Superparamagnetic Iron Oxide Concentration Research Articles

SPION based magnetic PLGA nanofibers for neural differentiation of mesenchymal stem cells

Recently, magnetic platforms have been widely investigated in diagnostic, therapeutic and research applications due to certain properties, such as cell and tissue tracking and imaging, thermal therapy and being dirigible. In this study, the incorporation of magnetic nanoparticles (MNPs) in nanofibers has been proposed to combine the advantages of both nanofibers and MNPs to induce neural differentiation of mesenchymal stem cells. Magnetic poly (lactic-co-glycolic acid) nanofibers (containing 0%, 5% and 10% SPION) were fabricated and utilized as the matrix for the differentiation of mesenchymal stem cells (MSCs). Morphological, magnetic and mechanical properties were analyzed using FESEM, VSM and tensile test, respectively. The expression of neural markers (TUJ-1, NSE, MAP-2) was assessed quantitative and qualitatively utilizing RT-PCR and immunocytochemistry. Results confirmed the incorporation of MNPs in nanofibrous scaffold, presenting a saturation magnetization of 9.73 emu g−1. Also, with increase in magnetic particle concentration (0%–10%), tensile strength increased from 4.08 to 5.85 MPa, whereas the percentage of elongation decreased. TUJ-1 expression was 3.8 and 1.8 fold for 10% and 5% magnetic scaffold (versus non-magnetic scaffold) respectively, and the expression of NSE was 6.3 and 1.2-fold for 10% and 5%, respectively. Consequently, it seems that incorporation of magnetic biomaterial can promote the neural differentiation of MSCs, during which the augmentation of super paramagnetic iron oxide concentration from 0% to 10% accelerates the neural differentiation process.

Decreasing iron susceptibility with temperature in quantitative susceptibility mapping: A phantom study

To clarify the temperature dependence of susceptibility estimated by quantitative susceptibility mapping (QSM) analysis, we investigated the relationship between temperature and susceptibility using a cylinder phantom with varying temperatures. Six solutions with various concentrations of superparamagnetic iron oxide (SPIO) nanoparticles were employed. These tubes were placed in a cylinder phantom and surrounded with water. The temperature of the circulated water was adjusted to change the temperature in the cylinder phantom from 25.8 °C to 42.5 °C. The cylinder phantom was scanned via a three-dimensional multiple spoiled gradient-echo sequence for R2* and QSM analyses with varying temperatures. The relationships between temperature, susceptibility, and R2* values were determined. Moreover, the temperature coefficients of susceptibility (χ-Tc) and (R2*-Tc) were calculated at each concentration and the linearities in these indices against each SPIO concentration were validated. Significant inverse correlations were found between temperature, susceptibility, and R2* values at each SPIO concentration due to the decrease in paramagnetic iron susceptibility that occurred with increasing temperature based on Curie's law. Moreover, although there were significant correlations between the susceptibility and R2* values at any temperature, the slopes of the regression lines grew in height with greater temperatures. The percentage of difference per Celsius degree in susceptibility in any SPIO concentration was lower than the corresponding finding among the R2* results. There were strong linearities between the SPIO concentration, χ-Tc (r = −0.994; p < 0.001), and R2*-Tc (r = −0.998; p < 0.001). The χ-Tc and R2*-Tc outcomes in a particular voxel varied considerably with the iron contents. Although there was an inverse correlation noted between temperature and susceptibility, the susceptibility analysis showed smaller temperature dependence relative to the R2* analysis. QSM analysis might be a more suitable option for magnetic resonance-based iron quantification in comparison with R2* relaxometry.

DeepSPIO: Super Paramagnetic Iron Oxide Particle Quantification Using Deep Learning in Magnetic Resonance Imaging.

The susceptibility of super paramagnetic iron oxide (SPIO) particles makes them a useful contrast agent for different purposes in MRI. These particles are typically quantified with relaxometry or by measuring the inhomogeneities they produced. These methods rely on the phase, which is unreliable for high concentrations. We present in this study a novel Deep Learning method to quantify the SPIO concentration distribution. We acquired the data with a new sequence called View Line in which the field map information is encoded in the geometry of the image. The novelty of our network is that it uses residual blocks as the bottleneck and multiple decoders to improve the gradient flow in the network. Each decoder predicts a different part of the wavelet decomposition of the concentration map. This decomposition improves the estimation of the concentration, and also it accelerates the convergence of the model. We tested our SPIO concentration reconstruction technique with simulated images and data from actual scans from phantoms. The simulations were done using images from the IXI dataset, and the phantoms consisted of plastic cylinders containing agar with SPIO particles at different concentrations. In both experiments, the model was able to quantify the distribution accurately.

