Secondary Axonal Damage Research Articles

A new mouse model of Charcot-Marie-Tooth 2J neuropathy replicates human axonopathy and suggest alteration in axo-glia communication.

Myelin is essential for rapid nerve impulse propagation and axon protection. Accordingly, defects in myelination or myelin maintenance lead to secondary axonal damage and subsequent degeneration. Studies utilizing genetic (CNPase-, MAG-, and PLP-null mice) and naturally occurring neuropathy models suggest that myelinating glia also support axons independently from myelin. Myelin protein zero (MPZ or P0), which is expressed only by Schwann cells, is critical for myelin formation and maintenance in the peripheral nervous system. Many mutations in MPZ are associated with demyelinating neuropathies (Charcot-Marie-Tooth disease type 1B [CMT1B]). Surprisingly, the substitution of threonine by methionine at position 124 of P0 (P0T124M) causes axonal neuropathy (CMT2J) with little to no myelin damage. This disease provides an excellent paradigm to understand how myelinating glia support axons independently from myelin. To study this, we generated targeted knock-in MpzT124M mutant mice, a genetically authentic model of T124M-CMT2J neuropathy. Similar to patients, these mice develop axonopathy between 2 and 12 months of age, characterized by impaired motor performance, normal nerve conduction velocities but reduced compound motor action potential amplitudes, and axonal damage with only minor compact myelin modifications. Mechanistically, we detected metabolic changes that could lead to axonal degeneration, and prominent alterations in non-compact myelin domains such as paranodes, Schmidt-Lanterman incisures, and gap junctions, implicated in Schwann cell-axon communication and axonal metabolic support. Finally, we document perturbed mitochondrial size and distribution along MpzT124M axons suggesting altered axonal transport. Our data suggest that Schwann cells in P0T124M mutant mice cannot provide axons with sufficient trophic support, leading to reduced ATP biosynthesis and axonopathy. In conclusion, the MpzT124M mouse model faithfully reproduces the human neuropathy and represents a unique tool for identifying the molecular basis for glial support of axons.

Prognostic Significance of Serial Nerve Conduction in GB Syndrome.

We evaluated dynamic changes in neurophysiology of Guillain-Barré syndrome (GBS) at different time points and the role of demyelination and axonal burden in predicting outcome. Nerve conduction study (NCS) was done in 44 GBS patients at admission and at 1 and 3 months, and were categorized into acute inflammatory demyelinating polyradiculoneuropathy (AIDP), acute motor axonal neuropathy (AMAN), acute motor sensory axonal neuropathy (AMSAN), equivocal and in-excitable motor nerve (IMN). The demyelinating and axonal burden on motor NCS at admission, 1 and 3 months were computed and correlated with disability at 3 and 6 months. Disability was assessed using Clinical Grading Scale. Twenty-four (54.3%) had AIDP, 5 (11.4%) AMAN, 12 (27.3%) equivocal and 3 (6.8%) had IMN at admission. Maximum instability was noted in equivocal group; majority of whom became AIDP at three months. Neurophysiological subtypes at different time points did not correlate with 6 months disability, but demyelination burden at admission (r = -0.42; P = 0.005) and axonal burden at one month (r = 0.43; P = 0.04) correlated with six months disability. Inverse correlation of axonal burden at one and three months with disability suggests role of secondary axonal damage in predicting outcome. Repeat NCS at one month helps in categorizing GBS and also in prognostication.

CIDP Refractory to Three Types of Mainstay Treatments: Differential Diagnosis and Treatment Strategies

Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is a syndrome constructed by several clinical phenotypes that share chronic inflammatory demyelination in the peripheral nervous system. While the detailed pathogenesis is not elucidated, mainstay induction therapies such as corticosteroids, IVIg, and plasma exchange, are effective for typical CIDP. However, most conventional treatments show inadequate responses in CIDP variants. Furthermore, patients with IgG4-predominant autoantibodies (anti-NF155 Ab, anti-CNTN1 Ab, and so on) show distal-predominant disability and are recognized as refractory CIDP (autoimmune nodopathy). Combining therapeutics with induction of plasma exchange following intermittent high-dose corticosteroids could be adequate for those patients. Besides, as a novel therapeutic option, rituximab is strongly expected to be a first-line for IgG4-positive autoimmune nodopathy. Some patients show relapses before the next IVIg maintenance. We can change from intravenous immunoglobulin per three weeks to weekly subcutaneous induction. Add on corticosteroids or immunosuppressants would also be helpful to the disease stability. Recently, serum NF-L has been a candidate biomarker for secondary axonal damage in CIDP. A high-level Nf-L suggests an active phase of the disease and could indicate the requirement for therapeutic intervention.

