GRS defective axonal distribution as a potential contributor to distal spinal muscular atrophy type V pathogenesis in a new model of GRS-associated neuropathy

Distal spinal muscular atrophy type V (dSMA-V), a hereditary axonal neuropathy, is a glycyl-tRNA synthetase (GRS)-associated neuropathy caused by a mutation in GRS. In this study, using an adenovirus vector system equipped with a neuron-specific promoter, we constructed a new GRS-associated neuropathy mouse model. We found that wild-type GRS (WT) is distributed in peripheral axons, dorsal root ganglion (DRG) cell bodies, central axon terminals and motor neuron cell bodies in the mouse model. In contrast, the L129P mutant GRS was localized in DRG and motor neuron cell bodies. Thus, we propose that the disease-causing L129P mutant is linked to a distribution defect in peripheral nerves in vivo.     

DGAT2 Mutation in a Family with Autosomal‐Dominant Early‐Onset Axonal Charcot‐Marie‐Tooth Disease

Charcot‐Marie‐Tooth disease (CMT) is the most common inherited peripheral neuropathy and is a genetically and clinically heterogeneous disorder. We examined a Korean family in which two individuals had an autosomal‐dominant axonal CMT with early‐onset, sensory ataxia, tremor, and slow disease progression. Pedigree analysis and exome sequencing identified a de novo missense mutation (p.Y223H) in the diacylglycerol O‐acyltransferase 2 (DGAT2) gene. DGAT2 encodes an endoplasmic reticulum‐mitochondrial‐associated membrane protein, acyl‐CoA:diacylglycerol acyltransferase, which catalyzes the final step of the triglyceride (TG) biosynthesis pathway. The patient showed consistently decreased serum TG levels, and overexpression of the mutant DGAT2 significantly inhibited the proliferation of mouse motor neuron cells. Moreover, the variant form of human DGAT2 inhibited the axonal branching in the peripheral nervous system of zebrafish. We suggest that mutation of DGAT2 is the novel underlying cause of an autosomal‐dominant axonal CMT2 neuropathy. This study will help provide a better understanding of the pathophysiology of axonal CMT and contribute to the molecular diagnostics of peripheral neuropathies.     

A Rare Motor Neuron Deleterious Missense Mutation in the DPYSL3 (CRMP4) Gene is Associated with ALS

The dihydropyrimidinase‐like 3 (DPYSL3) or Collapsin Response Mediator Protein 4a (CRMP4a) expression is modified in neurodegeneration and is involved in several ALS‐associated pathways including axonal transport, glutamate excitotoxicity, and oxidative stress. The objective of the study was to analyze CRMP4 as a risk factor for ALS. We analyzed the DPYSL3/CRMP4 gene in French ALS patients (n = 468) and matched‐controls (n = 394). We subsequently examined a variant in a Swedish population (184 SALS, 186 controls), and evaluated its functional effects on axonal growth and survival in motor neuron cell culture. The rs147541241:A>G missense mutation occurred in higher frequency among French ALS patients (odds ratio = 2.99) but the association was not confirmed in the Swedish population. In vitro expression of mutated DPYSL3 in motor neurons reduced axonal growth and accelerated cell death compared with wild type protein. Thus, the association between the rs147541241 variant and ALS was limited to the French population, highlighting the geographic particularities of genetic influences (risks, contributors). The identified variant appears to shorten motor neuron survival through a detrimental effect on axonal growth and CRMP4 could act as a key unifier in transduction pathways leading to neurodegeneration through effects on early axon development.     

Reduced evoked motor and sensory potential amplitudes in obstructive sleep apnea patients

It is unknown to what extent chronic intermittent hypoxaemia in obstructive sleep apnea causes damage to the motor and sensory peripheral nerves. It was hypothesized that patients with obstructive sleep apnea would have bilaterally significantly impaired amplitudes of both motor and sensory peripheral nerve‐evoked potentials of both lower and upper limbs. An observational study was conducted on 43 patients with obstructive sleep apnea confirmed by the whole‐night polysomnography, and 40 controls to assess the relationship between obstructive sleep apnea and peripheral neuropathy. All obstructive sleep apnea subjects underwent standardized electroneurographic testing, with full assessment of amplitudes of evoked compound muscle action potentials, sensory neural action potentials, motor and sensory nerve conduction velocities, and distal motor and sensory latencies of the median, ulnar, peroneal and sural nerves, bilaterally. All nerve measurements were compared with reference values, as well as between the untreated patients with obstructive sleep apnea and control subjects. Averaged compound muscle action potential and sensory nerve action potential amplitudes were significantly reduced in the nerves of both upper and lower limbs in patients with obstructive sleep apnea compared with controls (P < 0.001). These results confirmed that patients with obstructive sleep apnea had significantly lower amplitudes of evoked action potentials of both motor and sensory peripheral nerves. Clinical/subclinical axonal damage exists in patients with obstructive sleep apnea to a greater extent than previously thought.     