Endothelial progenitor cells in the peripheral blood of patients with moyamoya disease labeled with superparamagnetic iron oxide in vitro for MRI detection.

Moyamoya disease (MMD) is currently thought to involve endothelial progenitor cells (EPCs). We investigated whether superparamagnetic iron oxide (SPIO) can be used to label EPCs. Mononuclear cells from 10 moyamoya disease patients were isolated, and cluster of differentiation 133 (CD133) positive cells sorted by magnetic-activated cell sorting were cultured in vitro. The positive rates of CD133, vascular endothelial growth factor receptor (VEGFR)-2, and cluster of differentiation 34 (CD34) were detected by flow cytometry. The cells were co-cultured with fluorescence labeled Dil-acetylated-low-density lipoprotein (Dil-ac-LDL) and Ulex europaeus agglutinin-1 (UEA-1) to observe the endocytosis of Dil-ac-LDL and binding to UEA-1. Prussian blue staining and transmission electron microscopy were used to observe the endocytosis of different SPIO concentrations in EPCs, and CCK-8 was used to detect proliferation of cells transfected with different concentrations of SPIO. T2 weighted imaging (T2WI) signals from magnetic resonance imaging after SPIO endocytosis were compared. Positive rates of CD133, VEGFR-2, and CD34 on sorted mononuclear cells were 68.2±3.8, 57.5±4.2, and 36.8±6.5%, respectively. The double-positive expression rate of CD34 and VEGFR-2 was 19.6±4.7%, and 83.1±10.4% of cells, which showed the uptake of Dil-ac-LDL and binding with UEA-1. The labeling efficiencies of SPIO at concentrations of 25 and 50 μg/mL were higher than for 12.5 μg/mL. The proliferation of cells was not influenced by SPIO concentrations of 12.5 and 25 μg/mL. After labeling, the T2WI of EPCs was reduced. The concentration of 25 μg/mL SPIO had high labeling efficiency detected by magnetic resonance imaging (MRI) without decreased EPCs viability.

IMMU-25. MAGNETIC PARTICLE IMAGING FOR NON-INVASIVE TRACKING OF ADOPTIVE CELL TRANSFER IN CANCER IMMUNOTHERAPY

Abstract BACKGROUND Adoptive cell therapies (ACT) are strategies being explored to boost the immune response against cancer. ACT cancer immunotherapies are effective against metastatic melanoma, leukemia, and lymphoma, but face challenges in treating other solid tumors, such as in the brain. A critical step for the success of ACT in solid cancers is achieving trafficking and persistence of T-cells at tumor sites. Glioblastoma (GBM) is the most common and aggressive cancer of the central nervous system in adults, with a prognosis of 15-18-month average patient survival after diagnosis. Biomedical imaging is often used to track cell therapies. Magnetic Particle Imaging (MPI) is a novel biomedical imaging modality enabling non-invasive visualization of the distribution of biocompatible superparamagnetic iron oxide (SPIO) tracers. OBJECTIVE Label T-cells with SPIO to non-invasively track adoptive T cell transfer immunotherapy with MPI in the context of brain cancer. METHODS Murine pmel-DsRed T-cells were isolated from the spleen of a transgenic C57BL/6 mouse, and were exposed to different SPIO concentrations ex vivo. Cell viability, phenotype, and cytotoxic function were analyzed to determine if T-cells were affected by the SPIO labeling. Moreover, in vivo experiments were performed in a murine GBM model, and labeled T-cells were injected intravenously and tracked using MPI. RESULTS The SPIO-labeling of T-cells did not affected cell viability, phenotype, or cell cytotoxic function at all tested incubation conditions. The internalized SPIO can be quantified and spatially detected using MPI both in vitro and in vivo. In addition, MPI in vivo tracking shows T-cells accumulation in liver and lungs, as well in the spleen and brain, as showed ex vivo. CONCLUSIONS SPIO-labeling of T-cells did not affected its cytotoxic function and MPI allows for in vivo tracking of adoptively T-cell transfer. MPI will provide better understanding of ACT dynamics to accelerate development of novel treatments.

