Superparamagnetic Iron Oxide Nanoparticle Labeling Research Articles

MRI tracking of human Wharton's jelly stem cells seeded onto acellular dermal matrix labeled with superparamagnetic iron oxide nanoparticles in burn wounds.

In vivo cell tracking after transplantation in regenerative medicine remains an unmet challenge and limits current understanding of the wound healing mechanism through cell-based therapies. This study investigated tracking of human Wharton's jelly stem cells (hWJSCs) seeded onto an acellular dermal matrix (ADM) and labeled with superparamagnetic iron oxide nanoparticles (SPIONs) by magnetic resonance imaging (MRI) in burn injury. The hWJSCs were characterized and assessed for growth kinetics. A total of 30 rats were enrolled in three equal groups. Group 1 underwent scald burn injury left without treatment, the group 2 was treated by an ADM that was prepared from cosmetic surgery skin samples and the group 3 received hWJSCs labeled with SPIONs seeded onto an ADM. Tensile strength was evaluated before and after interventions, real time PCR assessed apoptosis, and Prussian blue staining, scanning electron microscopy (SEM) and MRI were used for the tracking of labeled cells. The hWJSCs exhibited mesenchymal stem cell properties. Population doubling time was 40.1 hours. SPIONs did not show any toxic effect. The hWJSCs seeded onto an ADM decreased Bax and increased Bcl-2 gene expression. Internalization of SPIONs within hWJSCs was confirmed by Prussian blue staining, SEM and MRI until day 21. There was a significant difference between the Young's moduli of normal skin and the group receiving hWJSCs seeded onto an ADM. Histological observations and SEM imaging confirmed that MRI is an accurate method to track SPION-labeled hWJSCs in vivo. This study showed that SPION labeling coupled with MRI can be used to further understand the fate of stem cells after transplantation in a burn model.

Research on treating atherosclerotic erectile dysfunction by magnetic adipose-derived stem cells

Objective To explore the effect of magnetic adipose-derived stem cells(ADSCs) on treating atherosclerotic erectile dysfunction(ASED). Methods ADSCs were extracted from para-epididymis fat of SD rats aged 2 to 3 months for culture and identification. ADSCs were labeled with the magnetic resonance contrast agent superparamagnetic iron oxide nanoparticles(SPIONs). Meanwhile, the labeling efficiency, as well as effect on cell viability and proliferation was evaluated using Prussian blue, trypan blue staining and MTT method. The paramagnetism of ADSCs after SPIONs labeling was evaluated through the action of the exterior magnetic field. The atherosclerosis rat model was constructed, and ADSCs were injected into the corpus cavernosum afterwards. Intracorporeal pressure(ICP) was determined in the normal group, atherosclerosis group, ADSCs treatment with no action of magnetic field group, and ADSCs treatment with action of exterior magnetic field group after 4 weeks.Contents of smooth muscle and endothelium in corpus cavernosum of rats were detected using immunohistochemical staining. In addition, SOD and NOS activities, as well as MDA content were detected by ELISA. Results ADSCs extracted from para-epididymis fat of rats had grown actively and had high purity. Besides, they showed multi-lineage differentiation. SPIONs could be effectively absorbed by ADSCs, which would not remarkably inhibit the proliferation and cell viability of ADSCs. Iron content in cell was positively correlated with the iron concentration in the labeled SPIONs, and ADSCs showed paramagnetism after SPIONs labeling. ADSCs transplantation could notably improve the erectile function of ASED rats and enhanced the endothelium and smooth muscle contents in corpus cavernosum. Moreover, it could also correct the oxidative stress state in corpus cavernosum tissue, and some ADSCs had differentiated into smooth muscle and endothelial cells. Conclusions ADSCs transplantation can markedly improve the erectile function of ASED rats. Furthermore, magnetic induction has promoted the colonization of ADSCs in the corpus cavernosum, which contributes to enhancing the therapeutic effect of ADSCs and provides a new thinking and concept for stem cell therapy. Key words: Plaque, Atherosclerotic; Erectile Dysfunction; Mesenchymal Stem Cells

MRI-Tracking of Dental Pulp Stem Cells In Vitro and In Vivo Using Dextran-Coated Superparamagnetic Iron Oxide Nanoparticles.

