Dual contrast CEST MRI for pH-weighted imaging in stroke.

PURPOSE: pH enhanced (pHenh ) CEST imaging combines the pH sensitivity from amide and guanidino signals, but the saturation parameters have not been optimized. We propose pHdual as a variant of pHenh that suppresses background signal variations, while enhancing pH sensitivity and potential for imaging ischemic brain injury of stroke. METHODS: Simulation and in vivo rodent stroke experiments of pHenh MRI were performed with varied RF saturation powers for both amide and guanidino protons to optimize the contrast between lesion/normal tissues, while simultaneously minimizing signal variations across different types of normal tissues. In acute stroke, contrast and volume ratio measured by pHdual imaging were compared with an amide-CEST approach, and perfusion and diffusion MRI. RESULTS: Simulation experiments indicated that amide and guanidino CEST signals exhibit unique sensitivities across different pH ranges, with pHenh producing greater sensitivity over a broader pH regime. The pHenh data of rodent stroke brain demonstrated that the lesion/normal tissue contrast was maximized for an RF saturation power pair of 0.5 µT at 2.0 ppm and 1.0 µT at 3.6 ppm, whereas an optimal contrast-to-variation ratio (CVR) was obtained with a 0.7 µT saturation at 2.0 ppm and 0.8 µT at 3.6 ppm. In acute stroke, CVR optimized pHenh (i.e., pHdual ) achieved a higher sensitivity than the three-point amide-CEST approach, and distinct patterns of lesion tissue compared to diffusion and perfusion MRI. CONCLUSION: pHdual MRI improves the sensitivity of pH-weighted imaging and will be a valuable tool for assessing tissue viability in stroke.

Mapping mesoscale connectivity within the human hippocampus.

The connectivity of the hippocampus is essential to its functions. To gain a whole system view of intrahippocampal connectivity, ex vivo mesoscale (100 µm isotropic resolution) multi-shell diffusion MRI (11.7T) and tractography were performed on entire post-mortem human right hippocampi. Volumetric measurements indicated that the head region was largest followed by the body and tail regions. A unique anatomical organization in the head region reflected a complex organization of the granule cell layer (GCL) of the dentate gyrus. Tractography revealed the volumetric distribution of the perforant path, including both the tri-synaptic and temporoammonic pathways, as well as other well-established canonical connections, such as Schaffer collaterals. Visualization of the perforant path provided a means to verify the borders between the pro-subiculum and CA1, as well as between CA1/CA2. A specific angularity of different layers of fibers in the alveus was evident across the whole sample and allowed a separation of afferent and efferent connections based on their origin (i.e. entorhinal cortex) or destination (i.e. fimbria) using a cluster analysis of streamlines. Non-canonical translamellar connections running along the anterior-posterior axis were also discerned in the hilus. In line with "dentations" of the GCL, mossy fibers were bunching together in the sagittal plane revealing a unique lamellar organization and connections between these. In the head region, mossy fibers projected to the origin of the fimbria, which was distinct from the body and tail region. Mesoscale tractography provides an unprecedented systems view of intrahippocampal connections that underpin cognitive and emotional processing.

Post-Stroke Timing of ECM Hydrogel Implantation Affects Biodegradation and Tissue Restoration.

Extracellular matrix (ECM) hydrogel promotes tissue regeneration in lesion cavities after stroke. However, a bioscaffold's regenerative potential needs to be considered in the context of the evolving pathological environment caused by a stroke. To evaluate this key issue in rats, ECM hydrogel was delivered to the lesion core/cavity at 7-, 14-, 28-, and 90-days post-stroke. Due to a lack of tissue cavitation 7-days post-stroke, implantation of ECM hydrogel did not achieve a sufficient volume and distribution to warrant comparison with the other time points. Biodegradation of ECM hydrogel implanted 14- and 28-days post-stroke were efficiently (80%) degraded by 14-days post-bioscaffold implantation, whereas implantation 90-days post-stroke revealed only a 60% decrease. Macrophage invasion was robust at 14- and 28-days post-stroke but reduced in the 90-days post-stroke condition. The pro-inflammation (M1) and pro-repair (M2) phenotype ratios were equivalent at all time points, suggesting that the pathological environment determines macrophage invasion, whereas ECM hydrogel defines their polarization. Neural cells (neural progenitors, neurons, astrocytes, oligodendrocytes) were found at all time points, but a 90-days post-stroke implantation resulted in reduced densities of mature phenotypes. Brain tissue restoration is therefore dependent on an efficient delivery of a bioscaffold to a tissue cavity, with 28-days post-stroke producing the most efficient biodegradation and tissue regeneration, whereas by 90-days post-stroke, these effects are significantly reduced. Improving our understanding of how the pathological environment influences biodegradation and the tissue restoration process is hence essential to devise engineering strategies that could extend the therapeutic window for bioscaffolds to repair the damaged brain.

