrow-derived mesenchymal stromal cells for clinical transplantation

Background aimsMesenchymal stromal cells (MSCs) have the ability to self-renew and differentiate into various cell types. Their plasticity and easy availability make them promising candidates for regenerative medicine. However, for successful clinical application, MSCs need to be expanded under a Good Manufacturing Practices-compliant system to obtain a large quantity of these cells. Although the viability and potency of these in vitro-expanded MSCs need to be maintained during preparation and transportation before transplantation, these characteristics have not thoroughly been examined. Our goal in this study was to standardize MSC preparation and storage before their clinical application to ensure reproducible quality and potency for their clinically intended purpose.MethodsWe examined the viability, self-renewal capacity and differentiation capability of MSCs on short-term in vitro storage in saline or dextrose solution at 4°C and room temperature.ResultsMSCs harvested and suspended in saline for 1–2 h showed >90% viability regardless of storage temperature. However, when cells were stored for >2 h in saline, their viability decreased gradually over time. The viability of cells in dextrose deteriorated rapidly. MSCs lost colony-forming unit and differentiation capacities rapidly as storage time increased. Collectively, we found that a storage period >2 h resulted in a significant decrease in cell viability, cell proliferation capacity and differentiation potency.ConclusionsStorage of culture-harvested MSCs for >2 h is likely to result in suboptimal MSC-mediated tissue regeneration because of decreased cell viability and differentiation capacity.     

Implantation of Ferumoxides Labeled Human Mesenchymal Stem Cells in Cartilage Defects

The field of tissue engineering integrates the principles of engineering, cell biology and medicine towards the regeneration of specific cells and functional tissue. Matrix associated stem cell implants (MASI) aim to regenerate cartilage defects due to arthritic or traumatic joint injuries. Adult mesenchymal stem cells (MSCs) have the ability to differentiate into cells of the chondrogenic lineage and have shown promising results for cell-based articular cartilage repair technologies. Autologous MSCs can be isolated from a variety of tissues, can be expanded in cell cultures without losing their differentiation potential, and have demonstrated chondrogenic differentiation in vitro and in vivo^{1, 2}.In order to provide local retention and viability of transplanted MSCs in cartilage defects, a scaffold is needed, which also supports subsequent differentiation and proliferation. The architecture of the scaffold guides tissue formation and permits the extracellular matrix, produced by the stem cells, to expand. Previous investigations have shown that a 2% agarose scaffold may support the development of stable hyaline cartilage and does not induce immune responses³.Long term retention of transplanted stem cells in MASI is critical for cartilage regeneration. Labeling of MSCs with iron oxide nanoparticles allows for long-term in vivo tracking with non-invasive MR imaging techniques⁴.This presentation will demonstrate techniques for labeling MSCs with iron oxide nanoparticles, the generation of cell-agarose constructs and implantation of these constructs into cartilage defects. The labeled constructs can be tracked non-invasively with MR-Imaging.Open in a separate windowClick here to view.^{(27M, flv)}     

Organically modified silica nanoparticles are biocompatible and can be targeted to neurons in vivo

The application of nanotechnology in biological research is beginning to have a major impact leading to the development of new types of tools for human health. One focus of nanobiotechnology is the development of nanoparticle-based formulations for use in drug or gene delivery systems. However most of the nano probes currently in use have varying levels of toxicity in cells or whole organisms and therefore are not suitable for in vivo application or long-term use. Here we test the potential of a novel silica based nanoparticle (organically modified silica, ORMOSIL) in living neurons within a whole organism. We show that feeding ORMOSIL nanoparticles to Drosophila has no effect on viability. ORMOSIL nanoparticles penetrate into living brains, neuronal cell bodies and axonal projections. In the neuronal cell body, nanoparticles are present in the cytoplasm, but not in the nucleus. Strikingly, incorporation of ORMOSIL nanoparticles into the brain did not induce aberrant neuronal death or interfered with normal neuronal processes. Our results in Drosophila indicate that these novel silica based nanoparticles are biocompatible and not toxic to whole organisms, and has potential for the development of long-term applications.     

Effect of polymer grafting density on silica nanoparticle toxicity

Nanoparticles are commonly engineered with a layer of polymers on the surface used to increase their stability and biocompatibility, as well as providing multifunctional properties. Formulating the nanoparticle size and surface properties with polymers directly affects the way these nanoparticles interact with a biological system. Many previous studies have emphasized the importance of nanoparticle size and surface charge in affecting their toxicity in cells. However, the potential weakness in many of these studies is that the polymer grafting densities on nanoparticles have been disregarded during toxicity evaluation. In the current study, we hypothesized that the density of polymers on nanoparticles will affect their toxicity to cells, especially for nanoparticle cores that are toxic themselves. To address this issue, we synthesized a range of RAFT (reversible addition fragmentation chain transfer) polymers bearing different surface charges and coated them onto silica nanoparticles (SiNPs) with different grafting densities. The in vitro cytotoxicity of these SiNPs was evaluated using the MTT (thiazolyl blue tetrazolium bromide) assay with Caco-2 cells. We found that neutral (biocompatible) polymers with a high grafting density on SiNPs were effective at protecting the cells from the toxicity of the silica core. High cellular toxicity was only observed for cationic polymer-SiNPs, while all other neutral and anionic polymer-SiNPs induced limited cellular toxicity. In contrast, the toxic effects induced by low density polymer-coated SiNPs were mostly attributed to the silica core, while the polymer coatings had a limited contribution. These findings are important indicators for the future evaluation of the toxicological profile of polymer-coated nanoparticles.     