The aim of this study was to track dental pulp stem cells (DPSCs) labeled with dextran-coated superparamagnetic iron oxide nanoparticles (SPIONs) using magnetic resonance imaging (MRI). Dental pulp was isolated from male Sprague Dawley rats and cultured in Dulbecco’s modified Eagle’s medium F12 (DMEM-F12) and 10% fetal bovine serum. Effects of SPIONs on morphology, viability, apoptosis, stemness, and osteogenic and adipogenic differentiation of DPSCs were assessed. Prussian blue staining and MRI were conducted to determine in vitro efficiency of SPIONs uptake by the cells. Both non-labeled and labeled DPSCs were adherent to culture plates and showed spindle-shape morphologies, respectively. They were positive for osteogenic and adipogenic induction and expression of cluster of differentiation (CD) 73 and CD90 biomarkers, but negative for expression of CD34 and CD45 biomarkers. The SPIONs were non-toxic and did not induce apoptosis in doses less than 25 mg/mL. Internalization of the SPIONs within the DPSCs was confirmed by Prussian blue staining and MRI. Our findings revealed that the MRI-based method could successfully monitor DPSCs labeled with dextran-coated SPIONs without any significant effect on osteogenic and adipogenic differentiation, viability, and stemness of DPSCs. We provided the in vitro evidence supporting the feasibility of an MRI-based method to monitor DPSCs labeled with SPIONs without any significant reduction in viability, proliferation, and differentiation properties of labeled cells, showing that internalization of SPIONs within DPSCs were not toxic at doses less than 25 mg/mL. In general, the SPION labeling does not seem to impair cell survival or differentiation. SPIONs are biocompatible, easily available, and cost effective, opening a new avenue in stem cell labeling in regenerative medicine.

Magnetic Iron Oxide Nanoparticle Labeling of Photoreceptor Precursors for Magnetic Resonance Imaging.

This study describes the methods and results of superparamagnetic iron oxide nanoparticle (SPION) labeling and magnetic resonance imaging (MRI) tracking of human embryonic stem cell-derived photoreceptor precursors transplanted into the subretinal space of Royal College of Surgeons rats. SPION labeling and MRI tracking provide information about the biodistribution of transplanted photoreceptor precursors, which is necessary for improving the functional benefits of cell therapy for degenerative retinal diseases.

Superparamagnetic Iron Oxide Nanoparticles as Magnetic Resonance Imaging Contrast Agents and Induced Autophagy Response in Endothelial Progenitor Cells.

Endothelial progenitor cells (EPCs) have been used in clinical trials for the treatment of patients with ischemic diseases, such as myocardial infarction and limb ischemia, as they could secrete angiogenic growth factors and promote neovascularization. However, the clinical employment of EPC therapy relies on the establishment of a noninvasive, timely, effective, and safe assessment system. In this study, we recruited superparamagnetic iron oxide nanoparticles (SPION) enhanced magnetic resonance imaging (MRI) to track EPCs in vitro. We evaluated the labeling efficiency of SPION in EPCs without cationic transfection reagents and determined the threshold value of the labeled cell quantity for MRI. Our data suggested that 1×105 EPCs incubated with SPION at a concentration of 100 μg Fe/mL for 24 h could be detected with a clinical 3T MRI T₂ scan. SPION were not cytotoxic for EPCs; however, SPION induced autophagy and impaired neovascularization. We then examined if the impairment could be alleviated through the inhibition of autophagy. The observed autophagy response and its function in EPCs may provide a strategy to improve the safety of SPION labeling. It may also verify that the assessment of autophagy may be a more sensitive detection method to evaluate the biocompatibility of tracking reagents.

Functionalized superparamagnetic iron oxide nanoparticles provide highly efficient iron-labeling in macrophages for magnetic resonance–based detection in vivo

