Silvia Lope-Piedrafita

Allopregnanolone treatment, both as a single injection or repetitively, delays demyelination and enhances survival of Niemann-Pick C mice.

Niemann‐Pick C disease (NPC) is an irreversible neurodegenerative disorder without current treatment. It is thought to result from deficient intracellular cholesterol and/or ganglioside trafficking. We have investigated the effects of allopregnanolone treatments on survival, weight loss, motor function, magnetic resonance imaging (MRI), and neuropathology in the mouse model of NPC (Npc1–/– mice). We confirmed previous results showing that a single injection of 250 μg of allopregnanolone on postnatal day 7 significantly extended the life span of Npc1–/– mice. This caused a marked difference in the weight curves of the treated mice but no statistical difference in the Rota‐Rod performance. T2‐weighted MRI and diffusion tensor imaging (DTI) of treated mice showed values of signal intensity and fractional anisotropy closer to those of wild‐type mice than those of untreated Npc1–/– mice. Neuropathology showed that day‐7 treatment markedly suppressed astrocyte reaction and significantly reduced microglial activation. Furthermore, the steroid treatment also increased myelination in brains of Npc1–/– mice. Similar effects of allopregnanolone treatment were observed in Npc1–/–, mdr1a–/– double‐mutant mice, which have a deficient blood–brain barrier, resulting in increased steroid uptake. The effects on survival and weight loss of a single injection on day 7 followed by injections every 2 weeks were also evaluated in Npc1–/– mice, and the beneficial effects were found to be greater than with the single injection at day 7. We conclude that allopregnanolone treatment significantly ameliorates several symptoms of NPC in Npc1–/– mice, presumably by effects on myelination or neuronal connectivity.

Factors Secreted by Endothelial Progenitor Cells Enhance Neurorepair Responses after Cerebral Ischemia in Mice

Cell therapy with endothelial progenitor cells (EPCs) has emerged as a promising strategy to regenerate the brain after stroke. Here, we aimed to investigate if treatment with EPCs or their secreted factors could potentiate angiogenesis and neurogenesis after permanent focal cerebral ischemia in a mouse model of ischemic stroke. BALB/C male mice were subjected to distal occlusion of the middle cerebral artery, and EPCs, cell-free conditioned media (CM) obtained from EPCs, or vehicle media were administered one day after ischemia. Magnetic resonance imaging (MRI) was performed at baseline to confirm that the lesions were similar between groups. Immunohistochemical and histological evaluation of the brain was performed to evaluate angio-neurogenesis and neurological outcome at two weeks. CM contained growth factors, such as VEGF, FGF-b and PDGF-bb. A significant increase in capillary density was noted in the peri-infarct areas of EPC- and CM-treated animals. Bielschowsky’s staining revealed a significant increase in axonal rewiring in EPC-treated animals compared with shams, but not in CM-treated mice, in close proximity with DCX-positive migrating neuroblasts. At the functional level, post-ischemia forelimb strength was significantly improved in animals receiving EPCs or CM, but not in those receiving vehicle media. In conclusion, we demonstrate for the first time that the administration of EPC-secreted factors could become a safe and effective cell-free option to be considered in future therapeutic strategies for stroke.

Longitudinal diffusion tensor imaging in a rat brain glioma model

In order to investigate the properties of water motion within and around brain tumors as a function of tumor growth, longitudinal diffusion tensor imaging (DTI) was carried out in a rat brain glioma (C6) model. As tumors grew in size, significant anisotropy of water diffusion was seen both within and around the tumor. The tissue water surrounding the tumor exhibited high planar anisotropy, as opposed to the linear anisotropy normally seen in white matter, indicating that cells were experiencing stress in a direction normal to the tumor border. When tumors were sufficiently large, significant anisotropy was also seen within the tumor because of longer‐range organization of cancer cells within the tumor borders. These findings have important implications for diffusion‐weighted MRI experiments examining tumor growth and response to therapy. Copyright

1H-MRSI pattern perturbation in a mouse glioma: the effects of acute hyperglycemia and moderate hypothermia.

MR spectroscopic Imaging (MRSI), with PRESS localization, is used here to monitor the effects of acute hyperglycemia in the spectral pattern of 11 mice bearing GL261 gliomas at normothermia (36.5–37.5°C) and at hypothermia (28.5–29.5°C). These in vivo studies were complemented by ex vivo high resolution magic angle spinning (HR‐MAS) analysis of GL261 tumor samples from 6 animals sacrificed by focused microwave irradiation, and blood glucose measurements in 12 control mice. Apparent glucose levels, monitored by in vivo MRSI in brain tumors during acute hyperglycemia, rose to an average of 1.6‐fold during hypothermia (p < 0.05), while no significant changes were detected at normothermia, or in control experiments performed at euglycemia, or in normal/peritumoral brain regions. Ex vivo analysis of glioma‐bearing mouse brains at hypothermia revealed higher glucose increases in distinct regions during the acute hyperglycemic challenge (up to 6.6‐fold at the tumor center), in agreement with maximal in vivo blood glucose changes (5‐fold). Phantom studies on taurine plus glucose containing solutions explained the differences between in vivo and ex vivo measurements. Our results also indicate brain tumor heterogeneity in the four animal tumors investigated in response to a defined metabolic challenge. Copyright

MRI detects therapeutic effects in weanling Niemann-Pick type C mice

To noninvasively evaluate the early effects of Niemann‐Pick type C (NPC) disease, diffusion tensor imaging (DTI) was carried out in the brains of very young (23‐day‐old) mice. The diffusion of water in white matter tracts of Npc1–/– mice at this young age was already abnormal, exhibiting decreased anisotropy, as quantified by fractional anisotropy (FA), compared with their wild‐type littermates, the controls. Postmortem histological staining revealed myelin deficiencies in Npc1–/– mice, consistent with the reduction in FA measured in vivo. Beneficial effects of treatment with allopregnanolone and/or 2 hydroxypropyl‐beta‐cyclodextrin was also detectable at this age by FA, which correlated with increased myelination as seen by histology. This is the earliest detection of a therapeutic effect in Npc1–/– mice.

