Modelling human CNS injury with human neural stem cells in 2- and 3-Dimensional cultures.

The adult human central nervous system (CNS) has very limited regenerative capability, and injury at the cellular and molecular level cannot be studied in vivo. Modelling neural damage in human systems is crucial to identifying species-specific responses to injury and potentially neurotoxic compounds leading to development of more effective neuroprotective agents. Hence we developed human neural stem cell (hNSC) 3-dimensional (3D) cultures and tested their potential for modelling neural insults, including hypoxic-ischaemic and Ca2+-dependent injury. Standard 3D conditions for rodent cells support neuroblastoma lines used as human CNS models, but not hNSCs, but in all cases changes in culture architecture alter gene expression. Importantly, response to damage differs in 2D and 3D cultures and this is not due to reduced drug accessibility. Together, this study highlights the impact of culture cytoarchitecture on hNSC phenotype and damage response, indicating that 3D models may be better predictors of in vivo response to damage and compound toxicity.

Immune Surveillance of the CNS following Infection and Injury.

The central nervous system (CNS) contains a sophisticated neural network that must be constantly surveyed in order to detect and mitigate a diverse array of challenges. The innate and adaptive immune systems actively participate in this surveillance, which is critical for the maintenance of CNS homeostasis and can facilitate the resolution of infections, degeneration, and tissue damage. Infections and sterile injuries represent two common challenges imposed on the CNS that require a prompt immune response. While the inducers of these two challenges differ in origin, the resultant responses orchestrated by the CNS share some overlapping features. Here, we review how the CNS immunologically discriminates between pathogens and sterile injuries, mobilizes an immune reaction, and, ultimately, regulates local and peripherally-derived immune cells to provide a supportive milieu for tissue repair.

Emerging potential of exosomes and noncoding microRNAs for the treatment of neurological injury/diseases.

Recent discoveries of cellular generation of exosomes, small (∼ 30 - 100 nm) complex lipid membrane structures which encapsulate and transport proteins, RNAs, including microRNAs (miRNAs) have provided new insight in how cells within organisms communicate. These discoveries will likely have a major impact on the treatment of disease, with cancers and neurological diseases as evident targets. Exosomes provide a major medium of intercellular communications and thereby, there being a potential by altering communications and instructions for protein production, we can employ exosomes to treat diseases. We now have an opportunity to treat neurological disease by modifying intercellular communication networks. Recent work demonstrating that the therapeutic benefit provided by stem cells for the treatments of stroke and traumatic brain injury depend on their generation and release of exosomes provides a foundation for exosome-based therapy. Cell-free exosomes have also been recently employed to effectively treat stroke and brain trauma. The content of exosomes, particularly their miRNA cargo which can concurrently impact the post-transcriptional regulation of many genes, can be regulated. We are at the cusp of capitalizing on this important means of intercellular communications for the treatment of diseases, such as cancers and neurological diseases, among many others.

The role of the Gadd45 family in the nervous system: a focus on neurodevelopment, neuronal injury, and cognitive neuroepigenetics.

The growth arrest and DNA damage-inducible (Gadd)45 proteins have been associated with numerous cellular mechanisms including cell-cycle control, DNA damage sensation and repair, genotoxic stress, neoplasia, and molecular epigenetics. The genes were originally identified in in vitro screens of irradiation- and interleukin-induced transcription and have since been implicated in a host of normal and aberrant central nervous system processes. These include early and postnatal development, injury, cancer, memory, aging, and neurodegenerative and psychiatric disease states. The proteins act through a variety of molecular signaling cascades including the MAPK cascade, cell-cycle control mechanisms, histone regulation, and epigenetic DNA demethylation. In this review, we provide a comprehensive discussion of the literature implicating each of the three members of the Gadd45 family in these processes.

AMIGO3 is an NgR1/p75 co-receptor signalling axon growth inhibition in the acute phase of adult central nervous system injury.

