Enhancement of gas‐filled microbubble R2* by iron oxide nanoparticles for MRI

Gas‐filled microbubbles have the potential to become a unique intravascular MR contrast agent due to their magnetic susceptibility effect, biocompatibility, and localized manipulation via ultrasound cavitation. However, microbubble susceptibility effect is relatively weak when compared with other intravascular MR susceptibility contrast agents. In this study, enhancement of microbubble susceptibility effect by entrapping monocrystalline iron oxide nanoparticles (MIONs) into polymeric microbubbles was investigated at 7 T in vitro. Apparent T₂ enhancement (ΔR₂*) induced by microbubbles was measured to be 79.2 ± 17.5 sec^?1 and 301.2 ± 16.8 sec^?1 for MION‐free and MION‐entrapped polymeric microbubbles at 5% volume fraction, respectively. ΔR₂* and apparent transverse relaxivities (r₂*) for MION‐entrapped polymeric microbubbles and MION‐entrapped solid microspheres (without gas core) were also compared, showing the synergistic effect of the gas core with MIONs. This is the first experimental demonstration of microbubble susceptibility enhancement for MRI application. This study indicates that gas‐filled polymeric microbubble susceptibility effect can be substantially increased by incorporating iron oxide nanoparticles into microbubble shells. With such an approach, microbubbles can potentially be visualized with higher sensitivity and lower concentrations by MRI. Magn Reson Med, 2010. © 2009 Wiley‐Liss, Inc.     

Uptake of dextran-coated monocrystalline iron oxides in tumor cells and macrophages

Although several dextran-coated iron oxide preparations are in preclinical and clinical use, little is known about the mechanism of uptake into cells. As these particles have been shown to accumulate in macrophages and tumor cells, we performed cellular uptake and inhibition studies with a prototypical monocrystalline iron oxide nanoparticle (MION). MION particles were labeled with fluorescein isothiocyanate or radioiodinated and purified by gel permeation chromatography. Two preparations of MION particles were used in cell experiments: nontreated MION and plasma-opsonized MION purified by gradient density purification. As determined by immunoblotting, opsonization resulted in C3, vitro-nectin, and fibronectin association with MION. Incubation of cells with fluorescent MION showed active uptake of particles in macrophages both before and after opsonization. In C6 tumor cells, however, intracellular MION was only detectable in dividing cells. Quantitatively, ¹²⁵I-labeled MION was internalized into cells with uptake values ranging from 17 ng (in 9L gliosarcoma) to 970 ng iron per million cells for peritoneal macrophages. Opsonization increased MION uptake into macrophages sixfold, whereas it increased the uptake in C6 tumor cells only twofold. Results from uptake inhibition assay suggest that cellular uptake of nonopsonized (dextran-coated) MION particles is mediated by fluid-phase endocytosis, whereas receptor-mediated endocytosis is presumably responsible for the uptake of opsonized (protein-coated) particles.     

Magnetically labeled cells can be detected by MR imaging

To determine the feasibility of MR imaging of magnetically labeled cells, different cell lines were labeled with monocrystalline iron oxide (MION) particles. Phantoms containing MION labeled cells were then assembled and imaged by MR at 1.5 T using T1-weighted and T2-weighted pulse sequences. MION uptake ranged from 8.5 × 10⁴ to 2.9 × 10⁵ particles/cell for tumor cells (9L and LX1, respectively) to 1.5 × 10⁶ to 4.8 × 10⁸ particles/cell for “professional phagocytes” (J774 and peritoneal macrophages, respectively). On the T1-weighted images, cell-internalized MION appeared hyperintense relative to agar and similar to MION in aqueous solution. On T2-weighted images, signal intensity varied according to concentration of MION within cells. Cell-internalized MION caused similar MR signal changes of cells as did free MION; however, at a dose that was an order of magnitude lower, depending on the pulse sequence used. The detectability of MION within cells was approximately 2 ng Fe, which corresponded to 10⁵ tumor cells/well or 5 × 10³ macrophages/well. We conclude that a variety of cells can be efficiently labeled with MION by simple incubation. Intracellular labeling may be used for MR imaging of in vivo cell tracking.     