Nerve Ultrasound Distinguishes Non-Inflammatory Axonal Polyneuropathy From Inflammatory Polyneuropathy With Secondary Axonal Damage.

IntroductionChronic inflammatory demyelinating polyneuropathy (CIDP) may have a similar clinical and electrophysiological presentation to non-inflammatory axonal polyneuropathies (NIAPs) when secondary axonal damage occurs. We aimed to investigate if nerve ultrasound can help to differentiate CIDP with additional secondary axonal damage from NIAP.MethodsIn a retrospective analysis, the cross-sectional area (CSA) of the peripheral nerves measured by ultrasound at six suitable nerve sites was compared in 95 patients with CIDP and 82 patients with NIAP. We developed the adjusted Bochum ultrasound score (aBUS) ranging from 0 to 6 resulting from the number of sites with enlarged CSA (median, ulnar, radial, and sural nerve).ResultsThe mean CSA of patients with CIDP was enlarged at all six nerve sites compared with the mean CSA of patients with NIAP. A total of 21 patients with CIDP did not meet 2010 electrophysiological diagnostic criteria (European Academy of Neurology/Peripheral Nerve Society Guideline, EFNS/PNS criteria) for CIDP at examination timepoint but only in further follow-up, while 25 patients with NIAP fulfilled electrophysiological EFNS/PNS criteria for CIDP as “possible” or “probable” CIDP. To increase diagnostic power, we included aBUS measured by ultrasound in patients classified as “possible” or “probable” resulting in an improved specificity of 94% and a sensitivity of 59%, compared to a specificity of the EFNS/PNS criteria alone of 60% and sensitivity of 78%.ConclusionUsing nerve ultrasound and the aBUS as a complementary method to distinguish CIDP from NIAP in case of secondary axonal damage can facilitate the diagnosis of CIDP.

Increased muscle echointensity correlates with clinical disability and muscle strength in chronic inflammatory demyelinating polyneuropathy.

We evaluated muscle echointensity as a marker for secondary axonal damage in patients with chronic inflammatory demyelinating polyneuropathy (CIDP) using ultrasonography. Findings were correlated with clinical disability and muscular strength. Eighty patients with CIDP (40 with typical and 40 with atypical CIDP) were examined clinically, including assessment of Medical Research Council (MRC) sum score and Inflammatory Neuropathy Cause and Treatment Overall Disability Sum Score (INCAT-ODSS). Echointensity in eight proximal and distal muscles of the arms and legs was evaluated by muscle ultrasonography using the Heckmatt scale. Alterations of echointensity occurred most frequently in the distal leg muscles, with a median (range) Heckmatt score of 1.5 (1-4). There were no differences between typical and atypical CIDP patients with regard to Heckmatt score. Alterations of echointensity correlated to disability and muscle strength. The arm score of the INCAT-ODSS correlated to Heckmatt score for the distal arm muscles (r=0.23, p=0.046) and the leg score of the INCAT-ODSS correlated to Heckmatt scores for the proximal (r=0.34, p=0.002) and distal leg muscles (r=0.33, p=0.004). MRC sum score, as well as individual MRC scores for arm and leg muscles, correlated to Heckmatt scores of the corresponding muscle groups (r=-0.25, p=0.02 for MRC sum score). Increased muscle echointensity, reflecting fibrosis and fatty infiltration due to secondary axonal damage, correlated to muscular strength and disability in a large cohort of CIDP patients. Alterations of echointensity occur in both typical and atypical CIDP patients and are pronounced in the distal leg muscles.

