Hindshaker,a Novel Myelin Mutant Showing Hypomyelination Preferentially Affecting the Spinal Cord

Animals with spontaneous mutations affecting myelin formation have provided useful information about the genetic and cellular mechanisms regulating normal and abnormal myelination. In this paper we describe a novel murine mutation termed hindshaker (hsh) which is inherited in an automosal recessive manner. Affected mice are characterised by a variable tremor of the hind end which commences at about 2 weeks of age and largely disappears in animals older than 6 weeks. There is hypomyelination affecting predominantly the spinal cord, although the optic nerves and brain are involved to a much lesser degree. The defect of thinly myelinated and naked axons is maximal at 20 days of age and largely resolves with time so that in the adult most axons are myelinated. The myelin structure appears normal and immunostains for the major proteins. Although the distribution of oligodendrocytes in the spinal cord is similar to normal during the period of hypomyelination, there are fewer mature cells. The hsh mutation appears to delay the maturation of oligodenrocytes, particularly in the spinal cord. Additionally, there is a considerable variation in phenotypic expression and in penetrance when the mutation is expressed on different genetic backgrounds, suggesting the hsh locus is subject to the influence of modifying gene(s). Identification of the hsh gene should identify a factor important in the development of oligodendrocytes, particularly those in the spinal cord.     

Novel Role of HAX-1 in Neurons Protection After Spinal Cord Injury Involvement of IRE-1

Neurochemical Research - Spinal cord injury (SCI) is one of the diseases with high probability of causing disability in human beings, and there is no reliable treatment at present. Neuronal...     

Retinoic Acid Induced-Autophagic Flux Inhibits ER-Stress Dependent Apoptosis and Prevents Disruption of Blood-Spinal Cord Barrier after Spinal Cord Injury

Spinal cord injury (SCI) induces the disruption of the blood-spinal cord barrier (BSCB) which leads to infiltration of blood cells, an inflammatory response, and neuronal cell death, resulting spinal cord secondary damage. Retinoic acid (RA) has a neuroprotective effect in both ischemic brain injury and SCI, however the relationship between BSCB disruption and RA in SCI is still unclear. In this study, we demonstrated that autophagy and ER stress are involved in the protective effect of RA on the BSCB. RA attenuated BSCB permeability and decreased the loss of tight junction (TJ) molecules such as P120, β-catenin, Occludin and Claudin5 after injury in vivo as well as in Brain Microvascular Endothelial Cells (BMECs). Moreover, RA administration improved functional recovery in the rat model of SCI. RA inhibited the expression of CHOP and caspase-12 by induction of autophagic flux. However, RA had no significant effect on protein expression of GRP78 and PDI. Furthermore, combining RA with the autophagy inhibitor chloroquine (CQ) partially abolished its protective effect on the BSCB via exacerbated ER stress and subsequent loss of tight junctions. Taken together, the neuroprotective role of RA in recovery from SCI is related to prevention of of BSCB disruption via the activation of autophagic flux and the inhibition of ER stress-induced cell apoptosis. These findings lay the groundwork for future translational studies of RA for CNS diseases, especially those related to BSCB disruption.     

The Effects of the Olig Family on the Regulation of Spinal Cord Development and Regeneration

Neurochemical Research - Neurons and glial cells in the central nervous system (CNS) are generated from neuroepithelial cells in the ventricular zone that surrounds the embryonic neural tube. The...     

Toward a Broader View of Ube3a in a Mouse Model of Angelman Syndrome: Expression in Brain,Spinal Cord,Sciatic Nerve and Glial Cells