Labeling efficacy of superparamagnetic iron oxide nanoparticles to human neural stem cells: comparison of ferumoxides, monocrystalline iron oxide, cross-linked iron oxide (CLIO)-NH2 and tat-CLIO.

OBJECTIVE: We wanted to compare the human neural stem cell (hNSC) labeling efficacy of different superparamagnetic iron oxide nanoparticles (SPIONs), namely, ferumoxides, monocrystalline iron oxide (MION), cross-linked iron oxide (CLIO)-NH(2) and tat-CLIO. MATERIALS AND METHODS: The hNSCs (5 x 10(5) HB1F3 cells/ml) were incubated for 24 hr in cell culture media that contained 25 microg/ml of ferumoxides, MION or CLIO-NH(2), and with or without poly-L-lysine (PLL) and tat-CLIO. The cellular iron uptake was analyzed qualitatively with using a light microscope and this was quantified via atomic absorption spectrophotometry. The visibility of the labeled cells was assessed with MR imaging. RESULTS: The incorporation of SPIONs into the hNSCs did not affect the cellular proliferations and viabilities. The hNSCs labeled with tat-CLIO showed the longest retention, up to 72 hr, and they contained 2.15+/-0.3 pg iron/cell, which are 59 fold, 430 fold and six fold more incorporated iron than that of the hNSCs labeled with ferumoxides, MION or CLIO-NH(2), respectively. However, when PLL was added, the incorporation of ferumoxides, MION or CLIO-NH(2) into the hNSCs was comparable to that of tat-CLIO. CONCLUSION: For MR imaging, hNSCs can be efficiently labeled with tat-CLIO alone or with a combination of ferumoxides, MION, CLIO-NH(2) and the transfection agent PLL.     

Early identification of aortic valve sclerosis using iron oxide enhanced MRI

Purpose:

To test the ability of MION‐47 enhanced MRI to identify tissue macrophage infiltration in a rabbit model of aortic valve sclerosis (AVS).

Materials and Methods:

The aortic valves of control and cholesterol‐fed New Zealand White rabbits were imaged in vivo pre‐ and 48 h post‐intravenous administration of MION‐47 using a 1.5 Tesla (T) MR clinical scanner and a CINE fSPGR sequence. MION‐47 aortic valve cusps were imaged ex vivo on a 3.0T whole‐body MR system with a custom gradient insert coil and a three‐dimensional (3D) FIESTA sequence and compared with aortic valve cusps from control and cholesterol‐fed contrast‐free rabbits. Histopathological analysis was performed to determine the site of iron oxide uptake.

Results:

MION‐47 enhanced the visibility of both control and cholesterol‐fed rabbit valves in in vivo images. Ex vivo image analysis confirmed the presence of significant signal voids in contrast‐administered aortic valves. Signal voids were not observed in contrast‐free valve cusps. In MION‐47 administered rabbits, histopathological analysis revealed iron staining not only in fibrosal macrophages of cholesterol‐fed valves but also in myofibroblasts from control and cholesterol‐fed valves.

Conclusion:

Although iron oxide labeling of macrophage infiltration in AVS has the potential to detect the disease process early, a macrophage‐specific iron compound rather than passive targeting may be required. J. Magn. Reson. Imaging 2010;31:110–116. © 2009 Wiley‐Liss, Inc.     

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. © 2010 Wiley‐Liss, Inc.     

Effectiveness of micron‐sized superparamagnetic iron oxide particles as markers for detection of migration of bone marrow‐derived mesenchymal stromal cells in a stroke model

Purpose:

To evaluate the feasibility of using micron‐sized superparamagnetic iron oxide particles (MPIOs) as an effective labeling agent for monitoring bone marrow‐derived mesenchymal stromal cell (BMSC) migration in the brain using magnetic resonance imaging (MRI) in a rat model of stroke and whether the accumulation of MPIO‐labeled BMSCs can be differentiated from the accumulation of free MPIO particles or hemoglobin breakdown at a site of neuronal damage.