Background aimsTracking cells during regenerative cytotherapy is crucial for monitoring their safety and efficacy. Macrophages are an emerging cell-based regenerative therapy for liver disease and can be readily labeled for medical imaging. A reliable, clinically applicable cell-tracking agent would be a powerful tool to study cell biodistribution. MethodsUsing a recently described chemical design, we set out to functionalize, optimize and characterize a new set of superparamagnetic iron oxide nanoparticles (SPIONs) to efficiently label macrophages for magnetic resonance imaging–based cell tracking in vivo. ResultsA series of cell health and iron uptake assays determined that positively charged SPIONs (+16.8 mV) could safely label macrophages more efficiently than the formerly approved ferumoxide (−6.7 mV; Endorem) and at least 10 times more efficiently than the clinically approved SPION ferumoxytol (−24.2 mV; Rienso). An optimal labeling time of 4 h at 25 µg/mL was demonstrated to label macrophages of mouse and human origin without any adverse effects on cell viability whilst providing substantial iron uptake (>5 pg Fe/cell) that was retained for 7 days in vitro. SPION labeling caused no significant reduction in phagocytic activity and a shift toward a reversible M1-like phenotype in bone marrow–derived macrophages (BMDMs). Finally, we show that SPION-labeled BMDMs delivered via the hepatic portal vein to mice are localized in the hepatic parenchyma resulting in a 50% drop in T2* in the liver. Engraftment of exogenous cells was confirmed via immunohistochemistry up to 3 weeks posttransplantation. DiscussionA positively charged dextran-coated SPION is a promising tool to noninvasively track hepatic macrophage localization for therapeutic monitoring.

137 Superparamagnetic Iron Oxide Nanoparticle Targeting of Adipose Tissue-Derived Stem Cells in Diabetes-Associated Erectile Dysfunction

To explore the effect of Adipose tissue-derived stem cells (ADSCs) labelled with superparamagnetic iron oxide nanoparticles (SPIONs) on improving the erectile function of streptozocin (STZ)-induced diabetic rats with an external magnetic field. Ten-week-old male Sprague Dawley rats were injected either with STZ to induce diabetes or citrate buffer as controls. Rat ADSCs were harvested and labelled with SPIONs, and then transplanted into corpus cavernosum of STZ-induced diabetic rats with or without an external magnetic field. At 28 days after transplantation, all rats were sacrificed for tracking of transplanted cells. Erectile response was assessed by cavernous nerve stimulation. Smooth muscle, endothelium and vascular endothelial growth factor (VEGF) expression in corpora cavernosum were assessed using immunohistochemistry. In vitro studies revealed that stem cell properties were minimally affected by SPION labeling. With magnetic targeting, SPION-labelled ADSCs were visibly attracted toward the magnet field. In vivo experiments, magnetic targeting of ADSCs contributed to long-term cell retention in corpus cavernosum and improved the erectile function of diabetic rats compared with ADSC injection alone. ADSCs were identified in the corpus cavernosums of diabetic rats at 28 days after transplantation, and they were negative for vWF and a-SMA, indicating that ADSCs did not differentiate into endothelial or smooth muscle cells. VEGF expression in the corpus cavernosum was significantly increased in both ADSC-treated groups compared with the PBS-treated group. The underlying mechanism of recovery appears to the paracrine mechanism of transplanted ADSCs.

Dose Dependent Side Effect of Superparamagnetic Iron Oxide Nanoparticle Labeling on Cell Motility in Two Fetal Stem Cell Populations

Multipotent stem cells (SCs) could substitute damaged cells and also rescue degeneration through the secretion of trophic factors able to activate the endogenous SC compartment. Therefore, fetal SCs, characterized by high proliferation rate and devoid of ethical concern, appear promising candidate, particularly for the treatment of neurodegenerative diseases. Super Paramagnetic Iron Oxide nanoparticles (SPIOn), routinely used for pre-clinical cell imaging and already approved for clinical practice, allow tracking of transplanted SCs and characterization of their fate within the host tissue, when combined with Magnetic Resonance Imaging (MRI). In this work we investigated how SPIOn could influence cell migration after internalization in two fetal SC populations: human amniotic fluid and chorial villi SCs were labeled with SPIOn and their motility was evaluated. We found that SPIOn loading significantly reduced SC movements without increasing production of Reactive Oxygen Species (ROS). Moreover, motility impairment was directly proportional to the amount of loaded SPIOn while a chemoattractant-induced recovery was obtained by increasing serum levels. Interestingly, the migration rate of SPIOn labeled cells was also significantly influenced by a degenerative surrounding. In conclusion, this work highlights how SPIOn labeling affects SC motility in vitro in a dose-dependent manner, shedding the light on an important parameter for the creation of clinical protocols. Establishment of an optimal SPIOn dose that enables both a good visualization of grafted cells by MRI and the physiological migration rate is a main step in order to maximize the effects of SC therapy in both animal models of neurodegeneration and clinical studies.