Ex vivo mesoscopic diffusion MRI correlates with seizure frequency in patients with uncontrolled mesial temporal lobe epilepsy.

The role of hippocampal connectivity in mesial temporal lobe epilepsy (mTLE) remains poorly understood. The use of ex vivo hippocampal samples excised from patients with mTLE affords mesoscale diffusion magnetic resonance imaging (MRI) to identify individual cell layers, such as the pyramidal (PCL) and granule cell layers (GCL), which are thought to be impacted by seizure activity. Diffusion tensor imaging (DTI) of control (n = 3) and mTLE (n = 7) hippocampi on an 11.7 T MRI scanner allowed us to reveal intra-hippocampal connectivity and evaluate how epilepsy affected mean (MD), axial (AD), and radial diffusivity (RD), as well as fractional anisotropy (FA). Regional measurements indicated a volume loss in the PCL of the cornu ammonis (CA) 1 subfield in mTLE patients compared to controls, which provided anatomical context. Diffusion measurements, as well as streamline density, were generally higher in mTLE patients compared to controls, potentially reflecting differences due to tissue fixation. mTLE measurements were more variable than controls. This variability was associated with disease severity, as indicated by a strong correlation (r = 0.87) between FA in the stratum radiatum and the frequency of seizures in patients. MD and RD of the PCL in subfields CA3 and CA4 also correlated strongly with disease severity. No correlation of MR measures with disease duration was evident. These results reveal the potential of mesoscale diffusion MRI to examine layer-specific diffusion changes and connectivity to determine how these relate to clinical measures. Improving the visualization of intra-hippocampal connectivity will advance the development of novel hypotheses about seizure networks.

Mesoscale diffusion magnetic resonance imaging of the ex vivo human hippocampus.

Mesoscale diffusion magnetic resonance imaging (MRI) endeavors to bridge the gap between macroscopic white matter tractography and microscopic studies investigating the cytoarchitecture of human brain tissue. To ensure a robust measurement of diffusion at the mesoscale, acquisition parameters were arrayed to investigate their effects on scalar indices (mean, radial, axial diffusivity, and fractional anisotropy) and streamlines (i.e., graphical representation of axonal tracts) in hippocampal layers. A mesoscale resolution afforded segementation of the pyramidal cell layer (CA1-4), the dentate gyrus, as well as stratum moleculare, radiatum, and oriens. Using ex vivo samples, surgically excised from patients with intractable epilepsy (n = 3), we found that shorter diffusion times (23.7 ms) with a b-value of 4,000 s/mm2 were advantageous at the mesoscale, providing a compromise between mean diffusivity and fractional anisotropy measurements. Spatial resolution and sample orientation exerted a major effect on tractography, whereas the number of diffusion gradient encoding directions minimally affected scalar indices and streamline density. A sample temperature of 15°C provided a compromise between increasing signal-to-noise ratio and increasing the diffusion properties of the tissue. Optimization of the acquisition afforded a system's view of intra- and extra-hippocampal connections. Tractography reflected histological boundaries of hippocampal layers. Individual layer connectivity was visualized, as well as streamlines emanating from individual sub-fields. The perforant path, subiculum and angular bundle demonstrated extra-hippocampal connections. Histology of the samples confirmed individual cell layers corresponding to ROIs defined on MR images. We anticipate that this ex vivo mesoscale imaging will yield novel insights into human hippocampal connectivity.

A systematic optimization of 19F MR image acquisition to detect macrophage invasion into an ECM hydrogel implanted in the stroke-damaged brain.