Materials and Methods:

In this study BMSCs were labeled with iron oxide and their pattern of migration following intravenous injection in a rat stroke model was monitored using a clinical MRI system followed by standard histopathology. The migration pattern was compared between intravenous injection of BMSCs alone, BMSCs labeled with MPIOs, and MPIO particles alone.

Results:

The results demonstrated that while MRI was highly sensitive in the detection of iron oxide particle‐containing cells in areas of neuronal ischemia, the true origin of cells containing iron oxide particles remains ambiguous. Therefore, detection of iron particles may not be a suitable strategy for the detection of BMSCs in the brain in a stroke model.

Conclusion:

This study suggests that the use of MPIOs as labeling agents are insufficient to conclusively determine the localization of iron within cells in regions of neuronal ischemia and hemorrhage. J. Magn. Reson. Imaging 2013;37:1409–1418. © 2012 Wiley Periodicals, Inc.     

Coating thickness of magnetic iron oxide nanoparticles affects R2 relaxivity

PURPOSE: To evaluate the effect of coating thickness on the relaxivity of iron oxide nanoparticles. MATERIALS AND METHODS: Monocrystalline superparamagnetic iron oxide nanoparticles (MIONs), coated with a polyethylene glycol (PEG)-modified, phospholipid micelle coating, with different PEG molecular weights, were prepared. The particle diameters were measured with dynamic light scattering (DLS) and electron microscopy (EM). The R1 and R2 of MIONs were measured using a bench-top nuclear magnetic resonance (NMR) relaxometer. pH was varied for some measurements. Monte Carlo simulations of proton movement in a field with nanometer-sized magnetic inhomogeneities were performed. RESULTS: Increasing the molecular weight of the PEG portion of the micelle coating increased overall particle diameter. As coating thickness increases, the R2 decreases and the R1 increases. Changing pH has no effect on relaxivity. The Monte Carlo simulations suggest that the effect of coating size on R2 relaxivity is determined by two competing factors: the physical exclusion of protons from the magnetic field and the residence time for protons within the coating zone. CONCLUSION: Coating thickness can significantly impact the R2, and the R2/R1 ratio, of a MION contrast agent. An understanding of the relationship between coating properties and changes in relaxivity is critical for designing magnetic nanoparticle probes for molecular imaging applications using MRI.     

Relaxation effects of ferucarbotran‐labeled mesenchymal stem cells at 1.5T and 3T: Discrimination of viable from lysed cells

Human mesenchymal stem cells (hMSCs) were labeled with Ferucarbotran by simple incubation and cultured for up to 14 d. Iron content was determined by spectrometry and the intracellular localization of the contrast agent uptake was studied by electron and confocal microscopy. At various time points after labeling, ranging from 1 to 14 d, samples with viable or lysed labeled hMSCs, as well as nonlabeled controls, underwent MRI. Spin‐echo (SE) and gradient‐echo (GE) sequences with multiple TRs and TEs were used at 1.5T and 3T on a clinical scanner. Spectrometry showed an initial iron oxide uptake of 7.08 pg per cell. Microscopy studies revealed lysosomal compartmentalization. Contrast agent effects of hMSCs were persistent for up to 14 d after labeling. A marked difference in the T₂ effect of compartmentalized iron oxides compared to free iron oxides was found on T₂‐weighted sequences, but not on T‐weighted sequences. The observed differences may be explained by the loss of compartmentalization of iron oxide particles, the uniformity of distribution, and the subsequent increase in dephasing of protons on SE images. These results show that viable cells with compartmentalized iron oxides may—in principle—be distinguished from lysed cells or released iron oxides. Magn Reson Med, 2009. © 2009 Wiley‐Liss, Inc.     