Silent susceptibility-weighted angiography to detect hemorrhagic lesions in the brain: a clinical and phantom study

To compare the effectiveness of silent susceptibility-weighted angiography (sSWAN), a new imaging technique with lower acoustic noise, with conventional susceptibility-weighted angiography (cSWAN) in the detection of intracranial hemorrhagic lesions. We measured the acoustic and background noise during sSWAN and cSWAN imaging and calculated the contrast-to-noise ratio (CNR) of the phantom consisting of eight chambers with different concentrations of superparamagnetic iron oxide. In the clinical study, we calculated the CNRs of hemorrhagic lesions in 15 patients and evaluated the images for conspicuity and artifact on each sequence and scored them on a 4-point scale. We also evaluated whether hypointense areas observed on sSWAN or cSWAN increased in size from those on T2*-weighted imaging (T2*-WI). Acoustic noise for sSWAN (57.9 ± 0.32 dB [background noise 51.3 dB]) was significantly less than that for cSWAN (89.0 ± 0.22 dB [background noise 50.9 dB]). The CNRs of phantoms for sSWAN were slightly but not significantly lower than those for cSWAN (P = 0.18). The CNRs of hemorrhagic lesions did not show significant differences between sSWAN and cSWAN (P = 0.17). There were no significant differences between sSWAN and cSWAN with respect to the scores for conspicuity, artifact, and change in size of hypointense areas from T2*-WI. sSWAN is equivalent to cSWAN with respect to the image quality for the detection of hemorrhagic lesions but has lower acoustic noise.

Synthesis and its targeting effect in vitro of disulfide-bonded hyaluronic acid-functionalized sodium-meter probe for hepatocellular carcinoma

Objective To explore the construction method and physicochemical properties of disulfide-bonded hyaluronic acid-functionalized sodium-meter probe for hepatocellular carcinoma,and its biological evaluation in vitro and feasibility of MRI. Methods Synthesis of hyaluronic acid-disulfide-bonded-poly e-caprolactone (HA-SS-PCL) by disulfide-bonded alkynyl-terminated polycaprolacton (alkyne-SS-PCL) and azido-terminated hyaluronic acid (HA-N3) by clicking chemical reaction,then doxorubicin (DOX) and superparamagnetic iron oxide (SPIO) were encapsulated in HA-SS-PCL core by dialysis method.HA-SS-PCL@DOX/SPIO was prepared and its particle size,DOX and SPIO loading rate were measured. With PBS as control group,the safety of HA-SS-PCL on human hepatocellular carcinoma cells HepG2 and normal liver cells LO2 was evaluated by the methylthiazolyl tetrazolium (MTT) assay, and the cytotoxicity of HA-PCL@DOX/SPIO and HA-SS-PCL@DOX/SPIO on human hepatocellular carcinoma cells HepG2 was evaluated. Immunofluorescence and flow cytometry were used to observe the expression of CD44 receptor on the surface of HepG2 cells in HA-PCL@DOX/SPIO and HA-SS-PCL@DOX/SPIO groups.Through in vitro MRI, PBS was used as the control group to observe the changes of T2 signal intensity of HA-PCL@DOX/SPIO and HA-SS-PCL@DOX/SPIO groups when SPIO concentration was 10,20,40,80 μg/ml. One way ANOVA test and t test were used. Results HA-SS-PCL@DOX/SPIO sodium-meter probes were successfully constructed. The particle size of HA-SS-PCL@DOX/SPIO was (126.9±6.3) nm,and they were spherical with uniform size. The loading rates of DOX and SPIO were 61.4% and 58.7%. MTT assay showed that the survival rate of HepG2 and LO2 cells was more than 80% even at 500 μg/ml of HA-SS-PCL,66.6% in HA-PCL@DOX/SPIO group and 55.2% in HA-SS-PCL@DOX/SPIO group. Immunofluorescence and flow cytometry showed that HA-PCL@DOX/SPIO and HA-SS-PCL@DOX/SPIO groups all have strong fluorescence,and the latter has stronger fluorescence intensity the former fluorescence intensity was 139.70±8.52,less than the latter 245.06±13.21. In vitro MRI showed that the T2 signal intensity of HA-PCL@DOX/SPIO and HA-SS-PCL@DOX/SPIO was significantly lower than that of the control group (F values were 613.591 and 569.234,P=0.000), the latter decline rate was more significant. Conclusion The disulfide-bonded hyaluronic acid-functionalized sodium-meter probe for hepatocellular carcinoma has excellent physicochemical properties,good targeting and MRI functions on human hepatoma cell HepG2 at the cellular level in vitro. Key words: Magnetic resonance imaging; Disulfide; Hyaluronicacid; Sodium-meter probe

Quantitative magnetic resonance imaging of brain iron deposition: comparison between quantitative susceptibility mapping and transverse relaxation rate (R2*) mapping

