Tracking of myelin-reactive T cells in experimental autoimmune encephalomyelitis (EAE) animals using small particles of iron oxide and MRI.

Myelin-reactive T cells are responsible for initiating the cascade of autoreactive immune responses leading to the development of multiple sclerosis. For better insights into the disease mechanism, it is of major importance to have knowledge on the sites at which these cells are active during disease progression. Herein, we investigated the feasibility of tracking myelin-reactive T cells, upon labelled with SPIO particles, in the central nervous system (CNS) of experimental autoimmune encephalomyelitis (EAE) animals by MRI. First, we determined the optimal labelling condition leading to a high particle uptake and minimal SPIO-Poly-l-lysine (PLL) aggregate formation using Prussian blue staining and inductively coupled plasma spectroscopy measurements. Results from labelling of myelin reactive T cells with low concentrations of SPIO particles (i.e. 25 microg/ml) combined with different concentrations of PLL (0-1.5 microg/ml) showed that increasing amounts of PLL led to augmented levels of free remnant SPIO-PLL aggregates. In contrast, a low PLL concentration (i.e. 0.5 microg/ml) combined with high concentrations of SPIO (i.e. 400 microg Fe/ml) led to a high labelling efficiency with minimal amounts of aggregates. Second, the labelled myelin-reactive T cells were transferred to control rats to induce EAE. At the occurrence of hindlimb paralysis, the SPIO labelled myelin-reactive T cells were detected in the sacral part of the spinal cord and shown to be highly confined to this region. However, upon transfer in already primed rats, T cells were more widely distributed in the CNS and shown present in the spinal cord as well as in the brain. Our study demonstrates the feasibility of tracking SPIO labelled myelin-reactive T cells in the spinal cord as well as the brain of EAE rats upon systemic administration. Furthermore, we provide data on the optimal labelling conditions for T cells leading to a high particle uptake and minimal aggregate formation.

A joined theoretical-experimental investigation on the 1H and 13C NMR signatures of defects in poly(vinyl chloride).

1H and 13C chemical shifts of PVC chains have been evaluated using quantum chemistry methods in order to evidence and interpret the NMR signatures of chains bearing unsaturated and branched defects. The geometrical structures of the stable conformers have been determined using molecular mechanics and the OPLS force field and then density functional theory with the B3LYP functional and the 6-311G(d) basis set. The nuclear shielding tensor has been calculated at the coupled-perturbed Kohn-Sham level (B3LYP exchange-correlation functional) using the 6-311+G(2d,p) basis set. The computational scheme accounts for the large number of stable conformers of the PVC chains, and average chemical shifts are evaluated using the Maxwell-Boltzmann distribution. Moreover, the chemical shifts are corrected for the inherent and rather systematic errors of the method of calculation by employing linear regression equations, which have been deduced from comparing experimental and theoretical results on small alkane model compounds containing Cl atoms and/or unsaturations. For each type of defect, PVC segments presenting different tacticities have been considered because it is known from linear PVC chains that the racemic (meso) dyads are characterized by larger (smaller) chemical shifts. NMR signatures of unsaturations in PVC chains have been highlighted for the internal -CH=CH- and -CH=CCl- units as well as for terminal unsaturations like the chloroallylic -CH=CH-CH2Cl group. In particular, the 13C chemical shifts of the two sp2 C atoms are very close for the chloroallylic end group. The CH2 and CHCl units surrounding an unsaturation present also specific 13C chemical shifts, which allow distinguishing them from the others. In the case of the proton, the CH2 unit of the -CHCl-CH2-CCl=CH- segment presents a larger chemical shift (2.6-2.7 ppm), while some CHCl units close to the -CH=CH- unsaturations appear at rather small chemical shifts (3.7 ppm). The -CH2Cl and -CHCl-CH2Cl branches also display specific signatures, which result in large part from modifications of the equilibrium conformations and their reduced number owing to the increased steric interactions. These branches lead to the appearance of 13C peaks at lower field associated either to the CH unit linking the -CH2Cl and -CHCl-CH2Cl branches (50 ppm) or to the CHCl unit of the ethyl branches (60 ppm). The corresponding protons resonate also at specific frequencies: 3.5-4.0 ppm for the -CH2Cl branch or 3.8-4.2 ppm for the terminal unit of the -CHCl-CH2Cl branch. Several of these signatures have been detected in the experimental 1H and 13C NMR spectra and are consistent with the reaction mechanisms.