Human superoxide dismutase 1 overexpression in motor neurons of Caenorhabditis elegans causes axon guidance defect and neurodegeneration

Strong evidence indicates that mutant Cu, Zn-superoxide dismutase 1 (SOD1) exerts toxic effect on motor neurons in amyotrophic lateral sclerosis (ALS). However, the nature of mutant SOD1-mediated motor neuron degeneration is poorly understood. To provide new insight into the mechanism by which mutant SOD1 induces motor neuron injury, we developed novel Caenorhabditis elegans models of ALS. Expression of human wild type or G93A SOD1 specifically in motor neurons of C. elegans caused progressive locomotion defect and paralytic phenotype, which recapitulate some characteristic features of ALS including age-dependent motor dysfunction and degeneration of motor neurons associated with SOD1 aggregation. In addition, the motor neuron loss is independent of cell death protein 3 (CED-3)/cell death protein 4 (CED-4) caspase pathway. We also found that before motor neurons began to die in adulthood, axon guidance defect of motor neuron appeared during the development stages. When green fluorescent protein (GFP)-tagged proteins related to axon guidance were examined in motor neurons, a significantly decreased density and number of GFP-tagged puncta were observed in the transgenic worms. Our models mimic axon developmental defect and the adult-onset degeneration of motor neurons in ALS. Using this model, we uncovered the cell-autonomous damage caused by human SOD1 to motor neurons in vivo, and provided a new insight into the developmental defect mechanism that may contribute to motor neuron degeneration in ALS.     

Increased accumulation of transferrin by motor neurons of the mouse mutant progressive motor neuronopathy (pmn/pmn)

Summary It has been suggested that iron-carrying transferrin exerts growth-factor-like influences on motor neurons. I have evaluated the distribution of proteins related to the intracerebral iron-homeostasis in the mouse mutant progressive motor neuronopathy (pmn/pmn); an autosomal recessive mutant with progressive caudo-cranial motor neuron degeneration. A higher immunoreactivity of transferrin and transferrin receptor in motor neurons of thepmn/pmn mutant compared to that in normal mice was demonstrated. Ferritin was not observed in motor neurons of thepmn/pmn mutant. Transferrin receptors were absent from axons and neuromuscular junctions, indicating that entry of blood-borne, liver-derived transferrin (liver transferrin) into motor neurons due to uptake and subsequent retrograde axonal transport was unspecific. Due to the selective presence of transferrin receptors on neuronal somata, a more likely mode of entry of transferrin into the motor neurons was by receptor-mediated uptake of brain-derived transferrin (brain transferrin) at the soma. This study provides data on transferrin accumulation and transferrin receptor expression in diseased motor neurons and adds further insights into influences of proteins related to iron-homeostasis in the diseased PNS.     

TARDBP and FUS Mutations Associated with Amyotrophic Lateral Sclerosis: Summary and Update

Mutations in the TAR DNA Binding Protein gene (TARDBP), encoding the protein TDP‐43, were identified in amyotrophic lateral sclerosis (ALS) patients. Interestingly, TDP‐43 positive inclusion bodies were first discovered in ubiquitin‐positive, tau‐negative ALS and frontotemporal dementia (FTD) inclusion bodies, and subsequently observed in the majority of neurodegenerative disorders. To date, 47 missense and one truncating mutations have been described in a large number of familial (FALS) and sporadic (SALS) patients. Fused in sarcoma (FUS) was found to be responsible for a previously identified ALS6 locus, being mutated in both FALS and SALS patients. TARDBP and FUS have a structural and functional similarity and most of mutations in both genes are also clustered in the C‐terminus of the proteins. The molecular mechanisms through which mutant TDP‐43 and FUS may cause motor neuron degeneration are not well understood. Both proteins play an important role in mRNA transport, axonal maintenance, and motor neuron development. Functional characterization of these mutations in in vitro and in vivo systems is helping to better understand how motor neuron degeneration occurs. This report summarizes the biological and clinical relevance of TARDBP and FUS mutations in ALS. All the data reviewed here have been submitted to a database based on the Leiden Open (source) Variation Database (LOVD) and is accessible online at www.lovd.nl/TARDBP , www.lovd.nl/FUS .     