19F-MR imaging of perfluorocarbon (PFC)-labeled macrophages can provide a unique insight into their participation and spatio-temporal dynamics of inflammatory events, such as the biodegradation of an extracellular matrix (ECM) hydrogel implanted into a stroke cavity. To determine the most efficient acquisition strategy for 19F-MR imaging, five commonly used sequences were optimized using a design of experiment (DoE) approach and compared based on their signal-to-noise ratio (SNR). The fast imaging with steady-state precession (FISP) sequence produced the most efficient detection of a 19F signal followed by the rapid acquisition with relaxation enhancement (RARE) sequence. The multi-slice multi-echo (MSME), fast low angle shot (FLASH), and zero echo time (ZTE) sequences were significantly less efficient. Imaging parameters (matrix/voxel size; slice thickness, number of averages) determined the accuracy (i.e. trueness and precision) of object identification by reducing partial volume effects, as determined by analysis of the point spread function (PSF). A 96 × 96 matrix size (0.35 mm3) produced the lowest limit of detection (LOD) for RARE (2.85 mM PFPE; 119 mM 19F) and FISP (0.43 mM PFPE; 18.1 mM 19F), with an SNR of 2 as the detection threshold. Imaging of a brain phantom with PFC-labeled macrophages invading an ECM hydrogel further illustrated the impact of these parameter changes. The systematic optimization of sequence and imaging parameters provides the framework for an accurate visualization of 19F-labeled macrophage distribution and density in the brain. This will enhance our understanding of the contribution of periphery-derived macrophages in bioscaffold degradation and its role in brain tissue regeneration.

Chemical exchange-sensitive spin-lock (CESL) MRI of glucose and analogs in brain tumors.

PURPOSE: Glucose uptake and metabolism can be measured by chemical exchange-sensitive spin-lock (CESL) MRI with an administration of glucose or its analogs. This study investigates the sensitivity, the spatiotemporal characteristics, and the signal source of glucoCESL with a 9L rat brain tumor model. METHODS: Dynamic CESL MRI with intravenous injection of D-glucose, 2-deoxy-D-glucose (2DG), and L-glucose were measured and compared with gadolinium-based dynamic contrast-enhanced (DCE) MRI. RESULTS: The CESL signals with an injection of glucose or analogs have faster and larger changes in tumors than normal brain tissue. In tumors, the CESL signal with 2DG injection has larger and slower peak response than that with D-glucose due to the accumulation of 2DG and 2DG-6-phosphate in the intracellular compartment, whereas L-glucose, which cannot be transported intracellularly by glucose transporters, only induces a small change. The initial glucoCESL maps (< 4 minutes) are qualitatively similar to DCE maps, whereas later maps (> 4 minutes) show more widespread responses. The rise times of D-glucose-CESL and 2DG-CESL signals in the tumor are slower than that of DCE. Our data suggest that the initial CESL contrast primarily reflects a passive increase of glucose content in the extracellular space of tumors due to a higher vascular permeability, whereas the later period may have a significant contribution from the uptake/metabolism of glucose in the intracellular compartment. CONCLUSIONS: Our results demonstrate that glucoCESL MRI has both extracellular and intracellular contributions, and can be a useful tool for measurements of both vascular permeability and glucose uptake in tumors.

Detection of aberrant hippocampal mossy fiber connections: Ex vivo mesoscale diffusion MRI and microtractography with histological validation in a patient with uncontrolled temporal lobe epilepsy.

Understanding the neurobiology and functional connectivity of hippocampal structures is essential for improving the treatment of mesial temporal lobe epilepsy. At the macroscale, in vivo MRI often reveals hippocampal atrophy and decreased fractional anisotropy, whereas at the microscopic scale, there frequently is evidence of neuronal loss and gliosis. Mossy fiber sprouting in the dentate gyrus (DG), with evidence of glutamatergic synapses in the stratum moleculare (SM) putatively originating from granule cell neurons, may also be observed. This aberrant connection between the DG and SM could produce a reverberant excitatory circuit. However, this hypothesis cannot easily be evaluated using macroscopic or microscopic techniques. We here demonstrate that the ex vivo mesoscopic MRI of surgically excised hippocampi can bridge the explanatory and analytical gap between the macro- and microscopic scale. Specifically, diffusion- and T2 -weighted MRI can be integrated to visualize a cytoarchitecture that is akin to immunohistochemistry. An appropriate spatial resolution to discern individual cell layers can then be established. Processing of diffusion tensor images using tractography detects extra- and intrahippocampal connections, hence providing a unique systems view of the hippocampus and its connected regions. Here, this approach suggests that there is indeed an aberrant connection between the DG and SM, supporting the sprouting hypothesis of a reverberant excitatory network. Mesoscopic ex vivo MR imaging hence provides an exciting new avenue to study hippocampi from treatment-resistant patients and allows exploration of existing hypotheses, as well as the development of new treatment strategies based on these novel insights. Hum Brain Mapp 37:780-795, 2016. © 2015 Wiley Periodicals, Inc.

