Clinical characteristics and pathophysiological properties of newly discovered LRRK2 variants associated with Parkinson's disease.

Leucine-rich repeat kinase 2 (LRRK2) is the most common gene responsible for familial Parkinson's disease (PD). The gene product of LRRK2 contains multiple protein domains, including armadillo repeat, ankyrin repeat, leucine-rich repeat (LRR), Ras-of-complex (ROC), C-terminal of ROC (COR), kinase, and WD40 domains. In this study, we performed genetic screening of LRRK2 in our PD cohort, detecting sixteen LRRK2 rare variants. Among them, we selected seven variants that are likely to be familial and characterized them in terms of LRRK2 protein function, along with clinical information and one pathological analysis. The seven variants were S1120P and N1221K in the LRR domain; I1339M, S1403R, and V1447M in the ROC domain; and I1658F and D1873H in the COR domain. The kinase activity of the LRRK2 variants N1221K, S1403R, V1447M, and I1658F toward Rab10, a well-known phosphorylation substrate, was higher than that of wild-type LRRK2. LRRK2 D1873H showed enhanced self-association activity, whereas LRRK2 S1403R and D1873H showed reduced microtubule-binding activity. Pathological analysis of a patient with the LRRK2 V1447M variant was also performed, which revealed Lewy pathology in the brainstem. No functional alterations in terms of kinase activity, self-association activity, and microtubule-binding activity were detected in LRRK2 S1120P and I1339M variants. However, the patient with PD carrying LRRK2 S1120P variant also had a heterozygous Glucosylceramidase beta 1 (GBA1) L444P variant. In conclusion, we characterized seven LRRK2 variants potentially associated with PD. Five of the seven variants in different LRRK2 domains exhibited altered properties in kinase activity, self-association, and microtubule-binding activity, suggesting that each domain variant may contribute to disease progression in different ways.

Serum IgG anti-GD1a antibody and mEGOS predict outcome in Guillain-Barré syndrome.

OBJECTIVE: Approximately 15%-20% of patients with Guillain-Barré syndrome (GBS) are unable to walk independently at 6 months from the onset of neurological symptom. The modified Erasmus GBS outcome score (mEGOS) has been reported as a prognostic tool.Herein we investigated the association between a poor outcome, inability to walk independently at 6 months and presence of antiganglioside antibodies. METHODS: The clinical and serological data of 177 patients with GBS were retrospectively collected in Japan to assess the associations between a poor outcome and serum IgG antibodies against each ganglioside (GM1, GD1a, GalNAc-GD1a, GQ1b and GT1a). In addition, we investigated whether the combination of mEGOS and serum IgG antibodies against gangliosides is useful in predicting a poor outcome. RESULTS: The patients with IgG anti-GD1a antibodies more frequently showed poor outcomes than those without these antibodies (9 (36%) of 25 vs 8 (6%) of 127 patients, p<0.001). Particularly, 80% showed a poor outcome when they had both serum IgG anti-GD1a antibody and a high mEGOS of ≥10 on day 7 of admission. CONCLUSIONS: The combination of serum IgG anti-GD1a antibodies and a high mEGOS could help in making a more accurate prognosis of patients than mEGOS alone, especially for predicting poor outcomes.

Clavariopsins C-I, Antifungal Cyclic Depsipeptides from the Aquatic Hyphomycete Clavariopsis aquatica.

Seven new cyclic depsipeptides, clavariopsins C-I (3-9), together with two known congeners, clavariopsins A and B (1 and 2), were isolated from the aquatic hyphomycete Clavariopsis aquatica. Their planar structures, which consist of nine amino acids and one α-hydroxy acid, were elucidated by NMR spectroscopy and HRESIMS. The absolute configurations were established by the advanced Marfey's method and chiral-phase HPLC analysis. Their antifungal and cytotoxic activities were evaluated against six plant pathogenic fungi (Botrytis cinerea, Magnaporthe oryzae, Colletotrichum orbiculare, Fusarium oxysporum, Alternaria alternata, and Aspergillus niger) and a cancer cell line (HeLa-S3), respectively. The majority of the compounds exhibited potent antifungal activity against the fungi tested (minimum inhibition dose = 0.01-10 µg/disk) and induced hyphal swelling in A. niger (minimum effective dose = 0.3-3 µg/disk), whereas the compounds exhibited no cytotoxicity toward the cancer cell line. The results suggest that the clavariopsins could be a promising class of antifungal agents.