Angelman Syndrome (AS) is a devastating neurodevelopmental disorder characterized by developmental delay, speech impairment, movement disorder, sleep disorders and refractory epilepsy. AS is caused by loss of the Ube3a protein encoded for by the imprinted Ube3a gene. Ube3a is expressed nearly exclusively from the maternal chromosome in mature neurons. While imprinting in neurons of the brain has been well described, the imprinting and expression of Ube3a in other neural tissues remains relatively unexplored. Moreover, given the overwhelming deficits in brain function in AS patients, the possibility of disrupted Ube3a expression in the infratentorial nervous system and its consequent disability have been largely ignored. We evaluated the imprinting status of Ube3a in the spinal cord and sciatic nerve and show that it is also imprinted in these neural tissues. Furthermore, a growing body of clinical and radiological evidence has suggested that myelin dysfunction may contribute to morbidity in many neurodevelopmental syndromes. However, findings regarding Ube3a expression in non-neuronal cells of the brain have varied. Utilizing enriched primary cultures of oligodendrocytes and astrocytes, we show that Ube3a is expressed, but not imprinted in these cell types. Unlike many other neurodevelopmental disorders, AS symptoms do not become apparent until roughly 6 to 12 months of age. To determine the temporal expression pattern and silencing, we analyzed Ube3a expression in AS mice at several time points. We confirm relaxed imprinting of Ube3a in neurons of the postnatal developing cortex, but not in structures in which neurogenesis and migration are more complete. This furthers the hypothesis that the apparently normal window of development in AS patients is supported by an incompletely silenced paternal allele in developing neurons, resulting in a relative preservation of Ube3a expression during this crucial epoch of early development.     

Longitudinal Evaluation of Mouse Hind Limb Bone Loss After Spinal Cord Injury using Novel,in vivo,Methodology

Spinal cord injury (SCI) is often accompanied by osteoporosis in the sublesional regions of the pelvis and lower extremities, leading to a higher frequency of fractures ¹. As these fractures often occur in regions that have lost normal sensory function, the patient is at a greater risk of fracture-dependent pathologies, including death. SCI-dependent loss in both bone mineral density (BMD, grams/cm²) and bone mineral content (BMC, grams) has been attributed to mechanical disuse ², aberrant neuronal signaling ³ and hormonal changes ⁴. The use of rodent models of SCI-induced osteoporosis can provide invaluable information regarding the mechanisms underlying the development of osteoporosis following SCI as well as a test environment for the generation of new therapies ^5-7 (and reviewed in ⁸). Mouse models of SCI are of great interest as they permit a reductionist approach to mechanism-based assessment through the use of null and transgenic mice. While such models have provided important data, there is still a need for minimally-invasive, reliable, reproducible, and quantifiable methods in determining the extent of bone loss following SCI, particularly over time and within the same cohort of experimental animals, to improve diagnosis, treatment methods, and/or prevention of SCI-induced osteoporosis.An ideal method for measuring bone density in rodents would allow multiple, sequential (over time) exposures to low-levels of X-ray radiation. This study describes the use of a new whole-animal scanner, the IVIS Lumina XR (Caliper Instruments) that can be used to provide low-energy (1-3 milligray (mGy)) high-resolution, high-magnification X-ray images of mouse hind limb bones over time following SCI. Significant bone density loss was seen in the tibiae of mice by 10 days post-spinal transection when compared to uninjured, age-matched control (naïve) mice (13% decrease, p<0.0005). Loss of bone density in the distal femur was also detectable by day 10 post-SCI, while a loss of density in the proximal femur was not detectable until 40 days post injury (7% decrease, p<0.05). SCI-dependent loss of mouse femur density was confirmed post-mortem through the use of Dual-energy X-ray Absorptiometry (DXA), the current gold standard for bone density measurements. We detect a 12% loss of BMC in the femurs of mice at 40 days post-SCI using the IVIS Lumina XR. This compares favorably with a previously reported BMC loss of 13.5% by Picard and colleagues who used DXA analysis on mouse femurs post-mortem 30 days post-SCI ⁹. Our results suggest that the IVIS Lumina XR provides a novel, high-resolution/high-magnification method for performing long-term, longitudinal measurements of hind limb bone density in the mouse following SCI.     