Axon regeneration in the injured adult CNS is reportedly inhibited by myelin-derived inhibitory molecules, after binding to a receptor complex comprised of the Nogo-66 receptor (NgR1) and two transmembrane co-receptors p75/TROY and LINGO-1. However, the post-injury expression pattern for LINGO-1 is inconsistent with its proposed function. We demonstrated that AMIGO3 levels were significantly higher acutely than those of LINGO-1 in dorsal column lesions and reduced in models of dorsal root ganglion neuron (DRGN) axon regeneration. Similarly, AMIGO3 levels were raised in the retina immediately after optic nerve crush, whilst levels were suppressed in regenerating optic nerves, induced by intravitreal peripheral nerve implantation. AMIGO3 interacted functionally with NgR1-p75/TROY in non-neuronal cells and in brain lysates, mediating RhoA activation in response to CNS myelin. Knockdown of AMIGO3 in myelin-inhibited adult primary DRG and retinal cultures promoted disinhibited neurite growth when cells were stimulated with appropriate neurotrophic factors. These findings demonstrate that AMIGO3 substitutes for LINGO-1 in the NgR1-p75/TROY inhibitory signalling complex and suggests that the NgR1-p75/TROY-AMIGO3 receptor complex mediates myelin-induced inhibition of axon growth acutely in the CNS. Thus, antagonizing AMIGO3 rather than LINGO-1 immediately after CNS injury is likely to be a more effective therapeutic strategy for promoting CNS axon regeneration when combined with neurotrophic factor administration.

Microfluidic chips for in vivo imaging of cellular responses to neural injury in Drosophila larvae.

With powerful genetics and a translucent cuticle, the Drosophila larva is an ideal model system for live imaging studies of neuronal cell biology and function. Here, we present an easy-to-use approach for high resolution live imaging in Drosophila using microfluidic chips. Two different designs allow for non-invasive and chemical-free immobilization of 3(rd) instar larvae over short (up to 1 hour) and long (up to 10 hours) time periods. We utilized these 'larva chips' to characterize several sub-cellular responses to axotomy which occur over a range of time scales in intact, unanaesthetized animals. These include waves of calcium which are induced within seconds of axotomy, and the intracellular transport of vesicles whose rate and flux within axons changes dramatically within 3 hours of axotomy. Axonal transport halts throughout the entire distal stump, but increases in the proximal stump. These responses precede the degeneration of the distal stump and regenerative sprouting of the proximal stump, which is initiated after a 7 hour period of dormancy and is associated with a dramatic increase in F-actin dynamics. In addition to allowing for the study of axonal regeneration in vivo, the larva chips can be utilized for a wide variety of in vivo imaging applications in Drosophila.

MicroRNA in central nervous system trauma and degenerative disorders.

MicroRNAs (miRNAs) are a novel class of small noncoding RNAs that negatively regulate gene expression at the posttranscriptional level by binding to the 3'-untranslated region of target mRNAs leading to their translational inhibition or sometimes degradation. MiRNAs are predicted to control the activity of at least 20-30% of human protein-coding genes. Recent studies have demonstrated that miRNAs are highly expressed in the central nervous system (CNS) including the brain and spinal cord. Although we are currently in the initial stages of understanding how this novel class of gene regulators is involved in neurological biological functions, a growing body of exciting evidence suggests that miRNAs are important regulators of diverse biological processes such as cell differentiation, growth, proliferation, and apoptosis. Moreover, miRNAs are key modulators of both CNS development and plasticity. Some miRNAs have been implicated in several neurological disorders such as traumatic CNS injuries and neurodegenerative diseases. Recently, several studies suggested the possibility of miRNA involvement in neurodegeneration. Identifying the roles of miRNAs and their target genes and signaling pathways in neurological disorders will be critical for future research. miRNAs may represent a new layer of regulators for neurobiology and a novel class of therapeutic targets for neurological diseases.

Alterations in neuronal gene expression profiles in response to experimental demyelination and axonal transection.