To evaluate the accuracy and sensitivity of quantitative susceptibility mapping (QSM) and transverse relaxation rate (R2*) mapping in the measurement of brain iron deposition. Super paramagnetic iron oxide (SPIO) phantoms and mouse models of Parkinson's disease (PD) related to iron deposition in the substantia nigra (SN) underwent 7.0 T magnetic resonance (MR) scans (Bruker, 70/16) with a multi-echo 3D gradient echo sequence, and the acquired data were processed to obtain QSM and R2*. Linear regression analysis was performed for susceptibility and R2* in the SPIO phantoms containing 5 SPIO concentrations (30, 15, 7.5, 3.75 and 1.875 µg/mL) to evaluate the accuracy of QSM and R2* in quantitative iron analysis. The sensitivities of QSM and R2* mapping in quantitative detection of brain iron deposition were assessed using mouse models of PD induced by 1-methyl-4-phenyl-1,2,3,6-tetrahy-dropyridine (MPTP) in comparison with the control mice. In SPIO phantoms, QSM provided a higher accuracy than R2* mapping and their goodness-of-fit coefficients (R2) were 0.98 and 0.89, respectively. In the mouse models of PD and control mice, the susceptibility of the SN was significantly higher in the PD models (5.19∓1.58 vs 2.98∓0.88, n=5; P<0.05), while the R2* values were similar between the two groups (20.22∓0.94 vs 19.74∓1.75; P=0.60). QSM allows more accurate and sensitive detection of brain iron deposition than R2*, and the susceptibility derived by QSM can be a potentially useful biomarker for studying PD.

In-vitro Labelling of Ovine Adipose-Derived Mesenchymal Stem Cells (oADMSCs) and Tracking Using MRI Technique

Abstract To understand the mechanisms standing behind a successful stem cell-based therapy, the monitoring of transplanted cell’s migration, homing as well as the engraftment efficiency and functional capability in-vivo has become a critical issue. The present study was designed to track the labelled oADMSCs in-vitro and its visualization through MRI technique. oADMSCs from passage 4 (P-4) to passage 6 (P-6) were labelled with superparamagnetic iron oxide (SPIO) conjugated with rhodamine (Molday Ion Rhodamine-B - MIRB) at the concentration of 25μg Fe/ml in DMEM. Internalized MIRB was observed under fluorescent microscope after 72 hrs of incubation. Labelled oADMSCs showed Prussian Blue positive reaction demonstrating the iron uptake of the cells. The viability of the MIRB-labelled oADMSCs ranged between 98-99 per cent and Trypan blue exclusion test showed no significant difference in viability between labelled and unlabelled oADMSCs. MR signal in control group of cells was similar to that of water. MR signals or fluorescence in MIRB-labelled cells decreased with increasing concentrations of iron. The T2 weighted images of MIRB-labelled oADMSCs increased with increasing concentrations of SPIOs. The MIRB was found to be nontoxic, and did not affect proliferation capacity in-vitro.

Research on biological activities and MRI of differentiations of neural-like cells induced by superparamagnetic iron oxide and green fluorescent protein double-labeled bone marrow-derived mesenchymal stem cells in vitro

Objective To explore the biological activities and the MR imaging signal intensities(SIs) characteristics of differentiations of neural-like cells induced by superparamagnetic iron oxide (SPIO) and green fluorescent protein (GFP) double-labeled bone marrow-derived mesenchymal stem cells (BMSCs) in vitro. Methods GFP-BMSCs were labeled with different concentrations of SPIO in vitro (the concentration of the A, B, C and D group was 25, 50, 75 and 100 ug/ml, respectively; the E group without labels of SPIO served as the control group). The Prussian blue stainings were used to detect the labeling rates of SPIO. The iron contents of cells were measured by atomic absorption spectrometer. The CCK8 experiments were used to detect the cell proliferation rates. The cell cycles were detect by PCR. Each of the A- E groups had a test tube with 1 × 108 cells.All test tubes underwent T2* weighted imaging (T2*WI)and susceptibility weighted imaging (SWI) in a MR imaging scanner. The optimal group was defined by comparing the measurements of SIs between T2*WI and SWI. The optimal group and the E group together induced the differentiations of osteogenesis and neural-like cells. The stainings of alizarin red, osteocalcin and Nissl, NeuN, and NF-200 were performed at 72 hours. Real-time quantitative PCR was used to detect the expression levels of RNA in tub3, nestin, NSE, MAP-2 and Syt1. The positive staining rates and the expression levels of RNA were compared between the two groups. Finally, SWI was used to analyse the changes of SIs. One-way analysis of variance (ANOVA) was used to the multi-group comparison. Using least significant difference (LSD) test to analyse the comparisons between the multi-groups. Results The labeling rates of the A- D groups were 100%. The iron contents of cells in the A- E groups were (14.36 ± 7.61), (21.73±3.42), (30.54±8.73), (33.65±9.62), and (2.31±0.32) pg/cell, respectively. The iron contents of cells in the A-D groups were significantly higher than those in the E group (F=3.852,P=0.003). There was no significant difference between the C and D groups (P=0.267). In all groups, the D group had the lowest OD value in the CCK-8 experiments (3.18 ± 0.46). In the A- E groups, the changes of SIs in SWI were significantly lower than those in T2* WI.There was no significant difference in SIs of SWI between the C group (145.89±14.31) and the D group (127.37±12.21). Except the comparison between the group C and D, the comparisons between all the groups had significant differences (P 0.05). The expression levels of tub3, nestin and NSE were significantly higher before than after induced differentiations(P<0.01). SIs of SWI had no significant difference between before and after induced differentiations in the C group (t=1.26,P=0.236). Conclusions SPIO and GFP double-labeled BMSCs can induce neural-like cells without influencing biologic activities. MR SIs are decreased by the increase of SPIO concentrations in cells. SWI was the most sensitive sequence. The SIs of SWI has no differnce between before and after induced differentiations. Key words: Mesenchymal stem cell; Magnetic resonance imaging; Molecular probe

