Expression of ATF3 and axonal outgrowth are impaired after delayed nerve repair

Background: A delay in surgical nerve repair results in impaired nerve function in humans, but mechanisms behind the weakened nerve regeneration are not known. Activating transcription factor 3 (ATF3) increases the intrinsic growth state of injured neurons early after injury, but the role of long-term changes and their relation to axonal outgrowth after a delayed nerve repair are not well understood. ATF3 expression was examined by immunohistochemistry in motor and sensory neurons and in Schwann cells in rat sciatic nerve and related to axonal outgrowth after transection and delayed nerve repair (repair 0, 30, 90 or 180 days post-injury). Expression of the neuronal cell adhesion molecule (NCAM), which is expressed in non-myelinating Schwann cells, was also examined. Results: The number of neurons and Schwann cells expressing ATF3 declined and the length of axonal outgrowth was impaired if the repair was delayed. The decline was more rapid in motor neurons than in sensory neurons and Schwann cells. Regeneration distances over time correlated to number of ATF3 stained neurons and Schwann cells. Many neurofilament stained axons grew along ATF3 stained Schwann cells. If nerve repair was delayed the majority of Schwann cells in the distal nerve segment stained for NCAM. Conclusion: Delayed nerve repair impairs nerve regeneration and length of axonal outgrowth correlates to ATF3 expression in both neurons and Schwann cells. Mainly non-myelinating Schwann cells (NCAM stained) are present in distal nerve segments after delayed nerve repair. These data provide a neurobiological basis for the poor outcomes associated with delayed nerve repair. Nerve trunks should, if possible, be promptly repaired

Systemic proteasome inhibition triggers neurodegeneration in a transgenic mouse model expressing human α-synuclein under oligodendrocyte promoter: implications for multiple system atrophy

Multiple system atrophy (MSA) is a progressive late onset neurodegenerative α-synucleinopathy with unclear pathogenesis. Recent genetic and pathological studies support a central role of α-synuclein (αSYN) in MSA pathogenesis. Oligodendroglial cytoplasmic inclusions of fibrillar αSYN and dysfunction of the ubiquitin–proteasome system are suggestive of proteolytic stress in this disorder. To address the possible pathogenic role of oligodendroglial αSYN accumulation and proteolytic failure in MSA we applied systemic proteasome inhibition (PSI) in transgenic mice with oligodendroglial human αSYN expression and determined the presence of MSA-like neurodegeneration in this model as compared to wild-type mice. PSI induced open field motor disability in transgenic αSYN mice but not in wild-type mice. The motor phenotype corresponded to progressive and selective neuronal loss in the striatonigral and olivopontocerebellar systems of PSI-treated transgenic αSYN mice. In contrast no neurodegeneration was detected in PSI-treated wild-type controls. PSI treatment of transgenic αSYN mice was associated with significant ultrastructural alterations including accumulation of fibrillar human αSYN in the cytoplasm of oligodendroglia, which resulted in myelin disruption and demyelination characterized by increased g-ratio. The oligodendroglial and myelin pathology was accompanied by axonal degeneration evidenced by signs of mitochondrial stress and dysfunctional axonal transport in the affected neurites. In summary, we provide new evidence supporting a primary role of proteolytic failure and suggesting a neurodegenerative pathomechanism related to disturbed oligodendroglial/myelin trophic support in the pathogenesis of MSA

Methodological advances in imaging intravital axonal transport.

Axonal transport is the active process whereby neurons transport cargoes such as organelles and proteins anterogradely from the cell body to the axon terminal and retrogradely in the opposite direction. Bi-directional transport in axons is absolutely essential for the functioning and survival of neurons and appears to be negatively impacted by both aging and diseases of the nervous system, such as Alzheimer's disease and amyotrophic lateral sclerosis. The movement of individual cargoes along axons has been studied in vitro in live neurons and tissue explants for a number of years; however, it is currently unclear as to whether these systems faithfully and consistently replicate the in vivo situation. A number of intravital techniques originally developed for studying diverse biological events have recently been adapted to monitor axonal transport in real-time in a range of live organisms and are providing novel insight into this dynamic process. Here, we highlight these methodological advances in intravital imaging of axonal transport, outlining key strengths and limitations while discussing findings, possible improvements, and outstanding questions