Guillain-Barré Syndrome.

Guillain-Barré syndrome (GBS) is an acute immune-mediated polyradiculoneuropathy, and pathophysiologically classified into acute inflammatory demyelinating polyneuropathy (AIDP), acute motor axonal neuropathy (AMAN), and acute motor and sensory axonal neuropathy (AMSAN). The main pathophysiological mechanism is complement-mediated nerve injury caused by antibody-antigen interaction in the peripheral nerves. Antiglycolipid antibodies are most pathogenic factors in the development of GBS, but not found in 40% of patients with GBS. One of the principal target regions in GBS is the node of Ranvier where functional molecules including glycolipids are assembled. Nodal dysfunction induced by the immune response in nodal axolemma, termed "nodopathy," can electrophysiologically show reversible conduction failure, axonal degeneration, or segmental demyelination. To detect new target molecules in antiglycolipid antibody-negative GBS and to elucidate the pathophysiology in the subacute and the subsequent phases of the disorder are the next problems.

Paranodal dissection in chronic inflammatory demyelinating polyneuropathy with anti-neurofascin-155 and anti-contactin-1 antibodies.

OBJECTIVE: To investigate the morphological features of chronic inflammatory demyelinating polyneuropathy (CIDP) with autoantibodies directed against paranodal junctional molecules, particularly focusing on the fine structures of the paranodes. METHODS: We assessed sural nerve biopsy specimens obtained from 9 patients with CIDP with anti-neurofascin-155 antibodies and 1 patient with anti-contactin-1 antibodies. 13 patients with CIDP without these antibodies were also examined to compare pathological findings. RESULTS: Characteristic light and electron microscopy findings in transverse sections from patients with anti-neurofascin-155 and anti-contactin-1 antibodies indicated a slight reduction in myelinated fibre density, with scattered myelin ovoids, and the absence of macrophage-mediated demyelination or onion bulbs. Teased-fibre preparations revealed that segmental demyelination tended to be found in patients with relatively higher frequencies of axonal degeneration and was tandemly found at consecutive nodes of Ranvier in a single fibre. Assessment of longitudinal sections by electron microscopy revealed that detachment of terminal myelin loops from the axolemma was frequently found at the paranode in patients with anti-neurofascin-155 and anti-contactin-1 antibody-positive CIDP compared with patients with antibody-negative CIDP. Patients with anti-neurofascin-155 antibodies showed a positive correlation between the frequencies of axo-glial detachment at the paranode and axonal degeneration, as assessed by teased-fibre preparations (p<0.05). CONCLUSIONS: Paranodal dissection without classical macrophage-mediated demyelination is the characteristic feature of patients with CIDP with autoantibodies to paranodal axo-glial junctional molecules.

Markers for Guillain-Barré syndrome with poor prognosis: a multi-center study.

Guillain-Barré syndrome (GBS) is an acute monophasic neuropathy. Prognostic tools include the modified Erasmus GBS outcome score (mEGOS), Erasmus GBS respiratory insufficiency score (EGRIS), and the increase in serum IgG levels (ΔIgG) 2 weeks after intravenous immunoglobulin (IVIg) treatment. Given that proportions of GBS subtypes differ between Western countries and Japan, the usefulness of these tools in Japan or other countries remains unknown. We enrolled 177 Japanese patients with GBS from 15 university hospitals and retrospectively obtained mEGOS and EGRIS for all and ΔIgG status for 79 of them. High mEGOS scores on admission or on day 7 were significantly associated with poorer outcomes (unable to walk independently at 6 months). High EGRIS scores (≥5 points) were associated with an increased risk for mechanical ventilation. Patients with ΔIgG <1,108 mg/dl had significantly poorer outcomes. We suggest that mEGOS, EGRIS, and ΔIgG in GBS are clinically relevant in Japan.

Efficacy of intravenous methylprednisolone pulse therapy in patients with multiple sclerosis and neuromyelitis optica.