Tracking mesenchymal stem cells using magnetic resonance imaging.

Recent translational studies in the fields of tissue regeneration and cell therapy have characterized mesenchymal stem cells (MSCs) as a potentially effective and accessible measure for treating ischemic cerebral and neurodegenerative disorders such as stroke, Parkinson's disease, and amyotrophic lateral sclerosis. Developing more efficient cell tracking techniques bear the potential to optimize MSC transplantation therapies by providing a more accurate picture of the fate and area of effect of implanted cells. Currently, determining the location of transplanted MSCs involves a histological approach, but magnetic resonance imaging (MRI) presents a noninvasive paradigm that permits repeat evaluations. To visualize MSCs using MRI, the implanted cells must be treated with an intracellular contrast agent. These are commonly paramagnetic compounds, many of which are based on superparamagnetic iron oxide (SPIO) nanoparticles. Recent research has set out characterize the effects of SPIO-uptake on the cellular activity of in vitro human MSCs and the resultant influence that respective SPIO concentration has on MRI sensitivity. As these studies reveal, SPIO-uptake has no effect on the cellular processes of proliferation and differentiation while producing high contrast MRI signals. Moreover, transplantation of SPIO-labeled MSCs in animal models encouragingly showed no loss in MRI contrast, suggesting that SPIO labeling may be an appealing regime for lasting MRI detection. This study is a review article. Referred literature in this study has been listed in the reference part. The datasets supporting the conclusions of this article are available online by searching the PubMed. Some original points in this article come from the laboratory practice in our research centers and the authors’ experiences.

A DNA hybridization system for labeling of neural stem cells with SPIO nanoparticles for MRI monitoring post-transplantation

Neural stem cells (NSCs) demonstrate encouraging results in cell replacement therapy for neurodegenerative disorders and traumatic injury in the central nervous system. Monitor the survival and migration of transplanted cells would provide us important information concerning the performance and integration of the graft during the therapy time course. Magnetic resonance imaging (MRI) allow us to monitor the transplanted cells in a non-invasive way. The only requirement is to use an appropriate contrast agent to label the transplanted cells. Superparamagnetic iron oxide (SPIO) nanoparticles are one of the most commonly used contrast agent for MRI detection of transplanted cells. SPIO nanoparticles demonstrated to be suitable for labeling several types of cells including NSCs. However, the current methods for SPIO labeling are non-specific, depending mostly on electrostatic interactions, demanding relatively high SPIO concentration, and long incubation time, which can affect the viability of cells. In this study, we propose a specific and relatively fast method to label NSCs with SPIO nanoparticles via DNA hybridization. Two short single stranded DNAs (ssDNAs), oligo[dT]20 and oligo[dA]20 were conjugated with a lipid molecule and SPIO nanoparticle respectively. The labeling process comprises two simple steps; first the cells are modified to present oligo[dT]20 ssDNA on the cell surface, then the oligo[dA]20 ssDNA conjugated with SPIO nanoparticles are presented to the modified cells to allow the oligo[dT]20-oligo[dA]20 hybridization. The method showed to be non-toxic at concentrations up to 50 μg/mL oligo[dA]20-SPIO nanoparticles. Presence of SPIO nanoparticles at cell surface and cell cytoplasm was verified by transmission electron microscopy (TEM). SPIO labeling via DNA hybridization demonstrated to not interfere on NSCs proliferation, aggregates formation, and differentiation. NSCs labeled with SPIO nanoparticles via DNA hybridization system were successfully detected by MRI in vitro as well in vivo. Cells transplanted into the rat brain striatum could be detected by MRI scanning up to 1 month post-transplantation.

