Frank Eibofner

Functional investigations on human mesenchymal stem cells exposed to magnetic fields and labeled with clinically approved iron nanoparticles.

BackgroundFor 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 MSCs 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.ResultsHuman 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.ConclusionsIn 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.

Positive contrast imaging of iron oxide nanoparticles with susceptibility-weighted imaging

Superparamagnetic iron oxide particles can be utilized to label cells for immune cell and stem cell therapy. The labeled cells cause significant field distortions induced in their vicinity, which can be detected with magnetic resonance imaging (MRI). In conventional imaging, the signal voids arising from the field distortions lead to negative contrast, which is not desirable, as detection of the cells can be masked by native low signal tissue. In this work, a new method for visualizing magnetically labeled cells with positive contrast is proposed and described. The technique presented is based on the susceptibility‐weighted imaging (SWI) post‐processing algorithm. Phase images from gradient‐echo sequences are evaluated pixel by pixel, and a mask is created with values ranging from 0 to 1, depending on the phase value of the pixel. The magnitude image is then multiplied by the mask. With an appropriate mask function, positive contrast in the vicinity of the labeled cells is created. The feasibility of this technique is proved using an agar phantom containing superparamagnetic iron oxide particles–labeled cells and an ex vivo bovine liver. The results show high potential for detecting even small labeled cell concentrations in structurally inhomogeneous tissue types. Magn Reson Med, 2010.

Quantitative diffusion imaging of adipose tissue in the human lower leg at 1.5 T

Diffusion‐weighted spin‐echo echo‐planar imaging was developed and applied for assessment of diffusion coefficients of adipose tissue in human lower leg on a 1.5‐T whole‐body MR scanner. Because of the higher molecular weight of triglycerides, apparent diffusion coefficients (ADCs) of adipose tissue are approximately two orders of magnitude smaller compared with water, leading to the necessity of using high b‐values up to 50,000 sec/mm2 and an echo time of 240 msec for sufficient diffusion‐related signal attenuation. ADC maps of adipose tissue in the human lower leg were derived for diffusion encoding along orthogonal spatial directions in six healthy volunteers. Mean diffusion coefficients in the tibial bone marrow amounted to (1.81 ± 0.10) × 10−5 mm2/sec (left–right), (1.96 ± 0.10) × 10−5 mm2/sec (anterior–posterior), and (1.96 ± 0.20) × 10−5 mm2/sec (head–feet), respectively. Pixel‐wise calculated ADC values of subcutaneous adipose tissue showed a distinctly higher variation with the smallest ADC values similar to those measured for tibial bone marrow. Some subcutaneous adipose tissue regions showed increased signal attenuation at higher b‐values resulting in ADC coefficients up to 4.2 × 10−5 mm2/sec. It must be noted that diffusion measurements with extremely high b‐values in vivo are extremely sensitive to incoherent motion effects in tissue. Nonetheless, it could be shown that in vivo diffusion imaging of adipose tissue in human lower leg is possible at 1.5 T in acceptable measurement time of a few minutes. Potential future applications of fat diffusion imaging are seen in temperature measurements in adipose tissue, detection of free fatty acids in white or brown adipose tissue in case of high lipolysis, differentiation of macro‐ and microvesicular steatosis, or assessment of the mobility of intramyocellular lipids. Magn Reson Med, 2010.

Viability and MR detectability of iron labeled mesenchymal stem cells used for endoscopic injection into the porcine urethral sphincter

Direct stem cell therapies for functionally impaired tissue require a sufficient number of cells in the target region and a method for verifying the fate of the cells in the subsequent time course. In vivo MRI of iron labeled mesenchymal stem cells has been suggested to comply with these requirements. The study was conducted to evaluate proliferation, migration, differentiation and adhesion effects as well as the obtained iron load of an iron labeling strategy for mesenchymal stem cells. After injection into the porcine urethral sphincter, the labeled cells were monitored for up to six months using MRI.

Anatomic-functional (perfusion-based) magnetic resonance imaging follow-up in patients with rheumatoid arthritis treated with anti-interleukin 6 antibodies: a comparison with clinical scores and serologic data.

Objective On a 3-T magnetic resonance scanner, morphologic and perfusion changes of 9 patients with rheumatoid arthritis were evaluated after start of anti–interleukin 6 receptor antibody Tocilizumab (TCZ) treatment. Methods Morphologic and perfusion-based magnetic resonance imaging (MRI) parameters were assessed before and 4, 12, and 24 after the start of TCZ treatment. Furthermore, serologic biomarkers and clinical assessment scores were evaluated 4, 12, 24, and 52 weeks after treatment initiation. Results Results of MRI parameters showed significant group differences between responders and nonresponders for synovial volume, transfer constant, and blood plasma volume fraction already at week 12 as well as relative enhancement and rate of early enhancement at week 24. Conclusions Considering the temporal change of perfusion-derived MRI parameters (transfer constant, blood plasma volume fraction, relative enhancement, and rate of early enhancement) as well as morphologic MRI parameters (synovial volume measurements), a quantifiable assessment of response to TCZ therapy in rheumatoid arthritis seems possible at an even earlier time point compared with clinical assessment scores, whereas serologic biomarkers proved nonspecific in this respect.