Isolation, proliferation, cytogenetic, and molecular characterization and in vitro differentiation potency of canine stem cells from foetal adnexa: a comparative study of amniotic fluid, amnion, and umbilical cord matrix

The possibility to isolate canine mesenchymal stem cells (MSCs) from foetal adnexa is interesting since several canine genetic disorders are reported to resemble similar dysfunctions in humans. In this study, we successfully isolated, cytogenetically and molecularly characterized, and followed the differentiation potency of canine MSCs from foetal adnexa, such as amniotic fluid (AF), amniotic membrane (AM), and umbilical cord matrix (UCM). In the three types of cell lines, the morphology of proliferating cells typically appeared fibroblast‐like, and the population doubling time (DT) significantly increased with passage number. For AF‐ and AM‐MSCs, cell viability did not change with passages. In UCM‐MSCs, cell viability remained at approximately constant levels up to P6 and significantly decreased from P7 (P < 0.05). Amnion and UCM‐MSCs expressed embryonic and MSC markers, such as Oct‐4 CD44, CD184, and CD29, whereas AF‐MSCs expressed Oct‐4, CD44. Expression of the hematopoietic markers CD34 and CD45 was not found. Dog leucocyte antigens (DLA‐DRA1 and DLA‐79) were expressed only in AF‐MSCs at P1. Isolated cells of the three cell lines at P3 showed multipotent capacity, and differentiated in vitro into neurocyte, adipocyte, osteocyte, and chondrocyte, as demonstrated by specific stains and expression of molecular markers. Cells at P4 showed normal chromosomal number, structure, and telomerase activity. These results demonstrate that, in dog, MSCs can be successfully isolated from foetal adnexa and grown in vitro. Their proven stemness and chromosomal stability indicated that MSCs could be used as a model to study stem cell biology and have an application in therapeutic programs. Mol. Reprod. Dev. 78:361–373, 2011. © 2011 Wiley‐Liss, Inc.     

Low-Intensity Ultrasound (LIUS) as an Innovative Tool for Chondrogenesis of Mesenchymal Stem Cells (MSCs)

Mesenchymal stem cells (MSCs) have a capacity to differentiate into the chondrogenic lineage and are a valuable allogenic source for cartilage tissue engineering. However, they still have critical limitations of relatively inefficient chondrogenic differentiation in vitro and of dedifferentiation and/or hypertrophic changes at late stages of differentiation. Numerous approaches using biochemical and mechanical factors have been tried but have so far failed to overcome these problems. Recent studies by other groups and ours have shown that low-intensity ultrasound (LIUS) is an efficient tool for promoting the chondrogenic differentiation of MSCs both in vitro and in vivo. A series of our experiments suggests that LIUS not only induces chondrogenic differentiation of MSCs but also has diverse additional activities that enhance the viability of MSCs, increase possibly the integrity of the differentiated tissues and delays hypertrophic changes during differentiation. Therefore, LIUS could be an innovative and versatile tool for chondrogenic differentiation of MSCs and for cartilage tissue engineering.     

Magnetic resonance imaging of ferumoxide-labeled mesenchymal stem cells in cartilage defects: in vitro and in vivo investigations

The purpose of this study was to (1) compare three different techniques for ferumoxide labeling of mesenchymal stem cells (MSCs), (2) evaluate if ferumoxide labeling allows in vivo tracking of matrix-associated stem cell implants (MASIs) in an animal model, and (3) compare the magnetic resonance imaging (MRI) characteristics of ferumoxide-labeled viable and apoptotic MSCs. MSCs labeled with ferumoxide by simple incubation, protamine transfection, or Lipofectin transfection were evaluated with MRI and histopathology. Ferumoxide-labeled and unlabeled viable and apoptotic MSCs in osteochondral defects of rat knee joints were evaluated over 12 weeks with MRI. Signal to noise ratios (SNRs) of viable and apoptotic labeled MASIs were tested for significant differences using t-tests. A simple incubation labeling protocol demonstrated the best compromise between significant magnetic resonance signal effects and preserved cell viability and potential for immediate clinical translation. Labeled viable and apoptotic MASIs did not show significant differences in SNR. Labeled viable but not apoptotic MSCs demonstrated an increasing area of T2 signal loss over time, which correlated to stem cell proliferation at the transplantation site. Histopathology confirmed successful engraftment of viable MSCs. The engraftment of iron oxide-labeled MASIs by simple incubation can be monitored over several weeks with MRI. Viable and apoptotic MASIs can be distinguished via imaging signs of cell proliferation at the transplantation site.     