Creatine Kinase and Progression Rate in Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease with no recognized clinical prognostic factor. Creatinine kinase (CK) increase in these patients is already described with conflicting results on prognosis and survival. In 126 ALS patients who were fast or slow disease progressors, CK levels were assayed for 16 months every 4 months in an observational case-control cohort study with prospective data collection conducted in Italy. CK was also measured at baseline in 88 CIDP patients with secondary axonal damage and in two mouse strains (129SvHSD and C57-BL) carrying the same SOD1G93A transgene expression but showing a fast (129Sv-SOD1G93A) and slow (C57-SOD1G93A) ALS progression rate. Higher CK was found in ALS slow progressors compared to fast progressors in T1, T2, T3, and T4, with a correlation with Revised Amyotrophic Lateral Sclerosis Functional Rating Scale (ALSFRS-R) scores. Higher CK was found in spinal compared to bulbar-onset patients. Transgenic and non-transgenic C57BL mice showed higher CK levels compared to 129SvHSD strain. At baseline mean CK was higher in ALS compared to CIDP. CK can predict the disease progression, with slow progressors associated with higher levels and fast progressors to lower levels, in both ALS patients and mice. CK is higher in ALS patients compared to patients with CIDP with secondary axonal damage; the higher levels of CK in slow progressors patients, but also in C57BL transgenic and non-transgenic mice designs CK as a predisposing factor for disease rate progression.

Anti-myelin-associated glycoprotein antibody positive IgM monoclonal gammopathy related peripheral neuropathy: 11 cases and literature review

目的提高对罕见的抗髓鞘相关糖蛋白（MAG）抗体阳性的IgM相关性周围神经病（IgM-PN）的认识。方法总结2014年1月至2019年4月北京协和医院诊断的11例抗MAG抗体阳性的IgM-PN患者的临床特点、实验室检查、治疗方案和预后。结果11例患者中，男8例，女3例，中位发病年龄63（52~77）岁。其中9例患者以远端肢体麻木起病，6例伴肌力减退。神经传导速度检查示，均为周围神经脱髓鞘损害，以下肢感觉神经损害为主，6例伴慢性轴索损害。11例患者均存在血清IgM型单克隆免疫球蛋白，6例为IgM κ型，3例为IgM λ型，2例为IgM κ/IgG κ双克隆型。3例患者继发于巨球蛋白血症。11例患者的血清抗MAG抗体均为阳性。9例患者接受利妥昔单抗单药或联合化疗，治疗后7例患者的神经症状稳定或改善。结论抗MAG抗体阳性的IgM-PN是一种罕见的M蛋白相关性疾病。对于伴IgM型M蛋白的周围神经病患者，应常规筛查抗MAG抗体。基于利妥昔单抗的治疗可作为其一线治疗方案。

CK prognostic factor in fast/slow progressive Amyotrophic Lateral Sclerosis (ALS)

To measure serum creatine kinase (CK) and myoglobin in ALS and Chronic Inflammatory Demyelinating Neuropathy (CIDP) patients with secondary axonal damage to investigate mechanisms of muscular damage and to assess whether CK in ALS is correlated with clinical features and disease progression. CK and myoglobin were quantified in 116 ALS and 46 CIDP. Patients with ALS were followed for 20 months, subdivided in fast or slow disease progressors according to mean monthly reduction of functional scale (ALSFRS score), whereas CK was measured at different times. Relationship with onset site, proximal and distal nerve compound action potential (cMAP) was analyzed. At baseline CK was raised in ALS patients with spinal compared to bulbar onset. CK in CIDP patients was normal. Myoglobin was normal in both diseases. Slow progressive patients showed higher CK than fast progressive (p

Neurofascin-155 IgM autoantibodies in patients with inflammatory neuropathies

ObjectivesRecently, IgG autoantibodies against different paranodal proteins have been detected and this has led to important advances in the management of inflammatory neuropathies. In contrast, not much is known on...