Targeted Ablation of Oligodendrocytes Triggers Axonal Damage

Glial dysfunction has been implicated in a number of neurodegenerative diseases. In this study we investigated the consequences of glial and oligodendrocyte ablation on neuronal integrity and survival in Drosophila and adult mice, respectively. Targeted genetic ablation of glia was achieved in the adult Drosophila nervous system using the GAL80-GAL4 system. In mice, oligodendrocytes were depleted by the injection of diphtheria toxin in MOGi-Cre/iDTR double transgenic animals. Acute depletion of oligodendrocytes induced axonal injury, but did not cause neuronal cell death in mice. Ablation of glia in adult flies triggered neuronal apoptosis and resulted in a marked reduction in motor performance and lifespan. Our study shows that the targeted depletion of glia triggers secondary neurotoxicity and underscores the central contribution of glia to neuronal homeostasis. The models used in this study provide valuable systems for the investigation of therapeutic strategies to prevent axonal or neuronal damage

Clonally expanded mitochondrial DNA deletions within the choroid plexus in multiple sclerosis

OBJECTIVE: Mitochondrial DNA deletions (Δ-mtDNA) are implicated in the pathogenesis of multiple sclerosis (MS), Parkinson’s disease (PD), Alzheimer’s disease (AD) and ageing. Given the diffuse nature of inflammation in MS, aim of this study was to determine whether Δ-mtDNA caused respiratory deficient cells in excess of age within choroid plexus (CP) and ongoing mutagenesis or clonal expansion accounted for the respiratory deficiency in MS. METHODS: Respiratory chain complex IV and complex II activity was determined sequentially using histochemistry. Δ-mtDNA were characterized using real time PCR, long range PCR, sequencing and single molecule PCR. Sources of reactive oxygen and nitrogen species (RONS) were explored using immunohistochemistry. RESULTS: Respiratory deficient cells (lacking complex IV and with intact complex II activity) within CP epithelium were in excess of age in MS, PD and AD. Subunit-I of complex IV was lacking to a greater extent in MS than controls. Percentage of respiratory deficient cells harboring >50% heteroplasmy level of Δ-mtDNA was significantly greater in MS than PD, AD and controls. Long range PCR and sequencing confirmed Δ-mtDNA. Single molecule PCR identified clonally expanded Δ-mtDNA in MS, despite an increase in sources of RONS. INTERPRETATION: Our findings establish clonal expansion of Δ-mtDNA causing respiratory deficiency in MS and the extraparenchymal intracranial location indicated the potential to involve multiple cell types. Understanding factors that influence clonal expansion of Δ-mtDNA, a molecular link between inflammation and delayed cellular energy failure, may identify potential therapeutic targets for progressive forms of MS as well as other neurodegenerative disorders

Discovery of widespread transcription initiation at microsatellites predictable by sequence-based deep neural network

Using the Cap Analysis of Gene Expression (CAGE) technology, the FANTOM5 consortium provided one of the most comprehensive maps of transcription start sites (TSSs) in several species. Strikingly, ~72% of them could not be assigned to a specific gene and initiate at unconventional regions, outside promoters or enhancers. Here, we probe these unassigned TSSs and show that, in all species studied, a significant fraction of CAGE peaks initiate at microsatellites, also called short tandem repeats (STRs). To confirm this transcription, we develop Cap Trap RNA-seq, a technology which combines cap trapping and long read MinION sequencing. We train sequence-based deep learning models able to predict CAGE signal at STRs with high accuracy. These models unveil the importance of STR surrounding sequences not only to distinguish STR classes, but also to predict the level of transcription initiation. Importantly, genetic variants linked to human diseases are preferentially found at STRs with high transcription initiation level, supporting the biological and clinical relevance of transcription initiation at STRs. Together, our results extend the repertoire of non-coding transcription associated with DNA tandem repeats and complexify STR polymorphism