Expression Profile and Role of EphrinA1 Ligand After Spinal Cord Injury

Spinal cord injury (SCI) triggers the re-expression of inhibitory molecules present in early stages of development, contributing to prevention of axonal regeneration. Upregulation of EphA receptor tyrosine kinases after injury suggest their involvement in the nervous system’s response to damage. However, the expression profile of their ephrinA ligands after SCI is unclear. In this study, we determined the expression of ephrinA ligands after contusive SCI. Adult Sprague-Dawley female rats were injured using the MASCIS impactor device at the T10 vertebrae, and levels of ephrinA mRNA and protein determined at different time points. Identification of the cell phenotype expressing the ephrin ligand and colocalization with Eph receptors was performed with immunohistochemistry and confocal microscopy. Behavioral studies were made, after blocking ephrinA1 expression with antisense (AS) oligonucleotides, to assess hindlimb locomotor activity. Real-time PCR demonstrated basal mRNA levels of ephrin (A1, A2, A3, and A5) in the adult spinal cord. Interestingly, ephrinA1 was the only ligand whose mRNA levels were significantly altered after SCI. Although ephrinA1 mRNA levels increased after 2 weeks and remain elevated, we did not observe this pattern at the protein level as revealed by western blot analysis. Immunohistochemical studies showed ephrinA1 expression in reactive astrocytes, axons, and neurons and also their colocalization with EphA4 and A7 receptors. Behavioral studies revealed worsening of locomotor activity when ephrinA1 expression was reduced. This study suggests that ephrinA1 ligands play a role in the pathophysiology of SCI.     

MicroRNAs参与调控中枢神经系统的发育及脊髓损伤修复

microRNAs（miRNAs）不仅参与神经系统的生长发育、功能完善,还参与脊髓损伤病理及损伤后修复过程。miRNAs能使中枢神经系统按正确的时序性和空间性顺序进行发育和分化,在维持生物体记忆及生物钟方面起着重要作用。miRNAs异常表达同脊髓损伤病理过程相关。目前,体内及体外实验均已证实,miRNAs不仅能够维持神经干细胞增殖,而且可以促进神经元轴突伸长,从而为脊髓损伤的治疗带来新的治疗策略。     

Dissection and Lateral Mounting of Zebrafish Embryos: Analysis of Spinal Cord Development

The zebrafish spinal cord is an effective investigative model for nervous system research for several reasons. First, genetic, transgenic and gene knockdown approaches can be utilized to examine the molecular mechanisms underlying nervous system development. Second, large clutches of developmentally synchronized embryos provide large experimental sample sizes. Third, the optical clarity of the zebrafish embryo permits researchers to visualize progenitor, glial, and neuronal populations. Although zebrafish embryos are transparent, specimen thickness can impede effective microscopic visualization. One reason for this is the tandem development of the spinal cord and overlying somite tissue. Another reason is the large yolk ball, which is still present during periods of early neurogenesis. In this article, we demonstrate microdissection and removal of the yolk in fixed embryos, which allows microscopic visualization while preserving surrounding somite tissue. We also demonstrate semipermanent mounting of zebrafish embryos. This permits observation of neurodevelopment in the dorso-ventral and anterior-posterior axes, as it preserves the three-dimensionality of the tissue.     

Fast and Accurate Semi-Automated Segmentation Method of Spinal Cord MR Images at 3T Applied to the Construction of a Cervical Spinal Cord Template

Objective

To design a fast and accurate semi-automated segmentation method for spinal cord 3T MR images and to construct a template of the cervical spinal cord.

Materials and Methods

A semi-automated double threshold-based method (DTbM) was proposed enabling both cross-sectional and volumetric measures from 3D T₂-weighted turbo spin echo MR scans of the spinal cord at 3T. Eighty-two healthy subjects, 10 patients with amyotrophic lateral sclerosis, 10 with spinal muscular atrophy and 10 with spinal cord injuries were studied. DTbM was compared with active surface method (ASM), threshold-based method (TbM) and manual outlining (ground truth). Accuracy of segmentations was scored visually by a radiologist in cervical and thoracic cord regions. Accuracy was also quantified at the cervical and thoracic levels as well as at C2 vertebral level. To construct a cervical template from healthy subjects’ images (n=59), a standardization pipeline was designed leading to well-centered straight spinal cord images and accurate probability tissue map.

Results

Visual scoring showed better performance for DTbM than for ASM. Mean Dice similarity coefficient (DSC) was 95.71% for DTbM and 90.78% for ASM at the cervical level and 94.27% for DTbM and 89.93% for ASM at the thoracic level. Finally, at C2 vertebral level, mean DSC was 97.98% for DTbM compared with 98.02% for TbM and 96.76% for ASM. DTbM showed similar accuracy compared with TbM, but with the advantage of limited manual interaction.

Conclusion

A semi-automated segmentation method with limited manual intervention was introduced and validated on 3T images, enabling the construction of a cervical spinal cord template.