Histopathology of the late-onset motor neuron degeneration (Mnd) mutant in the mouse

The motor neuron degeneration (Mnd) is characterized by a progressive deterioration of motor function (stiff-legged gait, abnormal limb placements and grasping, and finally paralysis; moving from rear to forelimbs). There is a dramatic degeneration of spinal cord motor neurons, more severe in the lumbosacral than in the other regions, as well as variable pathology in the lower cranial nerves. Upper motor neurons of the red nucleus, reticular formation of the pons and medulla, and restricted areas of the cerebral cortex are also affected. Degenerating motor neurons share many characteristics seen in the human disease amyotropic lateral sclerosis, including loss of Nissl substance, increases in lipofuscin and abnormal cytoplasmic inclusions. Additionally, Mnd, like ALS, is a disease of later life.     

Histopathology of the late-onset motor neuron degeneration (Mnd) mutant in the mouse

The motor neuron degeneration (Mnd) is characterized by a progressive deterioration of motor function (stiff-legged gait, abnormal limb placements and grasping, and finally paralysis; moving from rear to forelimbs). There is a dramatic degeneration of spinal cord motor neurons, more severe in the lumbosacral than in the other regions, as well as variable pathology in the lower cranial nerves. Upper motor neurons of the red nucleus, reticular formation of the pons and medulla, and restricted areas of the cerebral cortex are also affected. Degenerating motor neurons share many characteristics seen in the human disease amyotropic lateral sclerosis, including loss of Nissl substance, increases in lipofuscin and abnormal cytoplasmic inclusions. Additionally, Mnd, like ALS, is a disease of later life.     

Molecular Chaperones in the Pathogenesis of Amyotrophic Lateral Sclerosis: The Role of HSPB1

Genetic discoveries in amyotrophic lateral sclerosis (ALS) have a significant impact on deciphering molecular mechanisms of motor neuron degeneration but, despite recent advances, the etiology of most sporadic cases remains elusive. Several cellular mechanisms contribute to the motor neuron degeneration in ALS, including RNA metabolism, cellular interactions between neurons and nonneuronal cells, and seeding of misfolded protein with prion‐like propagation. In this scenario, the importance of protein turnover and degradation in motor neuron homeostasis gained increased recognition. In this study, we evaluated the role of the candidate gene HSPB1, a molecular chaperone involved in several proteome‐maintenance functions. In a cohort of 247 unrelated Italian ALS patients, we identified two variants (c.570G>C, p.Gln190His and c.610dupG, p.Ala204Glyfs^*6). Functional characterization of the p.Ala204Glyfs^*6 demonstrated that the mutant protein alters HSPB1 dynamic equilibrium, sequestering the wild‐type protein in a stable dimer and resulting in a loss of chaperone‐like activity. Our results underline the relevance of identifying rare but pathogenic variations in sporadic neurodegenerative diseases, suggesting a possible correlation between specific pathomechanisms linked to HSPB1 mutations and the associated neurological phenotype. Our study provides additional lines of evidence to support the involvement of HSPB1 in the pathogenesis of sporadic ALS.     

Loss of translation elongation factor (eEF1A2) expression in vivo differentiates between Wallerian degeneration and dying‐back neuronal pathology

Wallerian degeneration and dying‐back pathology are two well‐known cellular pathways capable of regulating the breakdown and loss of axonal and synaptic compartments of neurons in vivo. However, the underlying mechanisms and molecular triggers of these pathways remain elusive. Here, we show that loss of translation elongation factor eEF1A2 expression in lower motor neurons and skeletal muscle fibres in homozygous Wasted mice triggered a dying‐back neuropathy. Synaptic loss at the neuromuscular junction occurred in advance of axonal pathology and by a mechanism morphologically distinct from Wallerian degeneration. Dying‐back pathology in Wasted mice was accompanied by reduced expression levels of the zinc finger protein ZPR1, as found in other dying‐back neuropathies such as spinal muscular atrophy. Surprisingly, experimental nerve lesion revealed that Wallerian degeneration was significantly delayed in homozygous Wasted mice; morphological assessment revealed that ~80% of neuromuscular junctions in deep lumbrical muscles at 24 h and ~50% at 48 h had retained motor nerve terminals following tibial nerve lesion. This was in contrast to wild‐type and heterozygous Wasted mice where < 5% of neuromuscular junctions had retained motor nerve terminals at 24 h post‐lesion. These data show that eEF1A2 expression is required to prevent the initiation of dying‐back pathology at the neuromuscular junction in vivo. In contrast, loss of eEF1A2 expression significantly inhibited the initiation and progression of Wallerian degeneration in vivo. We conclude that loss of eEF1A2 expression distinguishes mechanisms underlying dying‐back pathology from those responsible for Wallerian degeneration in vivo and suggest that eEF1A2‐dependent cascades may provide novel molecular targets to manipulate neurodegenerative pathways in lower motor neurons.     