Temporal and noninvasive monitoring of inflammatory‐cell infiltration to myocardial infarction sites using micrometer‐sized iron oxide particles

Micrometer‐sized iron oxide particles (MPIO) are a more sensitive MRI contrast agent for tracking cell migration compared to ultrasmall iron oxide particles. This study investigated the temporal relationship between inflammation and tissue remodeling due to myocardial infarction (MI) using MPIO‐enhanced MRI. C57Bl/6 mice received an intravenous MPIO injection for cell labeling, followed by a surgically induced MI seven days later (n = 7). For controls, two groups underwent either sham‐operated surgery without inducing an MI post‐MPIO injection (n = 7) or MI surgery without MPIO injection (n = 6). The MRIs performed post‐MI showed significant signal attenuation around the MI site for the mice that received an intravenous MPIO injection for cell labeling, followed by a surgically induced MI seven days later, compared to the two control groups (P < 0.01). The findings suggested that the prelabeled inflammatory cells mobilized and infiltrated into the MI site. Furthermore, the linear regression of contrast‐to‐noise ratio at the MI site and left ventricular ejection function suggested a positive correlation between the labeled inflammatory cell infiltration and cardiac function attenuation during post‐MI remodeling (r² = 0.98). In conclusion, this study demonstrated an MRI technique for noninvasively and temporally monitoring inflammatory cell migration into the myocardium while potentially providing additional insight concerning the pathologic progression of a myocardial infarction. Magn Reson Med, 2010. © 2009 Wiley‐Liss, Inc.     

Monocrystalline iron oxide nanoparticles: possible solution to the problem of surgically induced intracranial contrast enhancement in intraoperative MR imaging

BACKGROUND AND PURPOSE: Intraoperative MR imaging is increasingly being used to control the extent of surgical resection; however, surgical manipulation itself causes intracranial contrast enhancement, which is a source of error. Our purpose was to investigate the potential of monocrystalline iron oxide nanoparticles (MIONs) to solve this problem in an animal model. METHODS: In male Wistar rats, surgical lesions of the brain were produced. The animals underwent MR examination immediately afterward. In the first group, a paramagnetic contrast agent was administered, whereas the second group of animals received MIONs 1 day before surgery. In a third group of animals, malignant glioma cells were stereotactically implanted in the caudoputamen. Two weeks later, MIONs were IV injected and the tumor was (partially) resected. Immediately after resection, MR examination was performed to determine the extent of residual tumor. RESULTS: Surgically induced intracranial contrast enhancement was seen in all animals in which a paramagnetic contrast agent was used. Conversely, when MIONs had been injected, no signal changes that could be confused with residual tumor were detected. In the animals that had undergone (partial) resection of experimental gliomas, MR assessment of residual tumor was possible without any interfering surgically induced phenomena. CONCLUSION: Because MIONs are stored in malignant brain tumor cells longer than they circulate in the blood, their use offers a promising strategy to avoid surgically induced intracranial contrast enhancement, which is known to be a potential source of error in intraoperative MR imaging.     

Monitoring bone marrow‐originated mesenchymal stem cell traffic to myocardial infarction sites using magnetic resonance imaging

How stem cells promote myocardial repair in myocardial infarction (MI) is not well understood. The purpose of this study was to noninvasively monitor and quantify mesenchymal stem cells (MSC) from bone marrow to MI sites using magnetic resonance imaging (MRI). MSC were dual‐labeled with an enhanced green fluorescent protein and micrometer‐sized iron oxide particles prior to intra‐bone marrow transplantation into the tibial medullary space of C57Bl/6 mice. Micrometer‐sized iron oxide particles labeling caused signal attenuation in T₂*‐weighted MRI and thus allowed noninvasive cell tracking. Longitudinal MRI demonstrated MSC infiltration into MI sites over time. Fluorescence from both micrometer‐sized iron oxide particles and enhanced green fluorescent protein in histology validated the presence of dual‐labeled cells at MI sites. This study demonstrated that MSC traffic to MI sites can be noninvasively monitored in MRI by labeling cells with micrometer‐sized iron oxide particles. The dual‐labeled MSC at MI sites maintained their capability of proliferation and differentiation. The dual‐labeling, intra‐bone marrow transplantation, and MRI cell tracking provided a unique approach for investigating stem cells' roles in the post‐MI healing process. This technique can potentially be applied to monitor possible effects on stem cell mobilization caused by given treatment strategies. Magn Reson Med, 2011. © 2011 Wiley‐Liss, Inc.     