Rapid, postmortem 9.4 T MRI of spinal cord injury: correlation with histology and survival times.

High field magnetic resonance imaging (MRI) has been increasingly used to assess experimental spinal cord injury (SCI). In the present investigation, after partial spinal cord injury and excision of the whole spine, pathological changes of the spinal cord were studied in spinal cord-spine blocks, from the acute to the chronic state (24 h to 5 months). Using proton density (PD) weighted imaging parameters at a magnetic field strength of 9.4 tesla (T), acquisition times ranging from <1 to 10 h per specimen were used. High in-plane pixel resolution (68 and 38 microm, respectively) was obtained, as well as high signal-to-noise ratio (SNR), which is important for optimal contrast settings. The quality of the resulting MR images was demonstrated by comparison with histology. The cord and the lesion were shown in their anatomical surroundings, detecting cord swelling in the acute phase (24 h to 1 week) and cord atrophy at the chronic stage. Haemorrhage was detected as hypo-intense signal. Oedema, necrosis and scarring were hyper-intense but could not be distinguished. Histology confirmed that the anatomical delimitation of the lesion extent by MRI was precise, both with high and moderate resolution. The present investigation thus demonstrates the precision of spinal cord MRI at different survival delays after compressive partial SCI and establishes efficient imaging parameters for postmortem PD MRI.

Visualisation of the kinetics of macrophage infiltration during experimental autoimmune encephalomyelitis by magnetic resonance imaging.

Macrophages are considered to be the predominant effector cells in multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). Ultra small particles of iron oxide (USPIO) can be used to detect macrophage infiltrates in the CNS with magnetic resonance imaging (MRI). Here, we investigated whether the kinetics of lesion formation in EAE can be visualised by altering the time point of USPIO injection and the time interval between particle injection and MRI. When USPIO are systemically injected 24 h before MRI, hypo intense regions are detected in different brain regions depending on the disease stage. These regions correspond to sites of macrophage infiltration. A more complete visualisation of sites of inflammation is accomplished by USPIO injection at disease onset and postponing MRI to top of disease. This study demonstrates that the distribution pattern and amount of inflammatory lesions detected with USPIO, depends on timing of USPIO administration and subsequent MRI. These findings are important for a correct application and interpretation of USPIO dependent contrast imaging of CNS inflammation.

The microanatomical environment of the subthalamic nucleus. Technical note.

High-frequency stimulation of the subthalamic nucleus (STN) is a widely performed method to treat advanced Parkinson disease. Due to the limitations of current imaging techniques, the 3D microanatomy of the STN and its surrounding structures in the mesencephalon are not well known. Using images they obtained using a 9.4-tesla magnetic resonance (MR) imaging unit, the authors developed a 3D reconstruction of the STN and its immediate surroundings. During the postmortem investigation of a human brain, a sample of tissue in the area around the STN was isolated. This brain tissue was scanned in the three orthogonal planes at 1-mm slice thickness. The images generated were compared with photographs of conventionally stained brain tissue slices in different neuroanatomical books, and a 3D reconstruction was made. High-field MR imaging is an appropriate method for visualizing the microanatomy of the STN and its surroundings. The images allow an optimal analysis of the microenvironment of the STN in the three orthogonal planes and can be used for 3D reconstructions of this area with possible clinical applications in the future.

Neurovascularization of the anterior jaw bones revisited using high-resolution magnetic resonance imaging.

The anterior jaw bones are often considered relatively safe surgical sites. Nonetheless, the increasing rate of surgical interventions in that area, such as oral implant placement and bone grafting, has highlighted the potential risks and has raised the reported complications. A careful documentation of all anatomic variations in anterior jaw bone neurovascularization has thus become necessary. The present report attempts to revisit jaw bone neurovascularization, addressing typical anatomic appearances and variations. We summarize the results of various microanatomical studies carried out by high-resolution magnetic resonance imaging (HR-MRI) of the human anterior jaw bones. These studies reveal that edentulous and dentate anterior jaws present significant variation in the occurrence of the mandibular incisive canal and genial spinal foramina, as well as the maxillary nasopalatine canal. All of these canal structures contain a neurovascular bundle, whose diameter may be large enough to cause clinically significant trauma. A careful presurgical radiographic analysis of the anterior jaw bones is therefore advised.