BACKGROUND: No large-scale studies have compared the efficacy of intravenous methylprednisolone pulse therapy (IVMP) for multiple sclerosis (MS) and neuromyelitis optica (NMO). OBJECTIVE: To explain differences in treatment responses of MS and NMO patients to IVMP. METHODS: Changes in neurological symptoms/signs and Expanded Disability Status Scale (EDSS) scores before and within 1 week of IVMP completion were obtained in 2010 at 28 institutions, and retrospectively collated from 271 MS (478 courses) and 73 NMO (118 courses) cases. RESULTS: In MS patients, decreased EDSS score was significant after the first (-0.8 ± 0.9), second (-0.7 ± 0.9), and third (-0.7 ± 0.8) courses (p < 0.05), but not after the fourth (-0.3 ± 0.7) and fifth (-0.5 ± 0.6). However, decreased EDSS score was only significant after the first course (-0.5 ± 1.5, p < 0.05) in NMO patients. EDSS score was significantly decreased in MS compared with NMO patients at the first course (p < 0.05), but not thereafter. Model analysis for EDSS score improvement at the first course, adjusting for covariates, showed significantly greater decreases in MS compared with NMO patients (p < 0.05). CONCLUSION: IVMP is effective in MS from the first to third courses, and in NMO at the first course. Additionally, IVMP is more efficacious in MS than NMO patients, even at the first course.

Overlapping demyelinating syndromes and anti­N-methyl-D-aspartate receptor encephalitis.

OBJECTIVE: To report the clinical, radiological, and immunological association of demyelinating disorders with anti­Nmethyl- D-aspartate receptor (NMDAR) encephalitis. METHODS: Clinical and radiological analysis was done of a cohort of 691 patients with anti-NMDAR encephalitis. Determination of antibodies to NMDAR, aquaporin-4 (AQP4), and myelin oligodendrocyte glycoprotein (MOG) was performed using brain immunohistochemistry and cell-based assays. RESULTS: Twenty-three of 691 patients with anti-NMDAR encephalitis had prominent magnetic resonance imaging (MRI) and/or clinical features of demyelination. Group 1 included 12 patients in whom anti-NMDAR encephalitis was preceded or followed by independent episodes of neuromyelitis optica (NMO) spectrum disorder (5 cases, 4 anti-AQP4 positive) or brainstem or multifocal demyelinating syndromes (7 cases, all anti-MOG positive). Group 2 included 11 patients in whom anti-NMDAR encephalitis occurred simultaneously with MRI and symptoms compatible with demyelination (5 AQ4 positive, 2 MOG positive). Group 3 (136 controls) included 50 randomly selected patients with typical anti-NMDAR encephalitis, 56 with NMO, and 30 with multiple sclerosis; NMDAR antibodies were detected only in the 50 anti-NMDAR patients, MOG antibodies in 3 of 50 anti-NMDAR and 1 of 56 NMO patients, and AQP4 antibodies in 48 of 56 NMO and 1 of 50 anti-NMDAR patients (p<0.0001 for all comparisons with Groups 1 and 2). Most patients improved with immunotherapy, but compared with anti-NMDAR encephalitis the demyelinating episodes required more intensive therapy and resulted in more residual deficits. Only 1 of 23 NMDAR patients with signs of demyelination had ovarian teratoma compared with 18 of 50 anti-NMDAR controls (p50.011). INTERPRETATION: Patients with anti-NMDAR encephalitis may develop concurrent or separate episodes of demyelinating disorders, and conversely patients with NMO or demyelinating disorders with atypical symptoms (eg, dyskinesias, psychosis) may have anti-NMDAR encephalitis.

Bioactive Maleic Anhydrides and Related Diacids from the Aquatic Hyphomycete Tricladium castaneicola.

Four maleic anhydride derivatives, tricladolides A-D (1-4), and three alkylidene succinic acid derivatives, tricladic acids A-C (5-7), were isolated from the aquatic hyphomycete Tricladium castaneicola. The structures of these compounds were determined by spectroscopic analysis, and all were found to be novel. The compounds exhibited inhibitory activity against fungi, particularly Phytophthora sp., a plant pathogen of oomycetes. The inhibitory activity of these metabolites revealed the importance of the cyclic anhydride structure and the lipophilicity of the alkyl side chain. On the other hand, the cytotoxicity of the compounds against B16 melanoma cells indicated that the cyclic anhydride structure was not essential.