A new complement evasion strategy: How S. pyogenes exploits IgGs to recruit more C4BP

Neurological symptoms of patients with p.Cys89Tyr mutation in the CD59 gene include recurrent peripheral neuropathy resembling Guillain-Barré syndrome, characterized by sensory-motor demyelinating neuropathy with secondary axonal damage and moderate enhancement of the nerve roots on spine MRI, together with recurrent strokes and retinal involvement. Three additional mutations in CD59, leading to loss of function, have been described, and overall, 12/12 (100%) of patients with any mutation presented with neurological symptoms; 11/12 (92%) patients presented with recurrent peripheral neuropathy, 6/12 (50%) with recurrent strokes, and 1/12 (8%) with retinal involvement. We review the possible thrombophilic profile associated with the mutations. In these patients, excessive intravascular hemolysis saturates scavenger mechanisms resulting in free hemoglobin in plasma that irreversibly reacts with nitric oxide to form nitrate and methemoglobin, leading to arterial thrombosis. CD59 loss of function is also one of the major thrombophilic mechanisms in patients with paroxysmal nocturnal hemoglobinuria. We then describe the relationship with demyelination. The lack of CD59 allows uncontrolled complement amplification following low-level spontaneous-, viral-, or post viral-induced complement activation, resulting in severe demyelination in the peripheral nervous system. It is interesting, and certainly encouraging, that after 3 years, following 4 patients with Cys89Tyr mutations who are treated with eculizumab, no strokes occurred and non-permanent neurological insults underwent resolution without any new neurological exacerbations.

The toll-like receptor 2 agonist Pam3CSK4 is neuroprotective after spinal cord injury

Microglia/macrophage activation and recruitment following spinal cord injury (SCI) is associated with both detrimental and reparative functions. Stimulation of the innate immune receptor Toll-like receptor-2 (TLR2) has shown to be beneficial following SCI, and it increases axonal regeneration following optic nerve crush. However, the mechanism(s) remain unclear. As microglia express high levels of TLR2, we hypothesized that modulating the microglial response to injury using a specific TLR2 agonist, Pam3CSK4, would prevent secondary-mediated white matter degeneration following SCI. To test this hypothesis, we documented acute changes in microglia, axons, and oligodendroglia over time using two-photon excitation and an ex vivo laser-induced SCI (LiSCI) model. We utilized double transgenic mice that express GFP in either microglia or oligodendroglia, and YFP in axons, and we applied the lipophilic fluorescent dye (Nile Red) to visualize myelin. We found that treatment with Pam3CSK4 initiated one hour after injury induced a significant increase in the extent and timing of the microglial response to injury compared to vehicle controls. This enhanced response was observed 2 to 4h following SCI and was most prominent in areas closer to the ablation site. In addition, Pam3CSK4 treatment significantly reduced axonal dieback rostral and caudal to the ablation at 6h post-SCI. This protective effect of Pam3CSK4 was also mirrored when assessing secondary bystander axonal damage (i.e., axons spared by the primary injury that then succumb to secondary degeneration), and when assessing the survival of oligodendroglia. Following these imaging experiments, custom microarray analysis of the ex vivo spinal cord preparations revealed that Pam3CSK4-treatment induced an alternative (mixed M1:M2) microglial activation profile. In summary, our data suggest that by providing a second “sterile” activation signal to microglia through TLR2/TLR1 signaling, the microglial response to injury can be modulated in situ and is highly neuroprotective.

An initial top-down proteomic analysis of the standard cuprizone mouse model of multiple sclerosis.

