Gerd Melkus

Bone marrow fat quantification in the presence of trabecular bone: initial comparison between water-fat imaging and single-voxel MRS.

The purpose of the present study was to test the relative performance of chemical shift‐based water‐fat imaging in measuring bone marrow fat fraction in the presence of trabecular bone, having as reference standard the single‐voxel magnetic resonance spectroscopy (MRS).

Combined Noninvasive Imaging and Modeling Approaches Reveal Metabolic Compartmentation in the Barley Endosperm

This research integrates dynamic NMR-based imaging and metabolic modeling to study the spatial arrangement of metabolism in the living grain of barley. Distinct metabolic compartments within the endosperm are uncovered and visualized using Ala as a marker. The beneficial role of metabolic compartmentation for carbon and nitrogen economy is discussed. The starchy endosperm of cereals is a priori taken as a metabolically uniform tissue. By applying a noninvasive assay based on 13C/1H-magnetic resonance imaging (MRI) to barley (Hordeum vulgare) grains, we uncovered metabolic compartmentation in the endosperm. 13C-Suc feeding during grain filling showed that the primary site of Ala synthesis was the central region of the endosperm, the part of the caryopsis experiencing the highest level of hypoxia. Region-specific metabolism in the endosperm was characterized by flux balance analysis (FBA) and metabolite profiling. FBA predicts that in the central region of the endosperm, the tricarboxylic acid cycle shifts to a noncyclic mode, accompanied by elevated glycolytic flux and the accumulation of Ala. The metabolic compartmentation within the endosperm is advantageous for the grains carbon and energy economy, with a prominent role being played by Ala aminotransferase. An investigation of caryopses with a genetically perturbed tissue pattern demonstrated that Ala accumulation is a consequence of oxygen status, rather than being either tissue specific or dependent on the supply of Suc. Hence the 13C-Ala gradient can be used as an in vivo marker for hypoxia. The combination of MRI and metabolic modeling offers opportunities for the noninvasive analysis of metabolic compartmentation in plants.

Dynamic 13C/1H NMR imaging uncovers sugar allocation in the living seed

Seed growth and accumulation of storage products relies on the delivery of sucrose from the maternal to the filial tissues. The transport route is hidden inside the seed and has never been visualized in vivo. Our approach, based on high-field nuclear magnetic resonance and a custom made (13)C/(1) H double resonant coil, allows the non-invasive imaging and monitoring of sucrose allocation within the seed. The new technique visualizes the main stream of sucrose and determines its velocity during the grain filling in barley (Hordeum vulgare L.). Quantifiable dynamic images are provided, which allow observing movement of (13)C-sucrose at a sub-millimetre level of resolution. The analysis of genetically modified barley grains (Jekyll transgenic lines, seg8 and Risø13 mutants) demonstrated that sucrose release via the nucellar projection towards the endosperm provides an essential mean for the control of seed growth by maternal organism. The sucrose allocation was further determined by structural and metabolic features of endosperm. Sucrose monitoring was integrated with an in silico flux balance analysis, representing a powerful platform for non-invasive study of seed filling in crops.

The metabolic role of the legume endosperm: a noninvasive imaging study.

Although essential for normal seed development in the legumes, the metabolic role of the endosperm remains uncertain. We designed noninvasive nuclear magnetic resonance tools for the in vivo study of key metabolites in the transient liquid endosperm of intact pea (Pisum sativum) seeds. The steady-state levels of sucrose, glutamine, and alanine could be monitored and their distribution within the embryo sac visualized. Seed structure was digitalized as a three-dimensional model, providing volume information for distinct seed organs. The nuclear magnetic resonance method, combined with laser microdissection, isotope labeling, in situ hybridization, and electron microscopy, was used to contrast the wild-type endosperm with that of a mutant in which embryo growth is retarded. Expression of sequences encoding amino acid and sucrose transporters was up-regulated earlier in the endosperm than in the embryo, and this activity led to the accumulation of soluble metabolites in the endosperm vacuole. The endosperm provides a temporary source of nutrition, permits space for embryo growth, and acts as a buffer between the maternal organism and its offspring. The concentration of sucrose in the endosperm vacuole is developmentally controlled, while the total amount accumulated depends on the growth of the embryo. The endosperm concentration of glutamine is a limiting factor for protein storage. The properties of the endosperm ensure that the young embryo develops within a homeostatic environment, necessary to sustain embryogenesis. We argue for a degree of metabolite-mediated control exerted by the endosperm on the growth of, and assimilate storage by, the embryo.