Study of the role of anaerobic metabolism in succinate production by Enterobacter aerogenes.

Succinate is a core biochemical building block; optimizing succinate production from biomass by microbial fermentation is a focus of basic and applied biotechnology research. Lowering pH in anaerobic succinate fermentation culture is a cost-effective and environmentally friendly approach to reducing the use of sub-raw materials such as alkali, which are needed for neutralization. To evaluate the potential of bacteria-based succinate fermentation under weak acidic (pH <6.2) and anaerobic conditions, we characterized the anaerobic metabolism of Enterobacter aerogenes AJ110637, which rapidly assimilates glucose at pH 5.0. Based on the profile of anaerobic products, we constructed single-gene knockout mutants to eliminate the main anaerobic metabolic pathways involved in NADH re-oxidation. These single-gene knockout studies showed that the ethanol synthesis pathway serves as the dominant NADH re-oxidation pathway in this organism. To generate a metabolically engineered strain for succinate production, we eliminated ethanol formation and introduced a heterogeneous carboxylation enzyme, yielding E. aerogenes strain ΔadhE/PCK. The strain produced succinate from glucose with a 60.5% yield (grams of succinate produced per gram of glucose consumed) at pH <6.2 and anaerobic conditions. Thus, we showed the potential of bacteria-based succinate fermentation under weak acidic conditions.

[Guillain-Barré syndrome].

Guillain-Barré syndrome (GBS), an acute immune-mediated neuropathy, occurs following immunological stimulation, such as infection, with complement-mediated neuropathy implicated in the pathophysiology of this condition. Glycolipid antibodies produced by molecular mimicry are detected in approximately 60% of cases. Recent studies have suggested the role of cell-mediated immunity in the pathogenesis of GBS. Intravenous immunoglobulin and plasma exchange are established immunotherapies. In this article, based on the latest knowledge, we describe the pathophysiology, diagnosis, treatment, prognosis, and prognostic prediction of GBS. Furthermore, we discuss some GBS guidelines published by the European Academy of Neurology/Peripheral Nerve Society.

Impact of Istradefylline on Levodopa Dose Escalation in Parkinson's Disease: ISTRA ADJUST PD Study, a Multicenter, Open-Label, Randomized, Parallel-Group Controlled Study.

INTRODUCTION: A higher levodopa dose is a risk factor for motor complications in Parkinson's disease (PD). Istradefylline (IST) is used as adjunctive treatment to levodopa in PD patients with off episodes, but its impact on levodopa dose titration remains unclear. The objective of this study was to investigate the effect of IST on levodopa dose escalation in PD patients with wearing-off. METHODS: This was a multicenter, open-label, randomized, parallel-group controlled study (ISTRA ADJUST PD) in which PD patients experiencing wearing-off (n = 114) who were receiving levodopa 300-400 mg/day were randomized to receive IST or no IST (control). Levodopa dose was escalated according to clinical severity. The primary endpoint was cumulative additional levodopa dose, and secondary endpoints were changes in symptom rating scales, motor activity determined by a wearable device, and safety outcomes. RESULTS: The cumulative additional levodopa dose throughout 37 weeks and dose increase over 36 weeks were significantly lower in the IST group than in the control group (both p < 0.0001). The Movement Disorder Society Unified Parkinson's Disease Rating Scale Part I and device-evaluated motor activities improved significantly from baseline to 36 weeks in the IST group only (all p < 0.05). Other secondary endpoints were comparable between the groups. Adverse drug reactions (ADRs) occurred in 28.8% and 13.2% of patients in the IST and control groups, respectively, with no serious ADRs in either group. CONCLUSION: IST treatment reduced levodopa dose escalation in PD patients, resulting in less cumulative levodopa use. Adjunctive IST may improve motor function more objectively than increased levodopa dose in patients with PD. TRIAL REGISTRATION: Japan Registry of Clinical Trials: jRCTs031180248.

A new maneuver for repetitive nerve stimulation testing in the trapezius muscle.