A physiological barrier distal to the anatomic blood-brain barrier in a model of transvascular delivery

BACKGROUND AND PURPOSE: Osmotic disruption of the blood-brain barrier (BBB) provides a method for transvascular delivery of therapeutic agents to the brain. The apparent global delivery of viral-sized iron oxide particles to the rat brain after BBB opening as seen on MR images was compared with the cellular and subcellular location and distribution of the particles. METHODS: Two dextran-coated superparamagnetic monocrystalline iron oxide nanoparticle contrast agents, MION and Feridex, were administered intraarterially in rats at 10 mg Fe/kg immediately after osmotic opening of the BBB with hyperosmolar mannitol. After 2 to 24 hours, iron distribution in the brain was evaluated first with MR imaging then by histochemical analysis and electron microscopy to assess perivascular and intracellular distribution. RESULTS: After BBB opening, MR images showed enhancement throughout the disrupted hemisphere for both Feridex and MION. Feridex histochemical staining was found in capillaries of the disrupted hemisphere. Electron microscopy showed that the Feridex particles passed the capillary endothelial cells but did not cross beyond the basement membrane. In contrast, after MION delivery, iron histochemistry was detected within cell bodies in the disrupted hemisphere, and the electron-dense MION core was detected intracellularly and extracellularly in the neuropil. CONCLUSION: MR images showing homogeneous delivery to the brain at the macroscopic level did not indicate delivery at the microscopic level. These data support the presence of a physiological barrier at the basal lamina, analogous to the podocyte in the kidney, distal to the anatomic (tight junction) BBB, which may limit the distribution of some proteins and viral particles after transvascular delivery to the brain.     

Release activation of iron oxide nanoparticles: (REACTION) A novel environmentally sensitive MRI paradigm

Smart contrast agents for MRI‐based cell tracking would enable the use of MRI methodologies to not only detect the location of cells but also gene expression. Here, we report on a new enzyme/contrast agent paradigm which involves the enzymatic degradation of the polymer coating of magnetic nanoparticles to release encapsulated magnetic cores. Cells were labeled with particles coated with a polymer, which is cleavable by a specific enzyme. This coat restricts the approach of water to the particle, preventing the magnetic core from efficiently relaxing protons. The reactive enzyme was delivered to cells and changes in cellular T₂ and T₂* relaxation times of ～ 35% and ～ 50% were achieved in vitro. Large enhancements of dark contrast volume (240%) and contrast‐to‐noise ratio (48%) within the contrast regions were measured, in vivo, for cells co‐labeled with enzyme and particles. These results warrant exploration of genetic avenues toward achieving release activation of iron oxide nanoparticles. Magn Reson Med, 2011. © 2011 Wiley‐Liss, Inc.     

Susceptibility gradient mapping (SGM): A new postprocessing method for positive contrast generation applied to superparamagnetic iron oxide particle (SPIO)‐labeled cells

Local susceptibility gradients result in a dephasing of the precessing magnetic moments and thus in a fast decay of the NMR signals. In particular, cells labeled with superparamagnetic iron oxide particles (SPIOs) induce hypointensities, making the in vivo detection of labeled cells from such a negative image contrast difficult. In this work, a new method is proposed to selectively turn this negative contrast into a positive contrast. The proposed method calculates the susceptibility gradient and visualizes it in a parametric map directly from a regular gradient‐echo image dataset. The susceptibility gradient map is determined in a postprocessing step, requiring no dedicated pulse sequences or adaptation of the sequence before and during image acquisition. Phantom experiments demonstrated that local susceptibility differences can be quantified. In vivo experiments showed the feasibility of the method for tracking of SPIO‐labeled cells. The method bears the potential also for usage in other applications, including the detection of contrast agents and interventional devices as well as metal implants. Magn Reson Med 60:595–603, 2008. © 2007 Wiley‐Liss, Inc.     