Development of starch-based pellets via extrusion/spheronisation.

A modified starch (high-amylose, crystalline and resistant starch) was evaluated as an alternative excipient to microcrystalline cellulose for pellets prepared via extrusion/spheronisation. Theophylline anhydrous (25%, w/w) was used as a model drug. A binder was necessary to obtain an acceptable yield and the addition of sorbitol improved the surface properties of the pellets. A surface response design with three formulation variables (binder, sorbitol and water level) and one process variable (spheronisation speed) was used to optimise the process and to evaluate pellet yield, sphericity (aspect ratio and two-dimensional shape factor, e(R)), size (mean Feret diameter), friability and disintegration properties. Mixer torque rheometry and solid-state NMR revealed a significant influence of sorbitol on wet mass consistency and pellet properties. A high pellet yield (>90%), acceptable sphericity (AR<1.2), low friability (<0.01%), fast disintegration (<10 min) and complete drug release in less than 20 min for all formulations, demonstrated the potential of this modified starch in formulations intended for extrusion/spheronisation.

Correlation of postmortem 9.4 tesla magnetic resonance imaging and immunohistopathology of the human thoracic spinal cord 7 months after traumatic cervical spine injury.

OBJECTIVE: To correlate high-resolution magnetic resonance imaging (MRI) with immunohistopathology in the injured human spinal cord. METHODS: Postmortem MRI scans at a field strength of 9.4 T, as well as standard histology and immunohistochemistry, were performed on an excised specimen of human high thoracic spinal cord, obtained 7 months after the initial trauma, several segments below a severe spinal cord lesion (C5). RESULTS: A precise correlation is described between MRI and immunohistochemistry of the long white matter tracts undergoing Wallerian degeneration and of an extension of the cervical lesion into the high thoracic cord. CONCLUSION: MRI, the only imaging technique that currently provides useful information on the spinal cord parenchyma after trauma, is rapidly evolving. High-field scanners of up to 9.4 T are being clinically tested. The present postmortem investigation of an isolated spinal cord specimen demonstrates the precise correlation that can be achieved between imaging and pathology. In future investigations, this type of technique can lead to a more precise description of spinal cord injuries and their consequences in remote tissue. Translation into the clinical setting will improve diagnosis and follow-up of spinal cord injured patients.

Femoral neck trabecular microstructure in ovariectomized ewes treated with calcitonin: MRI microscopic evaluation.

UNLABELLED: Ovariectomy induces deterioration of the trabecular structure in the femoral neck of ewes, as depicted by MR microscopic imaging. This structural deterioration is prevented by salmon calcitonin treatment. INTRODUCTION: This study evaluated the trabecular (Tb) microarchitecture of an ovariectomy (OVX)-induced osteoporotic model in ewes and determined the effects of salmon calcitonin (sCT), an osteoclast inhibitor, on the Tb structure. This is the first report of OVX-induced changes in the Tb structure in the femoral neck in the ewes and effect of sCT on the microarchitecture. MATERIALS AND METHODS: Ewes (5-8 years old, n = 28) were equally allocated into sham (Sham), OVX injected with vehicle, or OVX injected with sCT at 50 or 100 IU, three injections per week. They were killed 6 months after OVX. The femoral neck was examined with an MR imager at 9.4 T in axial, coronal, and sagittal planes. An internal calibration procedure as a means of standardizing image analysis was used to adjust the segmentation threshold. Data from all three planes were averaged. RESULTS AND CONCLUSIONS: Compared with Sham, OVX induced significant changes (p < 0.0125) in the MRI-derived femoral neck Tb structure: Tb bone volume fraction (BV/TV), -18%; Tb number, -20%; Tb separation, +23%; number of free ends, +28%; number of nodes, -39%; number of Tb branches, -23%; mean length of Tb branches, -19%. Compared with OVX, treatment of sCT at 100 IU significantly improved all the Tb structural parameters to the Sham level (p < 0.0001 approximately p = 0.0281), whereas 50 IU significantly increased the Tb number and the mean length of the Tb branches. BV/TV explained 74% of the variation of compressive stress of the trabecular cylinder cores of the femoral neck. Combining all structural parameters in a multivariate regression analysis significantly improved the explanation to 84%, and adding BMD further improved the predictive ability of the model to 92%. We conclude that OVX induces deterioration of the MRI-derived Tb microstructure in the femoral neck of ewes. sCT treatment prevents OVX-induced changes. The femoral neck microarchitecture significantly correlates with its biomechanical properties. Combining microstructural parameters with BMD further improves the prediction of bone biomechanical properties. The effects of sCT on OVX ewes may help explain reduced fracture risk in postmenopausal osteoporotic women treated with sCT.