Brain plasticity and recovery in preclinical models of stroke.

Post-stroke recovery relies on neurobiological changes that modify the organization and function of the brain under pathophysiological conditions. The changes can be adaptive (i.e. restoration of function) or maladaptive (i.e. worsening of function). Preclinical models of stroke exhibit adaptive plasticity that leads to a "spontaneous recov- ery" of functions. This recovery can be modulated through external factors, such as rehabilitation, pharmacology or other adjuvant strategies. Nevertheless, current interventions only result in a limited improvement of deficits and there is also potential for maladaptation. Hence, a better understanding of the mechanisms underlying recovery is essential for the design of more efficient and targeted treatment strategies. Here, we review the main features of adaptive plasticity that are thought to underlie the spontaneous and induced recovery of function in animal models of stroke. Within this context, therapeutic interventions, used in isolation or synergistically to modulate recovery, are discussed. It is hoped that a focus on neurobiological principles and their manipulation will enhance interven- tional strategies to maximize therapeutic benefit. To ensure translation of these interventions into a clinical setting, a close interaction between basic and applied research is required.

In vitro dose-dependent effects of matrix metalloproteinases on ECM hydrogel biodegradation.

Matrix metalloproteinases (MMPs) cause proteolysis of extracellular matrix (ECM) in tissues affected by stroke. However, little is known about how MMPs degrade ECM hydrogels implanted into stroke cavities to regenerate lost tissue. To establish a structure-function relationship between different doses of individual MMPs and isolate their effects in a controlled setting, an in vitro degradation assay quantified retained urinary bladder matrix (UBM) hydrogel mass as a measure of degradation across time. A rheological characterization indicated that lower ECM concentrations (<4 mg/mL) did not cure completely at 37 °C and had a high fraction of mobile proteins that were easily washed-out. Hydrolysis by dH2O caused a steady 2 % daily decrease in hydrogel mass over 14 days. An acceleration of degradation to 6 % occurred with phosphate buffered saline and artificial cerebrospinal fluid. MMPs induced a dose-dependent increase and within 14 days almost completely (>95 %) degraded the hydrogel. MMP-9 exerted the most significant biodegradation, compared to MMP-3 and -2. To model the in vivo exposure of hydrogel to MMPs, mixtures of MMP-2, -3, and -9, present in the cavity at 14-, 28-, or 90-days post-stroke, revealed that 14- and 28-days mixtures achieved an equivalent biodegradation, but a 90-days mixture exhibited a slower degradation. These results revealed that hydrolysis, in addition to proteolysis, exerts a major influence on the degradation of hydrogels. Understanding the mechanisms of ECM hydrogel biodegradation is essential to determine the therapeutic window for bioscaffold implantation after a stroke, and they are also key to determine optimal degradation kinetics to support tissue regeneration. STATEMENT OF SIGNIFICANCE: After implantation into a stroke cavity, extracellular matrix (ECM) hydrogel promotes tissue regeneration through the degradation of the bioscaffold. However, the process of degradation of an ECM hydrogel remains poorly understood. We here demonstrated in vitro under highly controlled conditions that hydrogel degradation is very dependent on its protein concentration. Lower protein concentration hydrogels were weaker in rheological measurements and particularly susceptible to hydrolysis. The proteolytic degradation of tissue ECM after a stroke is caused by matrix metalloproteinases (MMPs). A dose-dependent MMP-driven biodegradation of ECM hydrogel exceeded the effects of hydrolysis. These results highlight the importance of in vitro testing of putative causes of degradation to gain a better understanding of how these factors affect in vivo biodegradation.