An initial proteomic analysis of the cuprizone mouse model to characterise the breadth of toxicity by assessing cortex, skeletal muscle, spleen and peripheral blood mononuclear cells. Cuprizone treated vs. control mice for an initial characterisation. Select tissues from each group were pooled, analysed in triplicate using two-dimensional gel electrophoresis (2DE) and deep imaging and altered protein species identified using liquid chromatography tandem mass spectrometry (LC/MS/MS). Forty-three proteins were found to be uniquely detectable or undetectable in the cuprizone treatment group across the tissues analysed. Protein species identified in the cortex may potentially be linked to axonal damage in this model, and those in the spleen and peripheral blood mononuclear cells to the minimal peripheral immune cell infiltration into the central nervous system during cuprizone mediated demyelination. Primary oligodendrocytosis has been observed in type III lesions in multiple sclerosis. However, the underlying mechanisms are poorly understood. Cuprizone treatment results in oligodendrocyte apoptosis and secondary demyelination. This initial analysis identified proteins likely related to axonal damage; these may link primary oligodendrocytosis and secondary axonal damage. Furthermore, this appears to be the first study of the cuprizone model to also identify alterations in the proteomes of skeletal muscle, spleen and peripheral blood mononuclear cells. Notably, protein disulphide isomerase was not detected in the cuprizone cohort; its absence has been linked to reduced major histocompatibility class I assembly and reduced antigen presentation. Overall, the results suggest that, like experimental autoimmune encephalomyelitis, results from the standard cuprizone model should be carefully considered relative to clinical multiple sclerosis.

Proteomics-level analysis of myelin formation and regeneration in a mouse model for Vanishing White Matter disease.

Vanishing white matter (VWM) is a recessive neurodegenerative disease caused by mutations in translation initiation factor eIF2B and leading to progressive brain myelin deterioration, secondary axonal damage, and death in early adolescence. Eif2b5(R132H/R132H) mice exhibit delayed developmental myelination, mild early neurodegeneration and a robust remyelination defect in response to cuprizone-induced demyelination. In the current study we used Eif2b5(R132H/R132H) mice for mass-spectrometry analyses, to follow the changes in brain protein abundance in normal- versus cuprizone-diet fed mice during the remyelination recovery phase. Analysis of proteome profiles suggested that dysregulation of mitochondrial functions, altered proteasomal activity and impaired balance between protein synthesis and degradation play a role in VWM pathology. Consistent with these findings, we detected elevated levels of reactive oxygen species in mutant-derived primary fibroblasts and reduced 20S proteasome activity in mutant brain homogenates. These observations highlight the importance of tight translational control to precise coordination of processes involved in myelin formation and regeneration and point at cellular functions that may contribute to VWM pathology. Eif2b5(R132H/R132H) mouse model for vanishing white matter (VWM) disease was used for mass spectrometry of brain proteins at two time points under normal conditions and along recovery from cuprizone-induced demyelination. Comparisons of proteome profiles revealed the importance of mitochondrial function and tight coordination between protein synthesis and degradation to myelination formation and regeneration, pointing at cellular functions that contribute to VWM pathology.

PP09.10 – 2510: A novel case of periaxin-related neuropathy associated with cataract: Expanding the clinical and molecular spectrum of perxiaxin mutation

Objective Periaxin gene (PRX) is responsible for autosomal recessive early onset demyelinlating neuropathy: Charcot-Marie Tooth disease 4F (CMT4F) or its severe form Dejerine Sottas neuropathy (DSN). Periaxin protein is essential for maintenance of the peripheral nerve myelin probably in stabilizing its structure. It also functions as a scaffold protein that plays a role in lens membrane organization and cell adhesive interactions. Methods Exome sequencing. Results This report describes a child of non-consanguineous healthy parents with delayed motor milestones, frequent falls, and subcapsular cataract. His neurological exam revealed typical neuropathic features of the lower extremities: weakness of the lower limbs accompanied by absence of deep tendon reflexes. There was no sensory deficiency though he walked on abroad basis. Even so, his motor functions have improved over the years. Nerve conduction velocity (NCV) demonstrated demyelinating process with secondary axonal damage. Bilateral sub capsular cataract was discovered at one year old with no other ophthalmic pathology. A novel homozygote frame shift mutation c.418-419insG (P.m 149 fs) in the PRX gene was identified by whole exome sequencing and was confirmed by molecular analysis. Both his parents were found to be heterozygote carriers of this mutation. Conclusion Mutation in the periaxin protein is most probably the cause of peripheral distal neuropathy and cataract in this child. As of today, cataracts caused by mutations in the PRX gene were demonstrated only in laboratory animals. This is the first description of human congenital neuropathy coexistence with congenital cataract caused by mutations in the PRX gene. This is a new clinical presentation of PRX related hereditary neuropathy. It is suggested that careful eye examination should be done in every patients with congenital neuropathy. The coexistence of neuropathy and cataract may direct to the final diagnosis and its genetic cause.