MRI Detection of Nonproliferative Tumor Cells in Lymph Node Metastases Using Iron Oxide Particles in a Mouse Model of Breast Cancer

Cell tracking with magnetic resonance imaging (MRI) and iron nanoparticles is commonly used to monitor the fate of implanted cells in preclinical disease models. Few studies have employed these methods to study cancer cells because proliferative iron-labeled cancer cells will lose the label as they divide. In this study, we evaluate the potential for retention of the iron nanoparticle label, and resulting MRI signal, to serve as a marker for slowly dividing cancer cells. Green fluorescent protein-transfected MDA-MB-231 breast cancer cells were labeled with red fluorescent micron-sized superparamagnetic iron oxide (MPIO) nanoparticles. Cells were examined in vitro at multiple time points after labeling by staining for iron-labeled cells and by flow cytometric detection of the fluorescent MPIO. Severe combined immune deficiency (SCID) mice were implanted with 5 x 10⁵ MPIO-labeled or unlabeled cells in the mammary fat pad and MRI was performed weekly until 28 days after injection. Microscopy was performed to validate MRI. In vitro assays revealed a very small percentage of cells that retained MPIO at 14 days after labeling. Regions of signal loss were observed in MRI of primary tumors that developed from iron-labeled cancer cells. Small focal regions of signal loss were detected in images of the axillary and brachial nodes in six of eight mice, at day 14 or later, with microscopy confirming the presence of iron-labeled cancer cells. Our data suggest an interesting role for cell tracking with iron particles since label retention leads to persistent signal void, allowing proliferative status to be determined.     

Comparison of alternative mesenchymal stem cell sources for cell banking and musculoskeletal advanced therapies

With the continuous discovery of new alternative sources containing mesenchymal stem cells (MSCs), regenerative medicine therapies may find tailored applications in the clinics. Although these cells have been demonstrated to express specific mesenchymal markers and are able to differentiate into mesenchymal lineages in ad hoc culture conditions, it is still critical to determine the yield and differentiation potential of these cells in comparative studies under the same standardized culture environment. Moreover, the opportunity to use MSCs from bone marrow (BM) of multiorgan donors for cell banking is of relevant importance. In the attempt to establish the relative potential of alternative MSCs sources, we analyzed and compared the yield and differentiation potential of human MSCs from adipose and BM tissues of cadaveric origins, and from fetal annexes (placenta and umbilical cord) after delivery using standardized isolation and culture protocols. BM contained a significantly higher amount of mononuclear cells (MNCs) compared to the other tissue sources. Nonetheless, a higher cell seeding density was needed for these cells to successfully isolate MSCs. The MNCs populations were highly heterogeneous and expressed variable MSCs markers with a large variation from donor to donor. After MSCs selection through tissue culture plastic adhesion, cells displayed a comparable proliferation capacity with distinct colony morphologies and were positive for a pool of typical MSCs markers. In vitro differentiation assays showed a higher osteogenic differentiation capacity of adipose tissue and BM MSCs, and a higher chondrogenic differentiation capacity of BM MSCs.     

Dose-Response of Superparamagnetic Iron Oxide Labeling on Mesenchymal Stem Cells Chondrogenic Differentiation: A Multi-Scale In Vitro Study

Aim

The aim of this work was the development of successful cell therapy techniques for cartilage engineering. This will depend on the ability to monitor non-invasively transplanted cells, especially mesenchymal stem cells (MSCs) that are promising candidates to regenerate damaged tissues.

Methods

MSCs were labeled with superparamagnetic iron oxide particles (SPIO). We examined the effects of long-term labeling, possible toxicological consequences and the possible influence of progressive concentrations of SPIO on chondrogenic differentiation capacity.

Results

No influence of various SPIO concentrations was noted on human bone marow MSC viability or proliferation. We demonstrated long-term (4 weeks) in vitro retention of SPIO by human bone marrow MSCs seeded in collagenic sponges under TGF-β1 chondrogenic conditions, detectable by Magnetic Resonance Imaging (MRI) and histology. Chondrogenic differentiation was demonstrated by molecular and histological analysis of labeled and unlabeled cells. Chondrogenic gene expression (COL2A2, ACAN, SOX9, COL10, COMP) was significantly altered in a dose-dependent manner in labeled cells, as were GAG and type II collagen staining. As expected, SPIO induced a dramatic decrease of MRI T2 values of sponges at 7T and 3T, even at low concentrations.

Conclusions

This study clearly demonstrates (1) long-term in vitro MSC traceability using SPIO and MRI and (2) a deleterious dose-dependence of SPIO on TGF-β1 driven chondrogenesis in collagen sponges. Low concentrations (12.5–25 µg Fe/mL) seem the best compromise to optimize both chondrogenesis and MRI labeling.