Utilizing echo‐shifts in k‐space for generation of positive contrast in areas with marked susceptibility alterations

A technique for generation of positive contrast near susceptibility alterations utilizing echo‐shifts in k‐space is introduced, based on altered Larmor‐frequencies and resulting phase‐shifts accumulating during the echo‐time at the site of local magnetic field gradients. 3D gradient‐echo raw‐data is acquired and weighted with an inverse Hanning filter. The filter partly suppresses central raw‐data points, while maintaining outer areas. Reconstruction of the filtered raw‐data results in images where pixels with apparent magnetic field gradients are highlighted against homogeneous pixels. Further processing steps are introduced to remove remaining intensities in the homogeneous parts of the filtered image. Feasibility is shown by an agar phantom containing magnetically labeled cells, with concentrations of 25, 50, 100, and 250 cells/μL, and by images of the human head. The technique allows detection of echo‐shifted pixels with automatic suppression of magnetically homogeneous parts while keeping post‐processing time short. Fewer than four labeled cells per pixel were clearly displayed with positive contrast. Application to the human head shows bright veins and complete suppression of homogeneous regions. The presented technique has high potential for specific detection of low concentrations of labeled cells or susceptibility altered regions in vivo with positive contrast, whereas areas with low spin density are not highlighted. Magn Reson Med, 2012.

A fully automated trabecular bone structural analysis tool based on T2*-weighted magnetic resonance imaging

One major source affecting the precision of bone structure analysis in quantitative magnetic resonance imaging (qMRI) is inter- and intraoperator variability, inherent in delineating and tracing regions of interest along longitudinal studies. In this paper an automated analysis tool, featuring bone marrow segmentation, region of interest generation, and characterization of cancellous bone of articular joints is presented. In evaluation studies conducted at the knee joint the novel analysis tool significantly decreased the standard error of measurement and improved the sensitivity in detecting minor structural changes. It further eliminated the need of time-consuming user interaction, and thereby increasing reproducibility.

Technical Note: MR-visualization of interventional devices using transient field alterations and balanced steady-state free precession imaging.

PURPOSE In interventional magnetic resonance imaging, instruments can be equipped with conducting wires for visualization by current application. The potential of sequence triggered application of transient direct currents in balanced steady-state free precession (bSSFP) imaging is demonstrated. METHODS A conductor and a modified catheter were examined in water phantoms and in an ex vivo porcine liver. The current was switched by a trigger pulse in the bSSFP sequence in an interval between radiofrequency pulse and signal acquisition. Magnitude and phase images were recorded. Regions with transient field alterations were evaluated by a postprocessing algorithm. A phase mask was computed and overlaid with the magnitude image. RESULTS Transient field alterations caused continuous phase shifts, which were separated by the postprocessing algorithm from phase jumps due to persistent field alterations. The overlaid images revealed the position of the conductor. The modified catheter generated visible phase offset in all orientations toward the static magnetic field and could be unambiguously localized in the ex vivo porcine liver. CONCLUSIONS The application of a sequence triggered, direct current in combination with phase imaging allows conspicuous localization of interventional devices with a bSSFP sequence.

Quantitative Assessment of Iron-Labeled Stem-Cell Adhesion at the Vessel Wall in a Vascular Flow Model: Correlation of T2*-Weighted Imaging at 3 T and Histology.

PURPOSE To evaluate the distribution of superparamagnetic iron oxide (SPIO)-labeled cells in a perfused segment of a porcine artery and to estimate the number of adherent cells by means of magnetic resonance (MR) imaging. MATERIALS AND METHODS Six vessel specimens (diameters between 0.8 and 1.2 cm) were placed in a bioreactor system, and 2 × 10(4) to 1 × 10(6) SPIO-labeled endothelial colony-forming cells were injected into the artery within the perfused reactor. The area of resulting signal extinctions at the inner wall of the vessels was quantified on MR images by using a high-resolution T2*-weighted sequence with a slice-by-slice approach. After imaging, the labeled cells were quantified histologically. RESULTS The total iron load of each cell was 56.5 pg ± 14.4. In the applied range of 2 × 10(4) to 1 × 10(6) cells per vessel, the area of iron-induced signal extinction at the vessel wall on T2*-weighted imaging corresponded to the histologically detected cell number (r = 0.98, P < .001). CONCLUSIONS A correlation between the area of signal extinction and the number of labeled cells at the vessel wall was found. This might help to evaluate dose rates in further clinical applications of intravascular cell-based therapies.