Automated Tracking of Nanoparticle-labeled Melanoma Cells Improves the Predictive Power of a Brain Metastasis Model

Biologic and therapeutic advances in melanoma brain metastasis are hampered by the paucity of reproducible and predictive animal models. In this work, we developed a robust model of brain metastasis that empowers quantitative tracking of cellular dissemination and tumor progression. Human melanoma cells labeled with superparamagnetic iron oxide nanoparticles (SPION) were injected into the left cardiac ventricle of mice and visualized by MRI. We showed that SPION exposure did not affect viability, growth, or migration in multiple cell lines across several in vitro assays. Moreover, labeling did not impose changes in cell-cycle distribution or apoptosis. In vivo, several SPION-positive cell lines displayed similar cerebral imaging and histologic features. MRI-based automated quantification of labeled cells in the brain showed a sigmoid association between metastasis frequency and doses of inoculated cells. Validation of this fully automated quantification showed a strong correlation with manual signal registration (r(2) = 0.921, P < 0.001) and incidence of brain metastases (r(2) = 0.708, P < 0.001). Metastasis formation resembled the pattern seen in humans and was unaffected by SPION labeling (histology; tumor count, P = 0.686; survival, P = 0.547). In summary, we present here a highly reproducible animal model that can improve the predictive value of mechanistic and therapeutic studies of melanoma brain metastasis.

Functions of corneal endothelial cells do not change after uptake of superparamagnetic iron oxide nanoparticles

To avoid donor tissue shortages, ex vivo cultured human corneal endothelial cell (HCEC) transplantation is a promising therapeutic resource. Superparamagnetic iron oxide nanoparticle (SPION) cell labeling assists HCEC transplantation by attaching the posterior corneal stroma in ex vivo animal models. However, possible functional changes of the HCECs following SPION labeling remain to be determined. In this study, we used SPIONs to label cultured rabbit CECs (RCECs) in order to observe important cell functions and the levels of cell markers. The synthetic SPIONs exhibited superparamagnetism at room temperature, with saturation magnetization of 55.4 emu/g and negligible remanence or coercivity. The ζ-potential was -24.5 mV and the diameter was 101 ± 55 nm. Immunostaining demonstrated a normal density of zonula occluden-1 (ZO-1), nestin and Ki-67 at cellular junctions or in nuclei from RCECs following SPION labeling at 16 µg/ml. MTT cytotoxicity assay, homotypic adhesion assay, quantitative flow cytometric Ki-67 analysis and RCEC pump function measurement demonstrated no significant differences between the cells with or without SPION labeling (P<0.05, for all assays). Results of this study demonstrated successful labeled cultured RCECs with synthetic SPIONs. Labeled cells possessed several important characteristics required to maintain the transparency and refractive parameters of the cornea, including hexagonal cell morphology, higher cell adhesion ability and proliferative potential, cell pump function and the positive expression of several cell markers.

Magnetic labeling of pancreaticβ-cells modulates the glucose- and insulin-induced phosphorylation of ERK1/2 and AKT

This study was undertaken to investigate the effect of a magnetic resonance imaging (MRI) contrast agent, superparamagnetic iron oxide nanoparticle (SPIO), on signal transduction by glucose and insulin in pancreatic β-cells. INS-1 cells were labeled in culture medium containing clinically approved SPIO for 24 h. Labeled and unlabeled cells were stimulated with glucose (25 mM) or insulin (0.1-1 µM) for 12 h. The phosphorylation of extracellular signal-regulated kinase1/2 (ERK1/2) and protein kinase B (AKT) and intracellular insulin protein levels were assessed by Western blotting. After labeling with increasing amounts of SPIO, cytotoxicity was not observed, yet the intracellular iron concentration increased in a dose-dependent manner. SPIO labeling (200 µg Fe ml(-1)) induced a significant increase in ERK1/2 and AKT phosphorylation (labeled vs unlabeled, p < 0.05), but significantly reduced the glucose-stimulated phosphorylation of ERK1/2 and AKT and insulin-stimulated phosphorylation of AKT (labeled vs unlabeled, p < 0.05). The level of intracellular insulin protein was found to be lower in labeled cells than unlabeled cells (labeled vs unlabeled, p < 0.05). This study demonstrates that SPIO labeling alters some fundamental functional variables, at least in INS-1 cells, through modulation of the glucose- or insulin-induced activation of ERK1/2 and AKT, which leads to insulin biosynthesis.