X‐linked spinal muscular atrophy in mice caused by autonomous loss of ATP7A in the motor neuron

ATP7A is a copper‐transporting P‐type ATPase that is essential for cellular copper homeostasis. Loss‐of‐function mutations in the ATP7A gene result in Menkes disease, a fatal neurodegenerative disorder resulting in seizures, hypotonia and failure to thrive, due to systemic copper deficiency. Most recently, rare missense mutations in ATP7A that do not impact systemic copper homeostasis have been shown to cause X‐linked spinal muscular atrophy type 3 (SMAX3), a distal hereditary motor neuropathy. An understanding of the mechanistic and pathophysiological basis of SMAX3 is currently lacking, in part because the disease‐causing mutations have been shown to confer both loss‐ and gain‐of‐function properties to ATP7A, and because there is currently no animal model of the disease. In this study, the Atp7a gene was specifically deleted in the motor neurons of mice, resulting in a degenerative phenotype consistent with the clinical features in affected patients with SMAX3, including the progressive deterioration of gait, age‐dependent muscle atrophy, denervation of neuromuscular junctions and a loss of motor neuron cell bodies. Taken together, these data reveal autonomous requirements for ATP7A that reveal essential roles for copper in the maintenance and function of the motor neuron, and suggest that SMAX3 is caused by a loss of ATP7A function that specifically impacts the spinal motor neuron. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Bee venom attenuates neuroinflammatory events and extends survival in amyotrophic lateral sclerosis models

Background  

Amyotrophic lateral sclerosis (ALS) is a disease affecting the central nervous system that is either sporadic or familial origin and causing the death of motor neurons. One of the genetic factors contributing to the etiology of ALS is mutant SOD1 (mtSOD1), which induces vulnerability of motor neurons through protein misfolding, mitochondrial dysfunction, oxidative damage, cytoskeletal abnormalities, defective axonal transport, glutamate excitotoxicity, inadequate growth factor signaling, and neuroinflammation. Bee venom has been used in the practice of Oriental medicine and evidence from the literature indicates that BV plays an anti-inflammatory or anti-nociceptive role against inflammatory reactions associated with arthritis and other inflammatory diseases. The purpose of the present study was to determine whether bee venom suppresses motor neuron loss and microglial cell activation in hSOD1^G93A mutant mice.     

Embryonic motor axon development in the severe SMA mouse

Spinal muscular atrophy (SMA) is caused by reduced levels of survival motor neuron (SMN) protein. Previously, cultured SMA motor neurons showed reduced growth cone size and axonal length. Furthermore, reduction of SMN in zebrafish resulted in truncation followed by branching of motor neuron axons. In this study, motor neurons labeled with green fluorescent protein (GFP) were examined in SMA mice from embryonic day 10.5 to postnatal day 2. SMA motor axons showed no defect in axonal formation or outgrowth at any stage of development. However, a significant increase in synapses lacking motor axon input was detected in embryonic SMA mice. Therefore, one of the earliest detectable morphological defects in the SMA mice is the loss of synapse occupation by motor axons. This indicates that in severe SMA mice there are no defects in motor axon formation however, we find evidence of denervation in embryogenesis.     

皮神经感觉传导速度和波幅检测对糖尿病周围神经病的诊断作用

目的：探讨皮神经感觉传导速度（SCV）,波幅（Amp)检测对糖尿病患者周围神经远端受损的早期诊断价值。方法：受试者为糖悄病患者组和正常对照组各25例,对前臂外侧皮神经和腓肠神经共100条,就其感觉传导速度（SVC）及其诱发的波幅（Amp)进行分析,并与胫神经,正中神经进行比较,结果：混合神经的异常率明显低于皮神经,尤以无周围神经症状的糖尿病者更加明显（P＜0．01）,结论：皮神经的SCV,Amp检测,对糖尿病周围神经远端受损的早期诊断可靠,灵敏度高。     

Slowly progressing lower motor neuron disease caused by a novel duplication mutation in exon 1 of the SOD1 gene