Experimental study of endothelial progenitor cells labeled with superparamagnetic iron oxide in vitro.

Endothelial progenitor cells (EPCs) have an essential role in counteracting risk factor‑induced endothelial injury and protecting against the development of vascular injury, such as myocardial infarction. Magnetic resonance imaging (MRI) was reported to be effective in tracking transplanted stem cells following cell‑labeling with superparamagnetic iron oxide (SPIO) nanoparticles. SPIO has previously been used to label and track EPCs; however, the safest concentration of SPIO for labeling EPCs on a cellular level has remained to be elucidated. In addition, the optimum number of SPIO‑labeled cells required to produce the highest quality magnetic resonance images has not yet been determined. In the present study, EPCs were isolated from the bone marrow of minipigs using density gradient centrifugation. Their biological activity was then studied using flow cytometric analysis. Cells were incubated at different concentrations of SPIO for different durations and then the growth curve, apoptosis, morphology and labeling efficiency of the EPCs were detected using optical and electron microscopy. T2‑weighted fast spin‑echo (T2WITSE) MRI of the different numbers of SPIO‑labeled EPCs (35 µg/ml) were then obtained in axial and sagittal planes. The results of the present study demonstrated that EPCs were efficiently labeled with SPIO, with a labeling efficiency in each group of ~100% following incubation for 24 h. SPIO was found to be localized in the endosomal vesicles of EPCs, which was confirmed by electron microscopy. When the concentration of SPIO was <70 µg/ml, no significant differences were observed in cell viability, proliferative capability (P>0.05) and morphology between labeled and unlabeled EPCs. Furthermore, the T2WITSE signal intensity was significantly decreased in the groups of 5.0x105/ml and 1.0x105/ml compared with that of the control (P<0.05). In conclusion, the results of the present study indicated that 35 µg/ml was the most effective concentration of SPIO to label EPCs in vitro and acquire a high quality MRI. These findings may therefore contribute to the development of a promising novel therapeutic method for the treatment of myocardial infarction following autograft with SPIO‑labeled EPCs in vivo.

In vivo tracking and immunological properties of pulsed porcine monocyte-derived dendritic cells

Cellular therapies using immune cells and in particular dendritic cells (DCs) are being increasingly applied in clinical trials and vaccines. Their success partially depends on accurate delivery of cells to target organs or migration to lymph nodes. Delivery and subsequent migration of cells to regional lymph nodes is essential for effective stimulation of the immune system. Thus, the design of an optimal DC therapy would be improved by optimizing technologies for monitoring DC trafficking. Magnetic resonance imaging (MRI) represents a powerful tool for non-invasive imaging of DC migration in vivo. Domestic pigs share similarities with humans and represent an excellent animal model for immunological studies. The aim of this study was to investigate the possibility using pigs as models for DC tracking in vivo.Porcine monocyte derived DC (MoDC) culture with superparamagnetic iron oxide (SPIO) particles was standardized on the basis of SPIO concentration and culture viability. Phenotype, cytokine production and mixed lymphocyte reaction assay confirmed that porcine SPIO-MoDC culture were similar to mock MoDCs and fully functional in vivo. Alike, similar patterns were obtained in human MoDCs. After subcutaneous inoculation in pigs, porcine SPIO-MoDC migration to regional lymph nodes was detected by MRI and confirmed by Perls staining of draining lymph nodes. Moreover, after one dose of virus-like particles-pulsed MoDCs specific local and systemic responses were confirmed using ELISPOT IFN-γ in pigs. In summary, the results in this work showed that after one single subcutaneous dose of pulsed MoDCs, pigs were able to elicit specific local and systemic immune responses. Additionally, the dynamic imaging of MRI-based DC tracking was shown using SPIO particles. This proof-of-principle study shows the potential of using pigs as a suitable animal model to test DC trafficking with the aim of improving cellular therapies.

Superparamagnetic iron oxide nanoparticles function as a long-term, multi-modal imaging label for non-invasive tracking of implanted progenitor cells.