Diffusion-Tensor Imaging of Human Articular Cartilage Specimens with Early Signs of Cartilage Damage

PURPOSE To assess the use of diffusion-tensor (DT) imaging of articular cartilage to detect and grade early cartilage damage in human specimens with early signs of cartilage damage. MATERIALS AND METHODS This study was approved by the institutional review board. Forty-three cartilage-on-bone samples drilled from 21 human patellae were examined with 17.6-T magnetic resonance (MR) imaging and a diffusion-weighted spin-echo sequence (spatial resolution, 50 × 100 × 800 μm). Subsequently, samples underwent histologic analysis with safranin O staining. Cartilage damage on safranin O histologic slides was quantified with Osteoarthritis Research Society International (OARSI) grades; grades ranged from 0 (healthy) to 6 (bone remodeling). Maps of longitudinal diffusivity (λ(l)), transverse diffusivity (λ(t)), mean diffusivity (MD), and fractional anisotropy (FA) were calculated. Cartilage was segmented, and region of interest (ROI) analysis was performed and compared with histologic findings. Significant differences in MR parameters between the OARSI groups were assessed with the Tukey test. The value of DT imaging in the diagnosis and grading of cartilage damage was assessed with logistic regression analysis. RESULTS Samples had OARSI grades of 0 (n = 14), 1 (n = 11), 2 (n = 12), 3 (n = 4), and 4 (n = 2). Samples with an OARSI grade greater than 0 had significantly increased λ(l), λ(t), and MD (7%-25% increase) in the superficial cartilage growing deeper into cartilage with increasing OARSI grade. Samples with an OARSI grade greater than 0 showed significantly decreased FA in the deep cartilage (-25% to -35% decrease), suggesting that changes in the collagen architecture may occur early in cartilage degradation. DTI showed excellent performance in the detection of cartilage damage (accuracy, 0.95; 41 of 43 samples) and good performance in the grading of cartilage damage (accuracy, 0.74; 32 of 43 samples). CONCLUSION DT imaging of articular cartilage can enable physicians to detect and grade early cartilage damage.

Change of diffusion tensor imaging parameters in articular cartilage with progressive proteoglycan extraction

Objective:To investigate changes of diffusion tensor imaging (DTI) parameters (mean apparent diffusion coefficient [ADC], fractional anisotropy [FA], and first eigenvector) with increasing proteoglycan (PG) extraction of articular cartilage. Material and Methods:Twelve cylindrical cartilage-on-bone samples were drilled from 4 human patellae (3 per patella). Each sample was divided into 2 pieces. One piece underwent histologic examination to assess the PG content of the native sample by safranin-O staining and its collagen architecture by polarized light microscopy. The other underwent magnetic resonance imaging (MRI) at 17.6 T for DTI measurement. After MRI, 2 of the 3 samples from each patella were immersed in a dilute trypsin solution (0.1 mg/mL), whereas the third sample was kept as a negative control in physiological saline. After incubation (6, 48, 72, and 96 hours), the samples were reimaged, stained for PG content and for the collagen orientation. Maps of ADC, FA, and the orientation of the first eigenvector as well as histology were available for each sample before and after incubation. Results:PG loss led to increased ADC and reduced safranin-O staining from the articular surface to the bone-cartilage interface. A significant correlation (r2 = 0.86, P < 0.01) was observed between the change in bulk ADC and PG loss. Regional analysis from the articular surface to the tide mark demonstrated depth dependent significant correlations of ADC and PG loss. FA and first eigenvector as well as polarized light microscopy showed only small changes in the order of magnitude of measurement errors, not correlating with PG loss. Conclusion:Mean diffusivity evidence by the ADC is linearly correlated to progressive PG extraction in articular cartilage. FA and the first eigenvector seem to be specific to the collagen architecture of cartilage. DTI has the potential to become a valuable biomarker for the workup of cartilage degeneration in osteoarthritis, since evaluation of the PG content and collagen architectural properties of cartilage can be performed with a single, non–contrast-enhanced proton-based MRI measurement.

MRI quantification of fatty infiltration and muscle atrophy in a mouse model of rotator cuff tears

Rotator cuff pathology is the most common shoulder problem seen by orthopedic surgeons. Rotator cuff muscle fatty infiltration and muscle atrophy are common in larger tears and are considered predicting factors for the prognosis of cuff repair. Clinically, MRI is the gold standard in determining fatty infiltration and muscle atrophy; however, analysis for MRI imaging is primarily qualitative in nature with the results lacking further validation. We have recently developed a mouse model of rotator cuff tears. The goal of this study is to quantify and verify rotator cuff muscle atrophy and fatty infiltration using high‐resolution MRI in our mouse model. The rotator cuff muscles were analyzed for fat using a triglyceride quantification assay (TQA), muscle volume was measured through water displacement (WD), and histology. The study revealed that MRI had a high correlation with fat as measured with histology and TQA (R2 = 098). MRI also correlated well with atrophy measured with WD and wet weight. This suggests that MRI is a reliable modality in evaluating the progression of fatty infiltration and muscle atrophy following rotator cuff tears in a small animal model.