In vivo serial evaluation of superparamagnetic iron‐oxide labeled stem cells by off‐resonance positive contrast

MRI is emerging as a diagnostic modality to track iron‐oxide‐labeled stem cells. This study investigates whether an off‐resonance (OR) pulse sequence designed to generate positive contrast at 1.5T can assess the location, quantity, and viability of delivered stem cells in vivo. Using mouse embryonic stem cell transfected with luciferase reporter gene (luc‐mESC), multimodality validation of OR signal was conducted to determine whether engraftment parameters of superparamagnetic iron‐oxide labeled luc‐mESC (SPIO‐luc‐mESC) could be determined after cell transplantation into the mouse hindlimb. A significant increase in signal‐ and contrast‐to‐noise of the SPIO‐luc‐mESC was achieved with the OR technique when compared to a gradient recalled echo (GRE) sequence. A significant correlation between the quantity of SPIO‐luc‐mESC and OR signal was observed immediately after transplantation (R² = 0.74, P < 0.05). The assessment of transplanted cell viability by bioluminescence imaging (BLI) showed a significant increase of luciferase activities by day 16, while the MRI signal showed no difference. No significant correlation between BLI and MRI signals of cell viability was observed. In conclusion, using an OR sequence the precise localization and quantitation of SPIO‐labeled stem cells in both space and time were possible. However, the OR sequence did not allow evaluation of cell viability. Magn Reson Med 60:1269–1275, 2008. © 2008 Wiley‐Liss, Inc.     

Quantitative 19F imaging of nmol‐level F‐nucleotides/‐sides from 5‐FU with T2 mapping in mice at 9.4T

A unique acquisition method is proposed for quantitative, high‐sensitivity ¹⁹F MR spectroscopic imaging for the study of drug distribution aiming at nmol‐level metabolite information in mice. The use of fast spin echo (FSE) at 9.4T allowed us to obtain whole‐body images with minimal effect of magnetic susceptibility and to acquire several metabolite signals simultaneously by the method of interleaved multifrequency selection. Modified 2‐shot FSE was designed for simultaneous, high‐sensitivity ¹⁹F imaging and T₂ mapping. A time course study including all the main metabolites at 10‐minute resolution was attained with an oral dose of 1–2 mmol 5‐fluorouracil (5‐FU) (130–260 mg)/kg in mice. With acquisition parameters optimized for in vivo T₂ of 40 ms, images of F‐nucleotides/‐sides, effective anabolites of the anticancer drug 5‐FU, were obtained at the level of 200 nmol in the tumor for all the mice studied with a linear correlation (R = 0.96) between image intensity and the quantity determined in the excised tissue. The method exhibits potential capability of molecular imaging with a variety of ¹⁹F‐labeled compounds and drug evaluation. Magn Reson Med, 2009. © 2009 Wiley‐Liss, Inc.     

Chondrocyte gene expression is affected by very small iron oxide particles-labeling in long-term in vitro MRI tracking

Purpose:

To investigate the effect and dose response of very small iron oxide particles (VSOP) labeling of human chondrocytes for long‐term in vitro MRI tracking.

Materials and Methods:

Chondrocytes were isolated from cartilage biopsies from four patients. The cells for the dose–response study were labeled with 25, 50, or 100 μg/mL VSOP. Quantitative gene expression and cellular proliferation were compared with unlabeled controls at day 1, 3, and 7. The cells suited for MRI tracking were labeled with 50 μg/mL VSOP and embedded in alginate beads, followed by MRI (using T2‐weighted sequences) at day 0, 1, 3, 7, 14, 21, 28, and histology was performed at each time‐point.