The purpose of this study was to determine the ability of superparamagnetic iron oxide (SPIO) nanoparticles to function as a long-term tracking label for multi-modal imaging of implanted engineered tissues containing muscle-derived progenitor cells using magnetic resonance imaging (MRI) and X-ray micro-computed tomography (μCT). SPIO-labeled primary myoblasts were embedded in fibrin sealant and imaged to obtain intensity data by MRI or radio-opacity information by μCT. Each imaging modality displayed a detection gradient that matched increasing SPIO concentrations. Labeled cells were then incorporated in fibrin sealant, injected into the atrioventricular groove of rat hearts, and imaged in vivo and ex vivo for up to 1 year. Transplanted cells were identified in intact animals and isolated hearts using both imaging modalities. MRI was better able to detect minuscule amounts of SPIO nanoparticles, while μCT more precisely identified the location of heavily-labeled cells. Histological analyses confirmed that iron oxide particles were confined to viable, skeletal muscle-derived cells in the implant at the expected location based on MRI and μCT. These analyses showed no evidence of phagocytosis of labeled cells by macrophages or release of nanoparticles from transplanted cells. In conclusion, we established that SPIO nanoparticles function as a sensitive and specific long-term label for MRI and μCT, respectively. Our findings will enable investigators interested in regenerative therapies to non-invasively and serially acquire complementary, high-resolution images of transplanted cells for one year using a single label.

In vivo quantification of SPIO nanoparticles for cell labeling based on MR phase gradient images.

Along with the development of modern imaging technologies, contrast agents play increasingly important roles in both clinical applications and scientific research. Super-paramagnetic iron oxide (SPIO) nanoparticles, a negative contrast agent, have been extensively used in magnetic resonance imaging (MRI), such as in vivo labeling and tracking of cells. However, there still remain many challenges, such as in vivo quantification of SPIO nanoparticles. In this work, an MR phase gradient-based method was proposed to quantify the SPIO nanoparticles. As a calibration, a phantom experiment using known concentrations (10, 25, 50, 100, 150 and 250 µg/ml) of SPIO was first conducted to verify the proposed quantification method. In a following in vivo experiment, C6 glioma cells labeled with SPIO nanoparticles were implanted into flanks of four mice, which were scanned 1-3 days post-injection for in vivo quantification of SPIO concentration. The results showed that the concentration of SPIO nanoparticles could be determined in both phantom and in vivo experiments using the developed MR phase gradients approach.

Diagnosis of intracranial hemorrhagic lesions: comparison between 3D-SWAN (3D T2*-weighted imaging with multi-echo acquisition) and 2D-T2*-weighted imaging

3D-susceptibility-weighted angiography (SWAN) can produce high-resolution images that yield excellent susceptibility-weighted contrast at a relatively short acquisition time. To compare SWAN- and 2D-T2*-weighted gradient-echo images (T2*-WI) for their sensitivity in the depiction of cerebral hemorrhagic lesions. We subjected 75 patients with suspected cerebral hemorrhagic lesions to SWAN and T2*-WI at 3T. We first measured the contrast-to-noise ratio (CNR) using an agar phantom that contained different concentrations of superparamagnetic iron oxide (SPIO). The acquisition time for SWAN and T2*-WI was similar (182 vs. 196 s). Neuroradiologists compared the two imaging methods for lesion detectability and conspicuity. The CNR of the phantom was higher on SWAN images. Of the 75 patients, 50 were found to have a total of 278 cerebral hemorrhagic lesions (microbleeds, n = 229 [82.4%]; intracerebral hemorrhage, n = 18 [6.5%]; superficial siderosis, n = 13 [4.7%]; axonal injuries, n = 8 [2.9%]; subarachnoid hemorrhage [SAH] or brain contusion, n = 3 each [1.0%]; subdural hematoma, n = 2 [0.7%]; cavernous hemangioma or dural arterteriovenous fistula, n = 1 each [0.4%]). In none of the lesions was the SWAN sequence inferior to T2*-WI with respect to lesion detectability and conspicuity. In fact, SWAN yielded better lesion conspicuity in patients with superficial siderosis and SAH: it detected significantly more lesions than T2*-WI (P < 0.01) and it was particularly useful for the detection of microbleeds and lesions near the skull base. SWAN is equal or superior to standard T2*-WI for the diagnosis of various cerebral hemorrhagic lesions. Because its acquisition time is reasonable it may replace T2*-WI.