Synthesis and complete NMR spectral assignment of thiophene-substituted sulfinyl monomers.

This paper describes the synthesis of thiophene-substituted sulfinyl monomers. It comprises a four-step reaction by which the thiophene unit is built in via Suzuki coupling. These monomers could be used as building blocks for the preparation of conducting polymers via a new concept: the sulfinyl precursor route i.e. via thiophene substituted poly(p-phenylenevinylene) precursors. Furthermore, the complete 1H and 13C NMR signal assignment is presented. In addition to being essential for the characterization of the polymers concerned, it offers useful input information for further improvement of chemical shift prediction software. Furthermore, the T1C relaxation decay times are demonstrated to have the potential of being a fast and robust criterion for the spectral assignment of analogous monomers.

In vitro high-field magnetic resonance imaging-documented anatomy of a fetal myelomeningocele at 20 weeks' gestation. A contribution to the rationale of intrauterine surgical repair of spina bifida.

OBJECT: It remains uncertain if closure of a myelomeningocele at midgestation changes the neurological condition at birth in an infant born with spina bifida. The authors conducted a study to provide a detailed analysis of the morphology of the spinal cord with the myelomeningocele at the time fetal surgery usually is performed. METHODS: The myelomeningocele of a 20-week-gestation-age fetus was examined, and data were compared with those obtained in a neurologically intact specimen of the same age. In vitro high-field 9.4-tesla magnetic resonance (MR) microscopy was used to examine the fetal material. High-field MR spectroscopy provided images in the three orthogonal planes with a resolution comparable with low-power optical microscopy. The authors observed that the fetal cord of the myelomeningocele specimen was tapered and tethered at S3-4 while the conus medullaris in the normal fetus reaches L-4. No neurulation defects were noted. The axial MR images clearly revealed the nonfusion of the mesodermal structures. The absence of neurulation defects suggests that at least in some cases of spina bifida the spinal cord initially is well developed but is damaged later on chemically and mechanically. This might be an argument in favor of intrauterine myelomeningocele repair. By 20 weeks' gestation, however, the deformation of the cord inside the myelomeningocele is severe. An optimization of the preoperative assessment by means of MR imaging therefore might be considered a valuable contribution to intrauterine surgery. The in vitro high-field MR microscopic findings of this study could be used as references for clinical intrauterine MR imaging. CONCLUSIONS: The detailed in vitro high-field MR analysis of a 20-week-gestation-age fetus with spina bifida demonstrated that an improvement of the preoperative intrauterine imaging should be pursued to detect those cases without neurulation defects and with minimal deformation of the spinal cord.

Highly Selective Route for Producing Unsymmetrically Substituted Monomers toward Synthesis of Conjugated Polymers Derived from Poly(p-phenylene vinylene).

A new convenient route for producing unsymmetrically substituted sulfinyl monomers of precursor polymers toward poly(p-phenylene vinylene) is described. Upon treating a symmetrical bissulfonium salt with a thiolate anion, an unexpected high selectivity for the monosubstituted thioethers (90%) is obtained. Optimization of the reaction conditions showed that the stoichiometry of the reactants in this reaction is important to ensure the high selectivity and to prevent unwanted side reactions. Reaction of equimolar amounts of reagents at ambient temperature gave the best results. A mechanism consistent with these results, supported by UV-vis experiments, is presented. Selective oxidation of the thioethers yielded the sulfinyl monomers. By using this new route, it was possible to increase the overall yield by a factor of 2, as compared to the route previously used to obtain these compounds.