INTRODUCTION: The repetitive nerve stimulation (RNS) test in the trapezius muscle is used widely for the evaluation of myasthenia gravis. However, pseudofacilitation is often difficult to avoid in this muscle and may compromise the detection of small decremental responses. We have devised a new maneuver to reduce pseudofacilitation. METHODS: Using our maneuver, the shoulder of a supine subject is elevated passively and is held firmly by the examiner. Four conventional maneuvers as well as ours were compared with regard to pseudofacilitation that was maximal at the second wave in 14 control subjects. RESULTS: Pseudofacilitation at the second and fourth waves was the smallest using our maneuver. Up to 15% pseudofacilitation was observed using the other maneuvers. CONCLUSION: Pseudofacilitation in the trapezius muscle is mainly due to shortening of the muscle belly. It can be reduced greatly by shortening the muscle in advance.

Far-field potentials in hypothenar motor unit number estimation.

INTRODUCTION: Contamination by far-field potentials (FFPs) may interfere with motor unit number estimation (MUNE) in the ulnar nerve. METHODS: Surface motor unit potentials (SMUPs) from 29 spinal and bulbar muscular atrophy (SBMA) patients and 27 control subjects were classified into SMUPs from the abductor digiti minimi muscle (ADM SMUPs) or non-ADM SMUPs, based on the waveform patterns from 3-channel recordings. RESULTS: The mean areas of the ADM SMUPs and non-ADM SMUPs in control subjects were 219.0 ± 131.3 and 63.7 ± 48.5 µVms, respectively. In SBMA patients they were 1988.9 ± 999.4 and 222.7 ± 125.7 µVms, respectively. The percentages of non-ADM SMUPs were 68 ± 22% in controls and 84 ± 15% in SBMA patients. CONCLUSIONS: Non-ADM SMUPs generated mainly by FFPs often had a negative onset in the routine lead and were indistinguishable from ADM SMUPs. More frequent exclusion of smaller non-ADM SMUPs in controls by size criteria would reduce the diagnostic yield of MUNE.

[Antiganglioside complex antibody].

Antiganglioside antibodies are known to play a pathogenic role in development of Guillain-Barré syndrome (GBS) and Fisher syndrome (FS). Recently, antibodies to ganglioside complexes (GSCs) consisting of two different gangliosides were found in some patients with GBS and FS. Some anti-GSC antibodies such as anti-GD1a/GD1b, anti-GM1/GQ1b or anti-GM1/GalNAc-GD1a antibodies correlate with specific clinical features. Anti-GSCs antibodies can be found also in chronic inflammatory demyelinating polyneuropathy and neuropathy associated with IgM monoclonal gammopathy. Latest studies have revealed molecular basis of anti-GSCs antibody-mediated nerve injury. The concept of GSCs will provide new vistas on understanding of pathogenesis of GBS and related autoimmune diseases.

Guillain-Barré Syndrome Mimicking Lumbar Spinal Stenosis with Segmental Weakness in L5-S1 Myotomes: Two Case Reports.

We herein report two cases of Guillain-Barré syndrome (GBS) mimicking lumbar spinal stenosis (LSS). Both cases were initially diagnosed as LSS based on prominent segmental weakness in the L5 and S1 myotomes and coexisting LSS on magnetic resonance imaging. However, neurological and electrophysiological examinations revealed abnormalities that extended to the upper extremities, although slight, prompting us to suspect GBS. Subsequently, serum antiganglioside antibodies and remarkable responsiveness to intravenous immunoglobulin therapy confirmed GBS. We suspect that the focal blood-nerve barrier disruption due to preexisting LSS might have contributed to the segmental weakness in this atypical GBS case.

Anti-GM1/GD1a complex antibodies in GBS sera specifically recognize the hybrid dimer GM1-GD1a.