The main pathological features of multiple sclerosis, demyelination and axonal transection, are considered to cause reversible and irreversible neurological deficits, respectively. This study aimed to separately analyze the effects of these pathological hallmarks on neuronal gene expression in experimental paradigms. The pontocerebellar pathway was targeted with either lysolecithin-induced chemical demyelination or a complete pathway transection (axonal transection) in rats. Transcriptional changes in the pontocerebellar neurons were investigated with microarrays at days 4, 10 and 37 post-intervention, which was confirmed by immunohistochemistry on protein level. A common as well as unique set of injury-response genes was identified. The increased expression of activating transcription factor 3 (Atf3) and thyrotropin-releasing hormone (Trh) in both injury paradigms was validated by immunohistochemistry. The expression of Atf3 in a patient with Marburg's variant of multiple sclerosis was also detected, also confirming the activation of the Atf3 pathway in a human disease sample. It was concluded that this experimental approach may be useful for the identification of pathways that could be targeted for remyelinative or neuroprotective drug development.

Deficiency of the complement regulator CD59a exacerbates Wallerian degeneration.

The complement system is implicated in Wallerian degeneration (WD). We have previously shown that the membrane attack complex (MAC), the terminal activation product of the complement cascade, mediates rapid axonal degradation and myelin clearance during WD after peripheral nerve injury. In this study we analyzed the contribution of CD59a, a cell membrane negative regulator of the MAC, to WD. Following injury, the level of MAC deposition was higher in the CD59a deficient mice than wildtypes whereas the residual axonal content was lower in CD59a deficient mice than wildtypes, strongly implicating MAC as a determinant of axonal damage during WD. The number of endoneurial macrophages was significantly higher in CD59a deficient mice compared to wildtypes at 1 day post-injury. These findings are relevant to the understanding of the mechanisms of axon loss in injury and disease.

The proteomics of neurodegeneration.

The continuing improvement and refinement of proteomic and bioinformatic tools has made it possible to obtain increasing amounts of structural and functional information about proteins on a global scale. The emerging field of neuroproteomics promises to provide powerful strategies for further characterizing neuronal dysfunction and cell loss associated with neurodegenerative diseases. Neuroproteomic studies have thus far revealed relatively comprehensive quantitative changes and post-translational modifications (mostly oxidative damage) of high abundance proteins, confirming deficits in energy production, protein degradation, antioxidant protein function, and cytoskeletal regulation associated with neurodegenerative diseases such as Alzheimer and Parkinson disease. The identification of changes in low-abundance proteins and characterization of their functions based on protein-protein interactions still await further development of proteomic methodologies and more dedicated application of these technologies by neuroscientists. Once accomplished, however, the resulting information will certainly provide a truly comprehensive view of neurodegeneration-associated changes in protein expression, facilitating the identification of novel biomarkers for the early detection of neurodegenerative diseases and new targets for therapeutic intervention.

Genes and nerves.

The unpredictability of a brachial plexus graft, a median nerve repair, or a facial-nerve reconstruction is well known. No matter how precise the technical skills, a perfect recovery from a peripheral-nerve lesion is elusive. To resolve this problem, understanding of the normal development of the peripheral nervous system is needed. Presently, the development of the innervation in the upper limb is complex and not fully understood. However, many of the genes involved in this process are now known, and the link between anatomy and genetics is becoming clearer. This short review aims to acquaint the clinical surgeon with some of the main genes. The principal steps in the establishment of neural circuits will be summarized, in particular, the specification and development of neurons and glia, the pathfinding of cells and axons towards their target, and the downstream molecules that control the circuitry of these neurons.

Elevation of superoxide dismutase increases acoustic trauma from noise exposure.

The generation of superoxide has been implicated as a cause of cochlear damage from excessive noise. Cu/Zn superoxide dismutase (SOD1) generally will protect against superoxide-mediated tissue injury but protection by this enzyme against noise trauma is controversial. This study assessed auditory function in C57BL/6 mice overexpressing SOD1 or treated with lecithinized SOD1 (PC-SOD1). Noise exposure caused significantly higher threshold shifts in PC-SOD1-treated animals than physiological saline-treated animals. Cochlear tissues of PC-SOD1-treated animals exhibited significant elevation of the levels in the SOD activity, not in the catalase activity, in comparison with those of saline-treated animals. Likewise, transgenic mice overexpressing SOD1 tended to suffer higher threshold shifts than nontransgenic littermates from noise exposure. The findings indicate that increasing SOD1 enhances auditory dysfunction following noise exposure.