Uncovering of intracellular water in cultured cells

The complexity of biologic tissues, with multiple compartments each with its own diffusion and relaxation properties, requires complex formalisms to model water signal in most magnetic resonance imaging or magnetic resonance spectroscopy experiments. In this article, we describe a magnetic susceptibility‐induced shift in the resonance frequency of extracellular water by the introduction of a gadolinium contrast agent to medium perfusing a hollow fiber bioreactor. The frequency shift of the extracellular water (+185 Hz at 9.4 T) uncovers the intracellular water and allows direct measurement of motional and relaxation properties of the intracellular space. The proposed method provides a unique tool for understanding the mechanisms underlining diffusion and relaxation in the intracellular space. Magn Reson Med 54:79–86, 2005.

Brain magnetic resonance in experimental acute-on-chronic liver failure.

Acute‐on‐chronic liver failure is the term that refers to sustained liver injury with acute decompensation, usually induced by a precipitating factor. A common link between ensuing failures of various organs is impairment of the vascular supply, which may also induce vasogenic oedema in the brain. The aim of this study was to perform magnetic resonance (MR) study of the brain in a rat model combining bile duct ligation (BDL) and lipopolysaccharide (LPS) administration to investigate brain oedema in liver failure.

Solid-State Deuterium NMR Spectroscopy of Membranes

Michael F. Brown1,2, Silvia Lope-Piedrafita3, Gary V. Martinez1, and Horia I. Petrache4 1Department of Chemistry, University of Arizona, Tucson, AZ 85721, USA; 2Department of Physics, University of Arizona, Tucson, AZ 85721, USA; 3Department of Radiology, University of Arizona, Tucson, AZ 85724, USA; and 4Laboratory of Physical and Structural Biology, National Institutes of Health, NICHD, Bethesda, MD 20892, USA

A new ex vivo method to evaluate the performance of candidate MRI contrast agents: a proof-of-concept study

BackgroundMagnetic resonance imaging (MRI) plays an important role in tumor detection/diagnosis. The use of exogenous contrast agents (CAs) helps to improve the discrimination between lesion and neighbouring tissue, but most of the currently available CAs are non-specific. Assessing the performance of new, selective CAs requires exhaustive assays and large amounts of material. Accordingly, in a preliminary screening of new CAs, it is important to choose candidate compounds with good potential for in vivo efficiency. This screening method should reproduce as close as possible the in vivo environment. In this sense, a fast and reliable method to select the best candidate CAs for in vivo studies would minimize time and investment cost, and would benefit the development of better CAs.ResultsThe post-mortem ex vivo relative contrast enhancement (RCE) was evaluated as a method to screen different types of CAs, including paramagnetic and superparamagnetic agents. In detail, sugar/gadolinium-loaded gold nanoparticles (Gd-GNPs) and iron nanoparticles (SPIONs) were tested. Our results indicate that the post-mortem ex vivo RCE of evaluated CAs, did not correlate well with their respective in vitro relaxivities. The results obtained with different Gd-GNPs suggest that the linker length of the sugar conjugate could modulate the interactions with cellular receptors and therefore the relaxivity value. A paramagnetic CA (GNP (E_2)), which performed best among a series of Gd-GNPs, was evaluated both ex vivo and in vivo. The ex vivo RCE was slightly worst than gadoterate meglumine (201.9 ± 9.3% versus 237 ± 14%, respectively), while the in vivo RCE, measured at the time-to-maximum enhancement for both compounds, pointed to GNP E_2 being a better CA in vivo than gadoterate meglumine. This is suggested to be related to the nanoparticule characteristics of the evaluated GNP.ConclusionWe have developed a simple, cost-effective relatively high-throughput method for selecting CAs for in vivo experiments. This method requires approximately 800 times less quantity of material than the amount used for in vivo administrations.

In Vivo and Ex Vivo Magnetic Resonance Spectroscopy of the Infarct and the Subventricular Zone in Experimental Stroke

Ex vivo high-resolution magic-angle spinning (HRMAS) provides metabolic information with higher sensitivity and spectral resolution than in vivo magnetic resonance spectroscopy (MRS). Therefore, we used both techniques to better characterize the metabolic pattern of the infarct and the neural progenitor cells (NPCs) in the ipsilateral subventricular zone (SVZi). Ischemic stroke rats were divided into three groups: G0 (non-stroke controls, n = 6), G1 (day 1 after stroke, n = 6), and G7 (days 6 to 8 after stroke, n =12). All the rats underwent MRS. Three rats per group were analyzed by HRMAS. The remaining rats were used for immunohistochemical studies. In the infarct, both techniques detected significant metabolic changes. The most relevant change was in mobile lipids (2.80 ppm) in the G7 group (a 5.53- and a 3.95-fold increase by MRS and HRMAS, respectively). In the SVZi, MRS did not detect any significant metabolic change. However, HRMAS detected a 2.70-fold increase in lactate and a 0.68-fold decrease in N-acetylaspartate in the G1 group. None of the metabolites correlated with the 1.37-fold increase in NPCs detected by immunohistochemistry in the G7 group. In conclusion, HRMAS improves the metabolic characterization of the brain in experimental ischemic stroke. However, none of the metabolites qualifies as a surrogate biomarker of NPCs.