It is now emerging the new concept that the antibodies from some patients with Guillain-Barré syndrome (GBS) recognize an antigenic epitope formed by two different gangliosides, a ganglioside complex (GSC). We prepared the dimeric GM1-GD1a hybrid ganglioside derivative that contains two structurally different oligosaccharide chains to mimic the GSC. We use this compound to analyze sera from GBS patients by high-performance thin-layer chromatography immunostaining and enzyme-linked immunosorbent assay. We also synthesized the dimeric GM1-GM1 and GD1a-GD1a compounds that were used in control experiments together with natural gangliosides. The hybrid dimeric GM1-GD1a was specifically recognized by human sera from GBS patients that developed anti-oligosaccharide antibodies specific for grouped complex oligosaccharides, confirming the information that GBS patients developed antibodies against a GSC. High-resolution (1)H-(13)C heteronuclear single-quantum coherence-nuclear overhauser effect spectroscopy nuclear magnetic resonance experiments showed an interaction between the IV Gal-H1 of GM1 and the IV Gal-H2 of GD1a suggesting that the two oligosaccharide chains of the dimeric ganglioside form a single epitope recognized by a single-antibody domain. The availability of a method capable to prepare several hybrid gangliosides, and the availability of simple analytical approaches, opens new perspectives for the understanding and the therapy of several neuropathies.

Nationwide survey of patients in Japan with Bickerstaff brainstem encephalitis: epidemiological and clinical characteristics.

BACKGROUND: Information regarding the epidemiological background of Bickerstaff brainstem encephalitis (BBE) is limited. METHODS: We conducted a nationwide survey of BBE in the Japanese population in two steps: the first aimed to identify patients with brainstem encephalitis for the specified 3 year period and the second to evaluate whether the clinical picture met our diagnostic criteria for BBE. RESULTS: The number of patients with brainstem encephalitis was estimated as 704 (95% CI 478 to 930) over the 3 years. The annual onset of BBE was roughly estimated as 100 cases, which accounted for 43% of brainstem encephalitis. BBE was slightly male predominant and often young onset. Among brainstem encephalitis patients, BBE was characterised by antecedent infectious symptoms, oropharyngeal palsy and sensory disturbance at the distal extremities with absent or decreased tendon reflexes, in addition to a triad of symptoms (external ophthalmoplegia, ataxia and impaired level of consciousness) and shorter duration to the peak, with good outcome. Anti-GQ1b antibodies were present in 75% of cases. Several BBE patients with atypical clinical features or without anti-GQ1b antibodies were also identified. These cases often had marked CSF pleocytosis, abnormal brain MRI findings and a longer duration to peak symptoms, sometimes with considerable residual deficits. CONCLUSIONS: BBE is a rare disorder but accounts for a major proportion of brainstem encephalitis. BBE consists of typical and atypical cases. Typical BBE has similar neurological and serological features to Fisher syndrome and shows good recovery whereas atypical BBE is characterised by delayed recovery, negative anti-GQ1b antibodies, and abnormal CSF and brain MRI findings with other possible pathogeneses.

[CIDP Refractory to Three Types of Mainstay Treatments: Appropriate Induction Therapy and Evaluation of Therapeutic Effect].

Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is a disease with a heterogeneous pathology. The responsiveness to mainstay treatment differs depending on the type of CIDP. The treatment strategy is determined based on the type of CIDP, characteristics of the therapeutic agents and treatment methods, and patient background. For CIDPs that do not respond to the mainstay treatment, it is necessary to review whether the induction treatment was properly performed and whether the therapeutic effect was properly evaluated using objective indicators. In particular, treatment-resistant multifocal CIDP and distal CIDP require careful differential diagnoses. Intervention trials using rituximab and anti-neonatal Fcγ receptor monoclonal antibodies are underway, and it is expected that clear treatment guidelines will soon be developed for refractory CIDP.

[Epstein-Barr virus-associated subacute sensorimotor neuropathy with multiple cerebellar microbleeding: a case report].

We report the case of an 82-year-old male with subacute sensorimotor neuropathy associated with Epstein-Barr virus (EBV) infection, who presented with diplopia followed by gait disturbance due to limb weakness. Pathological findings of a biopsied cervical lymph node showed a high frequency of EBV-positive cells. EBV-DNA was detected in blood. A nerve conduction study suggested a mixture of axonal damage and demyelination. Brain MRI showed multiple microbleeds in cerebellar cortices, but cerebrospinal fluid EBV-PCR was negative, suggesting bleeding due to EBV-related vasculitis. Corticosteroid therapy improved the neurological symptoms and the patient was able to walk independently four months later. The main pathogenesis of the neuropathy in this case is likely to be a mixture of vasculitic neuropathy and immune-mediated demyelinating neuropathy, which are considered to be due to EBV reactivation.