Bioengineering solutions for neural repair and recovery in stroke.

PURPOSE OF REVIEW: This review discusses emerging bioengineering opportunities for the treatment of stroke and their potential to build on current rehabilitation protocols. RECENT FINDINGS: Bioengineering is a vast field that ranges from biomaterials to brain-computer interfaces. Biomaterials find application in the delivery of pharmacotherapies, as well as the emerging field of tissue engineering. For the treatment of stroke, these approaches have to be seen in the context of physical therapy in order to maximize functional outcomes. There is also an emergence of rehabilitation that engages engineering solutions, such as robot-assisted training, as well as brain-computer interfaces that can potentially assist in the case of paralysis. SUMMARY: Stroke remains the main cause of adult disability with rehabilitation therapy being the focus for chronic impairments. Bioengineering is offering new opportunities to both support and synergize with currently available treatment options, and also promises to potentially dramatically improve available approaches. VIDEO ABSTRACT AVAILABLE: See the Video Supplementary Digital Content 1 (http://links.lww.com/CONR/A21).

Implantation site and lesion topology determine efficacy of a human neural stem cell line in a rat model of chronic stroke.

Stroke remains one of the most promising targets for cell therapy. Thorough preclinical efficacy testing of human neural stem cell (hNSC) lines in a rat model of stroke (transient middle cerebral artery occlusion) is, however, required for translation into a clinical setting. Magnetic resonance imaging (MRI) here confirmed stroke damage and allowed the targeted injection of 450,000 hNSCs (CTX0E03) into peri-infarct tissue, rather than the lesion cyst. Intraparenchymal cell implants improved sensorimotor dysfunctions (bilateral asymmetry test) and motor deficits (footfault test and rotameter). Importantly, analyses based on lesion topology (striatal vs. striatal + cortical damage) revealed a more significant improvement in animals with a stroke confined to the striatum. However, no improvement in learning and memory (water maze) was evident. An intracerebroventricular injection of cells did not result in any improvement. MRI-based lesion, striatal and cortical volumes were unchanged in treated animals compared to those with stroke that received an intraparenchymal injection of suspension vehicle. Grafted cells only survived after intraparenchymal injection with a striatal + cortical topology resulting in better graft survival (16,026 cells) than in animals with smaller striatal lesions (2,374 cells). Almost 20% of cells differentiated into glial fibrillary acidic protein+ astrocytes, but <2% turned into FOX3+ neurons. These results indicate that CTX0E03 implants robustly recover behavioral dysfunction over a 3-month time frame and that this effect is specific to their site of implantation. Lesion topology is potentially an important factor in the recovery, with a stroke confined to the striatum showing a better outcome compared to a larger area of damage.

MR diffusion histology and micro-tractography reveal mesoscale features of the human cerebellum.

After 140 years from the discovery of Golgi's black reaction, the study of connectivity of the cerebellum remains a fascinating yet challenging task. Current histological techniques provide powerful methods for unravelling local axonal architecture, but the relatively low volume of data that can be acquired in a reasonable amount of time limits their application to small samples. State-of-the-art in vivo magnetic resonance imaging (MRI) methods, such as diffusion tractography techniques, can reveal trajectories of the major white matter pathways, but their correspondence with underlying anatomy is yet to be established. Hence, a significant gap exists between these two approaches as neither of them can adequately describe the three-dimensional complexity of fibre architecture at the level of the mesoscale (from a few millimetres to micrometres). In this study, we report the application of MR diffusion histology and micro-tractography methods to reveal the combined cytoarchitectural organisation and connectivity of the human cerebellum at a resolution of 100-µm (2 nl/voxel volume). Results show that the diffusion characteristics for each layer of the cerebellar cortex correctly reflect the known cellular composition and its architectural pattern. Micro-tractography also reveals details of the axonal connectivity of individual cerebellar folia and the intra-cortical organisation of the different cerebellar layers. The direct correspondence between MR diffusion histology and micro-tractography with immunohistochemistry indicates that these approaches have the potential to complement traditional histology techniques by providing a non-destructive, quantitative and three-dimensional description of the microstructural organisation of the healthy and pathological tissue.