Two faces of Schwann cell dedifferentiation in peripheral neurodegenerative diseases: pro-demyelinating and axon-preservative functions.

In vertebrates, Schwann cells are glial cells that form a myelin sheath, which produces the saltatory conduction of neural impulses in the peripheral nervous system (Sherman and Brophy, 2005). In the case of abnormal myelin sheath generation due to genetic mutations or myelin sheath destruction by acquired nerve damage induced by inflammation or drugs, problems arise in the peripheral nervous system, leading to muscle atrophy or sensory disorders (Sherman and Brophy, 2005; Jessen and Mirsky, 2008). During the embryonic period, the neural crest cells migrate along with growing peripheral axons. A Schwann cell precursor that has finished migrating and has attached to a growing nerve may wrap around a few or as many as dozens of axons, but ultimately only target a single axon that requires a myelin sheath in response to a certain stimulus (Sherman and Brophy, 2005; Jessen and Mirsky, 2008). In the period of Schwann cell generation, the cell is called Schwann cell progenitor or immature Schwann cell. Subsequently, the Schwann cell begins the process of myelin-related gene expression and wraps the axon with the elongated Schwann cell membrane, which continues throughout the postnatal period to complete the myelin sheath formation (Sherman and Brophy, 2005; Jessen and Mirsky, 2008). The mature myelin sheath will last a lifetime, except when damage, such as nerve injury, occurs. Acute and chronic demyelinating peripheral neuropathies are primary diseases that cause myelin destruction in adults (Jessen and Mirsky, 2008). When a peripheral nerve is physically severed, distal axons are completely degenerated, and the Schwann cells wrapped around the axons directly participate in the degeneration of the myelin sheath, which leads to demyelination (Jessen and Mirsky, 2008). Afterwards, macrophages that migrate from the blood assist in completely removing the degenerated axon and the myelin sheath (Martini et al., 2008). Meanwhile, in inflammatory demyelinating diseases, acute or chronic form of demyelination may occur regardless of axonal damage, and it is considered that demyelination occurs due to local inflammation caused by autoimmune mechanisms, which may attack components of the myelin sheath or junctional regions (Pollard and Armati, 2011).

A parkinsonian movement disorder with brain iron deposition and a haemochromatosis mutation

A 60-year-old, right-handed, Caucasian woman presented to the outpatient department with a three-month history of clumsiness and resting tremor in her right hand, associated with micrographia. She had no significant past medical history. Her family history included a brother who died at the age of 50 after a gastrointestinal bleed, attributed to alcoholic liver disease. Examination showed hypomimia, right hand resting tremor, and right arm and leg rigidity with bradykinesia. There was reduced arm swing during gait, and retropulsion on pull testing. The provisional diagnosis was idiopathic Parkinson’s disease. She was started on dopamine agonists and later on levodopa. However, the symptoms progressed rapidly, with worsening slowness and stiffness, development of postural hypotension and incomplete bladder emptying. She also reported daytime somnolence and snoring at night. As such, the diagnosis was revised to a Parkinson’s-plus syndrome: multiple system atrophy. Investigations revealed mildly raised serum alanine aminotransferase (63 IU/L), but the other liver function tests, full blood count, and copper studies (including serum ceruloplasmin) were all normal. No iron studies were performed. A 24-h cardiac monitor showed no arrhythmias. MRI-head (T1, T2, FLAIR and Gradient echo) at the time of presentation was normal. The patient did not attend for planned autonomic and respiratory function tests. Thirteen months after the first clinic appointment she died in her sleep. A post-mortem examination showed extraand intra-cellular deposition of iron in the globus pallidus, putamen, substantia nigra, locus coeruleus, red nuclei and dentate nuclei. This iron was visualised by Perl’s stain (Figs. 1 and 2), and was in fine granules, in moderate amounts. In addition, there was marked and diffuse neuronal loss from the substantia nigra and locus coeruleus with pale bodies within their neurons. However, no Lewy bodies were seen (alpha-synuclein negative). Axonal spheroids were present, dispersed through brain white matter tracts, consistent with secondary axonal damage. The liver was strongly positive for iron throughout (Perl’s stain). A DNA sample from post-mortem tissue was homozygous for the C282Y Hereditary Haemochromatosis mutation. This is the first case reported of a patient with a movement disorder and a haemochromatosis mutation, in which clear iron deposition in relevant parts of the brain has been seen at postmortem. Several idiopathic neurological conditions (including Parkinson’s disease) are associated with increased brain iron [1], but this is at the level of percentage changes in overall concentration, rather than the gross iron deposition seen in the case reported here. Very high brain iron does occur in conjunction with extrapyramidal disorders in neurodegeneration with brain iron accumulation (NBIA), a group of inherited diseases [2]. Our patient’s age, family history, liver iron, and C282Y mutation all go against this diagnosis. Hereditary haemochromatosis (HH) involves pathological iron deposition in a variety of organs [3]. However, central nervous system disorders are not traditionally listed among the sequelae of the disease. Previous postmortem studies in HH patients have shown excess iron in choroid plexus and pituitary, but not elsewhere in the brain [4], suggesting the blood brain barrier is protective. S. Williams (&) M. R. Vinjam A. Ismail A. Hassan Leeds Teaching Hospitals NHS Trust, Leeds, West Yorkshire, UK e-mail: stefanwilliams@doctors.org.uk