Compatibility of superparamagnetic iron oxide nanoparticle labeling for 1H MRI cell tracking with 31P MRS for bioenergetic measurements

Labeling of cells with superparamagnetic iron oxide nanoparticles permits cell tracking by (1)H MRI while (31)P MRS allows non-invasive evaluation of cellular bioenergetics. We evaluated the compatibility of these two techniques by obtaining (31)P NMR spectra of iron-labeled and unlabeled immobilized C2C12 myoblast cells in vitro. Broadened but usable (31)P spectra were obtained and peak area ratios of resonances corresponding to intracellular metabolites showed no significant differences between labeled and unlabeled cell populations. We conclude that (31)P NMR spectra can be obtained from cells labeled with sufficient iron to permit visualization by (1)H imaging protocols and that these spectra have sufficient quality to be used to assess metabolic status. This result introduces the possibility of using localized (31)P MRS to evaluate the viability of iron-labeled therapeutic cells as well as surrounding host tissue in vivo.

Superparamagnetic Iron Oxide Nanoparticles Labeling of Bone Marrow Stromal (Mesenchymal) Cells Does Not Affect Their “Stemness”

Superparamagnetic iron oxide nanoparticles (SPION) are increasingly used to label human bone marrow stromal cells (BMSCs, also called “mesenchymal stem cells”) to monitor their fate by in vivo MRI, and by histology after Prussian blue (PB) staining. SPION-labeling appears to be safe as assessed by in vitro differentiation of BMSCs, however, we chose to resolve the question of the effect of labeling on maintaining the “stemness” of cells within the BMSC population in vivo. Assays performed include colony forming efficiency, CD146 expression, gene expression profiling, and the “gold standard” of evaluating bone and myelosupportive stroma formation in vivo in immuncompromised recipients. SPION-labeling did not alter these assays. Comparable abundant bone with adjoining host hematopoietic cells were seen in cohorts of mice that were implanted with SPION-labeled or unlabeled BMSCs. PB+ adipocytes were noted, demonstrating their donor origin, as well as PB+ pericytes, indicative of self-renewal of the stem cell in the BMSC population. This study confirms that SPION labeling does not alter the differentiation potential of the subset of stem cells within BMSCs.

Efficient and Rapid Labeling of Transplanted Cell Populations with Superparamagnetic Iron Oxide Nanoparticles Using Cell Surface Chemical Biotinylation for in Vivo Monitoring by MRI

Determination of the dynamics of specific cell populations in vivo is essential for the development of cell-based therapies. For cell tracking by magnetic resonance imaging (MRI), cells need to internalize, or be surface labeled with a MRI contrast agent, such as superparamagnetic iron oxide nanoparticles (SPIOs): SPIOs give rise to signal loss by gradient-echo and T(2)-weighted MRI techniques. In this study, cancer cells were chemically tagged with biotin and then magnetically labeled with anti-biotin SPIOs. No significant detrimental effects on cell viability or death were observed following cell biotinylation. SPIO-labeled cells exhibited signal loss compared to non-SPIO-labeled cells by MRI in vitro. Consistent with the in vitro MRI data, signal attenuation was observed in vivo from SPIO-labeled cells injected into the muscle of the hind legs, or implanted subcutaneously into the flanks of mice, correlating with iron detection by histochemical and X-ray fluorescence (XRF) methods. To further validate this approach, human mesenchymal stem cells (hMSCs) were also employed. Chemical biotinylation and SPIO labeling of hMSCs were confirmed by fluorescence microscopy and flow cytometry. The procedure did not affect proliferation and multipotentiality, or lead to increased cell death. The SPIO-labeled hMSCs were shown to exhibit MRI signal reduction in vitro and was detectable in an in vivo model. In this study, we demonstrate a rapid, robust, and generic methodology that may be a useful and practical adjuvant to existing methods of cell labeling for in vivo monitoring by MRI. Further, we have shown the first application of XRF to provide iron maps to validate MRI data in SPIO-labeled cell tracking studies.