Acquisition and Analysis of Excised Neocortex from Pediatric Patients with Focal Cortical Dysplasia Using Mesoscale Diffusion MRI.

Non-invasive classification of focal cortical dysplasia (FCD) subtypes remains challenging from a radiology perspective. Quantitative imaging biomarkers (QIBs) have the potential to distinguish subtypes that lack pathognomonic features and might help in defining the extent of abnormal connectivity associated with each FCD subtype. A key motivation of diagnostic imaging is to improve the localization of a "lesion" that can guide the surgical resection of affected tissue, which is thought to cause seizures. Conversely, surgical resections to eliminate or reduce seizures provided unique opportunities to develop magnetic resonance imaging (MRI)-based QIBs by affording long scan times to evaluate multiple contrast mechanisms at the mesoscale (0.5 mm isotropic voxel dimensions). Using ex vivo hybrid diffusion tensor imaging on a 9.4 T MRI scanner, the grey to white matter ratio of scalar indices was lower in the resected middle temporal gyrus (MTG) of two neuropathologically confirmed cases of FCD compared to non-diseased control postmortem fixed temporal lobes. In contrast, fractional anisotropy was increased within FCD and also adjacent white matter tracts. Connectivity (streamlines/mm3) in the MTG was higher in FCD, suggesting that an altered connectivity at the lesion locus can potentially provide a tangible QIB to distinguish and characterize FCD abnormalities. However, as illustrated here, a major challenge for a robust tractographical comparison lies in the considerable differences in the ex vivo processing of bioptic and postmortem samples. Mesoscale diffusion MRI has the potential to better define and characterize epileptic tissues obtained from surgical resection to advance our understanding of disease etiology and treatment.

Ex vivo 100 µm isotropic diffusion MRI-based tractography of connectivity changes in the end-stage R6/2 mouse model of Huntington's disease.

Background: Huntington's disease is a progressive neurodegenerative disorder. Brain atrophy, as measured by volumetric magnetic resonance imaging (MRI), is a downstream consequence of neurodegeneration, but microstructural changes within brain tissue are expected to precede this volumetric decline. The tissue microstructure can be assayed non-invasively using diffusion MRI, which also allows a tractographic analysis of brain connectivity. Methods: We here used ex vivo diffusion MRI (11.7 T) to measure microstructural changes in different brain regions of end-stage (14 weeks of age) wild type and R6/2 mice (male and female) modeling Huntington's disease. To probe the microstructure of different brain regions, reduce partial volume effects and measure connectivity between different regions, a 100 µm isotropic voxel resolution was acquired. Results: Although fractional anisotropy did not reveal any difference between wild-type controls and R6/2 mice, mean, axial, and radial diffusivity were increased in female R6/2 mice and decreased in male R6/2 mice. Whole brain streamlines were only reduced in male R6/2 mice, but streamline density was increased. Region-to-region tractography indicated reductions in connectivity between the cortex, hippocampus, and thalamus with the striatum, as well as within the basal ganglia (striatum-globus pallidus-subthalamic nucleus-substantia nigra-thalamus). Conclusions: Biological sex and left/right hemisphere affected tractographic results, potentially reflecting different stages of disease progression. This proof-of-principle study indicates that diffusion MRI and tractography potentially provide novel biomarkers that connect volumetric changes across different brain regions. In a translation setting, these measurements constitute a novel tool to assess the therapeutic impact of interventions such as neuroprotective agents in transgenic models, as well as patients with Huntington's disease.

Implantation of undifferentiated and pre-differentiated human neural stem cells in the R6/2 transgenic mouse model of Huntington's disease.