Long Term Follow-Up of Clinical and Electromyoneurographical Abnormalities in Eight Croatian Patients with Triple A Syndrome (P01.126)

Objective: To analyse long term follow up of neurological abnormalities (during 2 years to 23 years) in eight Croatian patients with triple A syndrome. Background The triple A syndrome is caused by autosomal recessively inherited mutation in AAAS gene on chromosome 12q13 encoding the nuclear pore protein ALADIN. This multisystem disease is characterized by achalasia, alacrimia, adrenal insufficiency and neurological impairment. Design/Methods: Clinical examination, electromyoneurography (EMNG) and molecular genetic analysis were performed in eight patients with triple A syndrome (five males and three females). Results: At the time of diagnosis all patients (aged 2 years to 8 years) presented with alacrimia, latent or manifest adrenal insufficiency, anisocoria in 2, optic atrophy in 4, motor and sensory polyneuropathy on the first exam in 2 siblings, muscle weakness and hypotrophy of distal muscle groups, cavus feet, hyperreflexia but absent triceps surae jerks, talocrural joints contractures, tremor and dysmetria in 5 patients. First EMG findings were normal at the age 6-11 years in 6/8 patients. The follow up EMG showed chronic partial denervation in all patients. Spontaneous activity (fibrillations) was registered in 3/8, compound muscle action potentials (CMAP) were polyphasic in 6/8 patients. Absent or low CMAP amplitude (0-0,5mV) was obtained in 6/8, decreased motor conduction velocity (29-40 m/s) in 8/8, absent F-wave potentials in 7/8, absent neural potentials in 5/8, and proximal conduction block in 5 patients. Mutation pSer263Pro was identified in 5 of 8 patients. One is homozygous and four are compound heterozygous for this mutation. Genotype/phenotype analysis confirmed lack of correlation in patients with triple A syndrome. Conclusions: Long term follow up neurography in patients with triple A syndrome showed progressive mixed motor and sensory polyneuropathy development with signs of pronounced demyelination and/or probably secondary axonal damage. Molecular results support the hypothesis that the pSer263Pro mutation is founder mutation in Slavic population. Disclosure: Dr. Barisic has nothing to disclose. Dr. Dumic has nothing to disclose. Dr. Kusec has nothing to disclose. Dr. Stingl has nothing to disclose. Dr. Lehman has nothing to disclose. Dr. Bunoza has nothing to disclose. Dr. Grdan has nothing to disclose. Dr. Koehler has nothing to disclose. Dr. Heubner has nothing to disclose.