Monitoring Steady Flow Effects on Cell Distribution in Engineered Valve Tissues by Magnetic Resonance Imaging

In heart valve tissue engineering, assessment of cell migration under dynamic states can provide insights on the evolving tissue structure. We labeled human vascular smooth muscle (SMCs), endothelial (ECs), and bone marrow-derived mesenchymal stem cells (BMSCs) with superparamagnetic iron oxide (SPIO) microparticles and visualized them using magnetic resonance imaging (MRI) under steady flow. We determined that vascular cells were able to remain reasonably viable and proliferate well after being labeled with SPIO microparticles (200 μg/mL) for 48 hours. SPIO-labeled cells were successfully visualized using T2* contrast. When physiologically representative shear stresses (5-6 dynes/cm2) were applied to SMC-EC coculture-seeded scaffolds, hypointense regions seemed to have decreased after 2 weeks in some locations, whereas others revealed sustained levels of T2* contrast; similar observations were seen in the case of BMSC-seeded scaffolds. This could be attributable to increased out-of-plane cell migratory activity, which occurred from the fluid-induced mechanical cues received, which was not previously evidenced in static culture. Vascular cells and BMSCs were labeled with remarkably high concentrations of SPIO. Moreover, steady fluid flow enhanced intrascaffold cell migration of vascular SMCs and ECs as well as BMSCs, which, in turn, significantly improved construct cellularity and extracellular collagen content.

Magnetic Resonance Imaging of Human Dental Pulp Stem Cells in Vitro and in Vivo

Recent advances in stem cell research have shown the promising nature of mesenchymal stem cells as plausible candidates for cell-based regenerative medicine. Many studies reported the use of human dental pulp stem cells (hDPSCs), which possess self-renewal capacity, high proliferation potential, and the ability to undergo multilineage differentiation. Together with this therapeutic approach, development of effective, noninvasive and nontoxic imaging techniques for visualizing and tracking the cells in vivo is crucial for the evaluation and improvement of stem cell therapy. Magnetic resonance imaging (MRI) is one of the most powerful diagnostic imaging techniques currently available for in vivo diagnosis and has been proposed as the most attractive modality for monitoring stem cell migration. The aim of this study was to investigate the labeling efficiency of hDPSCs using superparamagnetic iron oxide (SPIO) particles in order to allow visualization using in vitro and in vivo MRI without influencing cellular metabolism. MRI and transmission electron microscopy (TEM) showed optimal uptake with low SPIO concentrations of 15 µg/ml in combination with 0.75 µg/ml poly-L-lysine (PLL) resulting in more than 13 pg iron/cell and an in vitro detection limit of 50 labeled cells/µl. Very low SPIO concentrations in the culture medium resulted in extremely high labeling efficiency not reported before. For these conditions, tetrazolium salt assays showed no adverse effects on cell viability. Furthermore, in vivo MRI was performed to detect labeled hDPSCs transplanted into the brain of Rag 2-γ C immune-deficient mice. Transplanted cells did not show any signs of tumorgenecity or teratoma formation during the studied time course. We have reported on a labeling and imaging strategy to visualize human dental pulp stem cells in vivo using MRI. These data provide a solid base to allow cell tracking in future regenerative studies in the brain longitudinally.

Tracing type 1 diabeticTibet miniature pig's bone marrow mesenchymal stem cells in vitro by magnetic resonance imaging (1型糖尿病西藏小型猪骨髓间充质干细胞体外磁共振成像示踪)

Traditional cell-tracking methods fail to meet the needs of preclinical or clinical research. Thus, the aim of the present study was to establish a new method of double labeling bone marrow mesenchymal stem cells (BMSCs) from type 1 diabetic (T1D) minipigs with super-paramagnetic iron oxide (SPIO) and enhanced green fluorescent protein (eGFP) and tracing them using MRI in vitro. Isolated BMSCs from T1D minipigs were labeled with eGFP and different concentrations of SPIO. The effects of lentivirus (LV)-eGFP transfection and SPIO on the viability and growth curves of BMSCs were determined by Trypan blue exclusion, the 3-(4,5-dimethyl-2 thiazoyl)-2,5-diphenyl-2H-tetrazolium bromide assay and flow cytometry. Cellular ultrastructure was evaluated by transmission electron microscopy. Magnetic resonance imaging was used to evaluate BMSCs labeled with SPIO-eGFP complexes 6 weeks after labeling. Expression of eGFP in BMSCs peaked 96 h after transfection with LV-eGFP. Prussian blue staining revealed scattered blue granules in the cytoplasm of SPIO-labeled cells. Transmission electron microscopy revealed that the dense granules aggregated mainly in secondary lysosomes. On MRI, T2* -weighted imaging was far more sensitive for SPIO-labeled BMSCs than other image sequences 3 and 6 weeks after the cells had been labeled with SPIO-eGFP. We have developed a relatively simple and safe method for double labeling of BMSCs from T1D minipigs using SPIO and LV-eGFP and tracing them in vitro by MRI for 6 weeks.