BACKGROUND: Cell therapy is a potential therapeutic approach for several neurodegenetative disease, including Huntington Disease (HD). To evaluate the putative efficacy of cell therapy in HD, most studies have used excitotoxic animal models with only a few studies having been conducted in genetic animal models. Genetically modified animals should provide a more accurate representation of human HD, as they emulate the genetic basis of its etiology. RESULTS: In this study, we aimed to assess the therapeutic potential of a human striatal neural stem cell line (STROC05) implanted in the R6/2 transgenic mouse model of HD. As DARPP-32 GABAergic output neurons are predominately lost in HD, STROC05 cells were also pre-differentiated using purmorphamine, a hedgehog agonist, to yield a greater number of DARPP-32 cells. A bilateral injection of 4.5x105 cells of either undifferentiated or pre-differentiated DARPP-32 cells, however, did not affect outcome compared to a vehicle control injection. Both survival and neuronal differentiation remained poor with a mean of only 161 and 81 cells surviving in the undifferentiated and differentiated conditions respectively. Only a few cells expressed the neuronal marker Fox3. CONCLUSIONS: Although the rapid brain atrophy and short life-span of the R6/2 model constitute adverse conditions to detect potentially delayed treatment effects, significant technical hurdles, such as poor cell survival and differentiation, were also sub-optimal. Further consideration of these aspects is therefore needed in more enduring transgenic HD models to provide a definite assessment of this cell line's therapeutic relevance. However, a combination of treatments is likely needed to affect outcome in transgenic models of HD.

Magnetic resonance imaging of pathological processes in rodent models of amyotrophic lateral sclerosis.

Non-human models of neurodegenerative diseases have potential for the identification of key pathways in pathogenesis and for the more rapid assessment of therapeutic candidates. While there are legitimate concerns about the physiological differences between the rodent and human motor systems, mice expressing the 'G93A' superoxide dismutase-1 gene mutation are a predictable and robustly-characterized model for amyotrophic lateral sclerosis (ALS). This model has provided evidence for an important role of inflammatory processes during the pre-clinical phase, a stage currently inaccessible for human study in what is largely a sporadic disease. While magnetic resonance imaging is now an established and leading modality for the identification of ALS biomarkers in humans, it can also be increasingly applied to rodent models to probe structural, functional and biochemical changes throughout the course of the disease, with additional potential to generate surrogate markers for the efficacy of therapeutic interventions. Targeted MRI contrast agents, through tagging of various cell types and even individual molecules, will deliver an era of in vivo molecular neuroimaging, with greater specificity for the most relevant pathological processes. These are potentially important steps towards the ultimate goal of human therapeutic translation.

Targeting 17ß-estradiol biosynthesis in neural stem cells improves stroke outcome.

Dax-1 (dosage-sensitive sex reversal adrenal hypoplasia congenital region on X-chromosome gene 1) blocks 17ß-estradiol biosynthesis and its knockdown would be expected to increase 17ß-estradiol production. We hypothesized that knockdown of Dax-1 in a conditionally immortalized neural stem cell (NSC) line, MHP36, is a useful approach to increase 17ß-estradiol production. Short hairpin (sh) RNA targeted to Dax-1 in NSCs, namely MHP36-Dax1KD cells, resulted in the degradation of Dax-1 RNA and attenuation of Dax-1 protein expression. In vitro, MHP36-Dax1KD cells exhibited overexpression of aromatase and increased 17ß-estradiol secretion compared to MHP36 cells. As 17ß-estradiol has been shown to promote the efficacy of cell therapy, we interrogated the application of 17ß-estradiol-enriched NSCs in a relevant in vivo disease model. We hypothesized that MHP36-Dax1KD cells will enhance functional recovery after transplantation in a stroke model. C57BL/6 male adult mice underwent ischemia/reperfusion by left middle cerebral artery occlusion for 45 min using an intraluminal thread. Two days later male mice randomly received vehicle, MHP36 cells, MHP36-Dax1KD cells, and MHP36 cells suspended in 17ß-estradiol (100 nm) or 17ß-estradiol alone (100 nm) with serial behavioral testing over 28 days followed by post-mortem histology and blinded analysis. Recovery of sensorimotor function was accelerated and enhanced, and lesion volume was reduced by MHP36-Dax1KD transplants. Regarding mechanisms, immunofluorescence indicated increased synaptic plasticity and neuronal differentiation after MHP36-Dax1KD transplants. In conclusion, knockdown of Dax-1 is a useful target to increase 17ß-estradiol biosynthesis in NSCs and improves functional recovery after stroke in vivo, possibly mediated through neuroprotection and improved synaptic plasticity. Therefore, targeting 17ß-estradiol biosynthesis in stem cells may be a promising therapeutic strategy for enhancing the efficacy of stem cell-based therapies for stroke.