Spatially localized intermolecular zero-quantum coherence spectroscopy for in vivo applications

Magnetic resonance spectroscopy (MRS) techniques that use the distant dipolar field (DDF) to locally refocus inhomogeneous line‐broadening promise improved spectral resolution in spatially varying fields. We investigated three possible implementations of localized DDF spectroscopy. Theoretical analysis and phantom experiments at 17.6 T showed that only localization immediately prior to acquisition provides sufficient spatial selectivity and sensitivity for in vivo applications. Spectra from an (8 mm)3 voxel of the rat brain were acquired in 25 min, and three major metabolites were resolved. In a tumor mouse model, DDF spectra with well‐resolved lines can be obtained from significantly larger voxels compared to conventional localized spectroscopy. From an inhomogeneous voxel, improved spectral resolution can be obtained with DDF techniques when a sufficient number of increments are sampled along the second spectral dimension. With fewer increments, measurement time is significantly shortened, and DDF techniques can provide higher signal‐to‐noise ratio (SNR) efficiency. Magn Reson Med, 2006.

A Noninvasive Platform for Imaging and Quantifying Oil Storage in Submillimeter Tobacco Seed

Summary: This study describes a non-invasive NMR approach that enables counting of seeds inside the intact tobacco capsule, to measure seed sizes, to model the seed interior in three dimensions, to quantify the lipid content, and to visualize lipid gradients. While often thought of as a smoking drug, tobacco (Nicotiana spp.) is now considered as a plant of choice for molecular farming and biofuel production. Here, we describe a noninvasive means of deriving both the distribution of lipid and the microtopology of the submillimeter tobacco seed, founded on nuclear magnetic resonance (NMR) technology. Our platform enables counting of seeds inside the intact tobacco capsule to measure seed sizes, to model the seed interior in three dimensions, to quantify the lipid content, and to visualize lipid gradients. Hundreds of seeds can be simultaneously imaged at an isotropic resolution of 25 µm, sufficient to assess each individual seed. The relative contributions of the embryo and the endosperm to both seed size and total lipid content could be assessed. The extension of the platform to a range of wild and cultivated Nicotiana species demonstrated certain evolutionary trends in both seed topology and pattern of lipid storage. The NMR analysis of transgenic tobacco plants with seed-specific ectopic expression of the plastidial phosphoenolpyruvate/phosphate translocator, displayed a trade off between seed size and oil concentration. The NMR-based assay of seed lipid content and topology has a number of potential applications, in particular providing a means to test and optimize transgenic strategies aimed at the manipulation of seed size, seed number, and lipid content in tobacco and other species with submillimeter seeds.

Diffusion tensor imaging and T2 relaxometry of bilateral lumbar nerve roots: feasibility of in-plane imaging

Lower back pain is a common problem frequently encountered without specific biomarkers that correlate well with an individual patients pain generators. MRI quantification of diffusion and T2 relaxation properties may provide novel insight into the mechanical and inflammatory changes that occur in the lumbosacral nerve roots in patients with lower back pain. Accurate imaging of the spinal nerve roots is difficult because of their small caliber and oblique course in all three planes. Two‐dimensional in‐plane imaging of the lumbosacral nerve roots requires oblique coronal imaging with large field of view (FOV) in both dimensions, resulting in severe geometric distortions using single‐shot echo planar imaging (EPI) techniques. The present work describes initial success using a reduced‐FOV single‐shot spin‐echo EPI acquisition to obtain in‐plane diffusion tensor imaging (DTI) and T2 mapping of the bilateral lumbar nerve roots at the L4 level of healthy subjects, minimizing partial volume effects, breathing artifacts and geometric distortions. A significant variation in DTI and T2 mapping metrics is also reported along the course of the normal nerve root. The fractional anisotropy is statistically significantly lower in the dorsal root ganglia (0.287 ± 0.068) than in more distal regions in the spinal nerve (0.402 ± 0.040) (p < 10–5). The T2 relaxation value is statistically significantly higher in the dorsal root ganglia (78.0 ± 11.9 ms) than in more distal regions in the spinal nerve (59.5 ± 7.4 ms) (p < 10–5). The quantification of nerve root DTI and T2 properties using the proposed methodology may identify the specific site of any degenerative and inflammatory changes along the nerve roots of patients with lower back pain. Copyright