Results:

Histology revealed that VSOP particles were intracellularly confined at all time‐points, whereas no extracellular VSOPs were observed. A mean reduction in T2‐value of 25.1 ms (±SD 3.5 ms) was found on T2‐maps. The chondrocyte‐specific genes aggrecan, collagen type 2, and sox9 were all affected by labeling, the two latter in a dose‐dependent manner. VSOPs had no effect on proliferation.

Conclusion:

VSOP labeling of chondrocytes affected gene expression but not proliferation. The labeled chondrocytes could be recognized by MRI for 4 weeks without significant changes in the T2 relaxation time. J. Magn. Reson. Imaging 2011;33:724–730. © 2011 Wiley‐Liss, Inc.     

Mechanisms of tissue–iron relaxivity: Nuclear magnetic resonance studies of human liver biopsy specimens

MRI is becoming an increasingly important tool to assess iron overload disorders, but the complex nature of proton–iron interactions has troubled noninvasive iron quantification. Intersite and intersequence variability as well as methodological inaccuracies have been limiting factors to its widespread clinical use. It is important to understand the underlying proton relaxation mechanisms within the (human) tissue environment to address these differences. In this respect, NMR relaxometry was performed on 10 fresh human liver biopsy specimens taken from patients with transfusion‐dependent anemia. T₁ (1/R₁) inversion recovery, T₂ (1/R₂) single echo, and multiecho T₂ CPMG measurements were performed on a 60‐MHz Bruker Minispectrometer. NMR parameters were compared to quantitative iron levels and tissue histology. Relaxivities R₁ and R₂ both increased linearly with hepatic iron content, with R₂ being more sensitive to iron. CPMG data were well described by a chemical‐exchange model and predicted effective iron center dimensions consistent with hemosiderin‐filled lysosomes. Nonexponential relaxation was evident at short refocusing intervals with R₂ and amplitude behavior suggestive of magnetic susceptibility‐based compartmentalization rather than anatomic subdivisions. NMR relaxometry of human liver biopsy specimens yields unique insights into the mechanisms of tissue–iron relaxivity. Magn Reson Med, 2005. © 2005 Wiley‐Liss, Inc.     

Hyperpolarized 13C spectroscopy and an NMR‐compatible bioreactor system for the investigation of real‐time cellular metabolism

The purpose of this study was to combine a three‐dimensional NMR‐compatible bioreactor with hyperpolarized ¹³C NMR spectroscopy in order to probe cellular metabolism in real time. JM1 (immortalized rat hepatoma) cells were cultured in a three‐dimensional NMR‐compatible fluidized bioreactor. ³¹P spectra were acquired before and after each injection of hyperpolarized [1‐¹³C] pyruvate and subsequent ¹³C spectroscopy at 11.7 T. ¹H and two‐dimensional ¹H‐¹H‐total correlation spectroscopy spectra were acquired from extracts of cells grown in uniformly labeled ¹³C‐glucose, on a 16.4 T, to determine ¹³C fractional enrichment and distribution of ¹³C label. JM1 cells were found to have a high rate of aerobic glycolysis in both two‐dimensional culture and in the bioreactor, with 85% of the ¹³C label from uniformly labeled ¹³C‐glucose being present as either lactate or alanine after 23 h. Flux measurements of pyruvate through lactate dehydrogenase and alanine aminotransferase in the bioreactor system were 12.18 ± 0.49 nmols/sec/10⁸ cells and 2.39 ± 0.30 nmols/sec/10⁸ cells, respectively, were reproducible in the same bioreactor, and were not significantly different over the course of 2 days. Although this preliminary study involved immortalized cells, this combination of technologies can be extended to the real‐time metabolic exploration of primary benign and cancerous cells and tissues prior to and after therapy. Magn Reson Med, 2010. © 2010 Wiley‐Liss, Inc.