Genetically Induced Adult Oligodendrocyte Cell Death Is Associated with Poor Myelin Clearance, Reduced Remyelination, and Axonal Damage

Loss of oligodendrocytes is a feature of many demyelinating diseases including multiple sclerosis. Here, we have established and characterized a novel model of genetically induced adult oligodendrocyte death. Specific primary loss of adult oligodendrocytes leads to a well defined and highly reproducible course of disease development that can be followed longitudinally by magnetic resonance imaging. Histological and ultrastructural analyses revealed progressive myelin vacuolation, in parallel to disease development that includes motor deficits, tremor, and ataxia. Myelin damage and clearance were associated with induction of oligodendrocyte precursor cell proliferation, albeit with some regional differences. Remyelination was present in the mildly affected corpus callosum. Consequences of acutely induced cell death of adult oligodendrocytes included secondary axonal damage. Microglia were activated in affected areas but without significant influx of B-cells, T-helper cells, or T-cytotoxic cells. Analysis of the model on a RAG-1 (recombination activating gene-1)-deficient background, lacking functional lymphocytes, did not change the observed disease and pathology compared with immune-competent mice. We conclude that this model provides the opportunity to study the consequences of adult oligodendrocyte death in the absence of primary axonal injury and reactive cells of the adaptive immune system. Our results indicate that if the blood-brain barrier is not disrupted, myelin debris is not removed efficiently, remyelination is impaired, and axonal integrity is compromised, likely as the result of myelin detachment. This model will allow the evaluation of strategies aimed at improving remyelination to foster axon protection.

Role of n‐type voltage‐dependent calcium channels in autoimmune optic neuritis

The aim of this study was to investigate the role of voltage-dependent calcium channels (VDCCs) in axon degeneration during autoimmune optic neuritis. Calcium ion (Ca(2+)) influx into the optic nerve (ON) through VDCCs was investigated in a rat model of optic neuritis using manganese-enhanced magnetic resonance imaging and in vivo calcium imaging. After having identified the most relevant channel subtype (N-type VDCCs), we correlated immunohistochemistry of channel expression with ON histopathology. In the confirmatory part of this work, we performed a treatment study using omega-conotoxin GVIA, an N-type specific blocker. We observed that pathological Ca(2+) influx into ONs during optic neuritis is mediated via N-type VDCCs. By analyzing the expression of VDCCs in the inflamed ONs, we detected an upregulation of alpha(1B), the pore-forming subunit of N-type VDCCs, in demyelinated axons. However, high expression levels were also found on macrophages/activated microglia, and lower levels were detected on astrocytes. The relevance of N-type VDCCs for inflammation-induced axonal degeneration and the severity of optic neuritis was corroborated by treatment with omega-conotoxin GVIA. This blocker led to decreased axon and myelin degeneration in the ONs together with a reduced number of macrophages/activated microglia. These protective effects were confirmed by analyzing the spinal cords of the same animals. We conclude that N-type VDCCs play an important role in inflammation-induced axon degeneration via two mechanisms: First, they directly mediate toxic Ca(2+) influx into the axons; and second, they contribute to macrophage/microglia function, thereby promoting secondary axonal damage. Ann Neurol 2009;66:81-93.

Mechanisms of axonal degeneration in EAE—lessons from CNTF and MHC I knockout mice

The major pathological hallmarks of multiple sclerosis (MS) comprise inflammation, demyelination with associated gliosis and axonal damage, which most likely correlates with persisting disability. Axonal damage can occur by several mechanisms. This article focuses on myelin disintegration and direct immune attack on axons by CD8-positive T-cells as two possible scenarios for axonal injury. As protoypic models, we investigated experimental autoimmune encephalomyelitis (EAE) in ciliary neurotrophic factor gene knockout mice (CNTF−/− mice) with severe myelin pathology and EAE in beta-2 microglobulin gene knockout mice (β2m−/− mice) lacking CD8-positive T-cells. The results from these studies indicate that the trigger attack for axonal injury even in a well-defined experimental design can be multi-facetted. No single factor seems to be absolutely necessary for the initiation of the process, but they rather act in concert and orchestrate tissue destruction, inflammation and regeneration. Some mechanisms of primary or secondary axonal damage may be shared between inflammatory and degenerative diseases of the nervous system, thereby establishing a link which might be of importance for future therapeutic strategies.