Familial amyotrophic lateral sclerosis accounts for about 5% of all cases of the neurodegenerative disorder amyotrophic lateral sclerosis. Genetic mutations in Cu/Zn superoxide dismutase (SOD1) have been associated with one kind of familial amyotrophic lateral sclerosis (ALS1). We identified a novel duplication mutation in exon 1 of the SOD1 gene in a Japanese family whose members had lower motor neuron diseases. The patients showed slow disease progression, with the onset of lower limb muscle weakness and exertional dyspnea. Some patients had mild motor and sensory neuropathy and/or bladder dysfunction, which is further evidence that SOD1 mutation results in a predominantly lower motor neuron phenotype.     

The neuroprotective factor Wld fails to mitigate distal axonal and neuromuscular junction (NMJ) defects in mouse models of spinal muscular atrophy

Spinal muscular atrophy (SMA) is a common autosomal recessive neurodegenerative disorder in humans. Amongst the earliest signs of neurodegeneration are severe and progressive defects of the neuromuscular synapse. These defects, characterized by poor terminal arborization and immature motor endplates, presumably result in a loss of functional synapses. The slow Wallerian degeneration (Wld^s) mutation in rodents has been shown to have a protective effect on mouse models of motor neuron disease by retarding axonal die-back and preventing neuromuscular synapse loss. In this study we tested the effects of the Wld^s mutation on the disease phenotype of SMA model mice. Consistent with previous reports, the mutation slows axon and neuromuscular synapse loss following nerve injury in wild-type as well as in SMA mice. However, the synaptic defects found in severely affected SMA patients and model mice persist in the double (Wld^s;SMA) mutants. No delay in disease onset was observed and survival was not significantly altered. Finally, Wld^s had no effect on the striking phrenic nerve projection defects that we discovered in SMA model mice. Our results indicate that the reported protective effects of Wld^s are insufficient to mitigate the neuromuscular phenotype due to reduced SMN protein, and that the mechanisms responsible for distal defects of the motor unit in SMA are unlikely to be similar to those causing neurodegeneration in genetic mutants such as the pmn mouse which is partially rescued by the Wld^s protein.     

Extension of the phenotype of biallelic loss‐of‐function mutations in SLC25A46 to the severe form of pontocerebellar hypoplasia type I

Biallelic mutations in SLC25A46, encoding a modified solute transporter involved in mitochondrial dynamics, have been identified in a wide range of conditions such as hereditary motor and sensory neuropathy with optic atrophy type VIB (OMIM: *610826) and congenital lethal pontocerebellar hypoplasia (PCH). To date, 18 patients from 13 families have been reported, presenting with the key clinical features of optic atrophy, peripheral neuropathy, and cerebellar atrophy. The course of the disease was highly variable ranging from severe muscular hypotonia at birth and early death to first manifestations in late childhood and survival into the fifties. Here we report on 4 patients from 2 families diagnosed with PCH who died within the first month of life from respiratory insufficiency. Patients from 1 family had pathoanatomically proven spinal motor neuron degeneration (PCH1). Using exome sequencing, we identified biallelic disease‐segregating loss‐of‐function mutations in SLC25A46 in both families. Our study adds to the definition of the SLC25A46‐associated phenotypic spectrum that includes neonatal fatalities due to PCH as the severe extreme.     

Novel Mutations in the DYNC1H1 Tail Domain Refine the Genetic and Clinical Spectrum of Dyneinopathies

The heavy chain 1 of cytoplasmic dynein (DYNC1H1) is responsible for movement of the motor complex along microtubules and recruitment of dynein components. Mutations in DYNC1H1 are associated with spinal muscular atrophy (SMA), hereditary motor and sensory neuropathy (HMSN), cortical malformations, or a combination of these. Combining linkage analysis and whole‐exome sequencing, we identified a novel dominant defect in the DYNC1H1 tail domain (c.1792C>T, p.Arg598Cys) causing axonal HMSN. Mutation analysis of the tail region in 355 patients identified a de novo mutation (c.791G>T, p.Arg264Leu) in an isolated SMA patient. Her phenotype was more severe than previously described, characterized by multiple congenital contractures and delayed motor milestones, without brain malformations. The mutations in DYNC1H1 increase the interaction with its adaptor BICD2. This relates to previous studies on BICD2 mutations causing a highly similar phenotype. Our findings broaden the genetic heterogeneity and refine the clinical spectrum of DYNC1H1, and have implications for molecular diagnostics of motor neuron diseases.