Ayumi Nishiyama

GNE myopathy associated with congenital thrombocytopenia: A report of two siblings

GNE myopathy is an autosomal recessive muscular disorder caused by mutations in the gene encoding the key enzyme in sialic acid biosynthesis, UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE/MNK). Here, we report two siblings with myopathy with rimmed vacuoles and congenital thrombocytopenia who harbored two compound heterozygous GNE mutations, p.V603L and p.G739S. Thrombocytopenia, which is characterized by shortened platelet lifetime rather than ineffective thrombopoiesis, has been observed since infancy. We performed exome sequencing and array CGH to identify the underlying genetic etiology of thrombocytopenia. No pathogenic variants were detected among the known causative genes of recessively inherited thrombocytopenia; yet, candidate variants in two genes that followed an autosomal recessive mode of inheritance, including previously identified GNE mutations, were detected. Alternatively, it is possible that the decreased activity of GNE/MNK itself, which would lead to decreased sialic content in platelets, is associated with thrombocytopenia in these patients. Further investigations are required to clarify the association between GNE myopathy and the pathogenesis of thrombocytopenia.

Comprehensive targeted next-generation sequencing in Japanese familial amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disease characterized by loss of motor neurons. We have recently identified SOD1 and FUS mutations as the most common causes in a consecutive series of 111 familial ALS pedigrees in Japan. To reveal possible genetic causes for the remaining 51 patients with familial ALS (45 pedigrees), we performed targeted next-generation sequencing of 35 known ALS/motor neuron diseases-related genes. Known variants in ANG, OPTN, SETX, and TARDBP were identified in 6 patients. A novel likely pathogenic homozygous variant in ALS2 was identified in 1 patient. In addition, 18 patients harbored 1-3 novel variants of uncertain significance, whereas hexanucleotide repeat expansions in C9ORF72 were not detected using repeat-primed polymerase chain reaction. Collectively, in our Japanese cohort, the frequencies of SOD1, FUS, SETX, TARDBP, ANG, and OPTN variants were 32%, 11%, 2%, 2%, 1%, and 1%, respectively. These findings indicate considerable differences in the genetic variations associated with familial ALS across populations. Further genetic analyses and functional studies of novel variants are warranted.

Postural leg tremor in X-linked spinal and bulbar muscular atrophy

X-linked spinal and bulbar muscular atrophy (SBMA) is an adult-onset neuromuscular disorder caused by a CAG repeat expansion in the androgen receptor gene. Postural hand tremor is well known as a non-motor neuron sign, but to our knowledge postural leg tremor has not been reported. We studied the occurrence and physiological features of postural leg tremor in 12 male patients (38-64 years old) with genetically proven SBMA. Three patients had postural leg tremor with a frequency of 4-7Hz. In these patients, sensory nerve action potential (SNAP) was not detected in the lower limbs. There were significant differences between the patients with postural leg tremor and those without postural leg tremor in both the SNAP of the sural nerve and the length of the CAG repeat. Phenotypical differences between shorter CAG repeats, which indicate a sensory-dominant phenotype, and longer CAG repeats, which indicate a motor-dominant phenotype, have been previously reported. In the present study, 60% of patients with shorter CAG repeats (<47) showed leg tremor and none of the patients with longer CAG repeats (≥47) did. Postural leg tremor could be a clinical feature that predicts shorter CAG repeats of the androgen receptor gene.

Isolated inclusion body myopathy caused by a multisystem proteinopathy–linked hnRNPA1 mutation

Objective: To identify the genetic cause of isolated inclusion body myopathy (IBM) with autosomal dominant inheritance in 2 families. Methods: Genetic investigations were performed using whole-exome and Sanger sequencing of the heterogeneous nuclear ribonucleoprotein A1 gene (hnRNPA1). The clinical and pathologic features of patients in the 2 families were evaluated with neurologic examinations, muscle imaging, and muscle biopsy. Results: We identified a missense p.D314N mutation in hnRNPA1, which is also known to cause familial amyotrophic lateral sclerosis, in 2 families with IBM. The affected individuals developed muscle weakness in their 40s, which slowly progressed toward a limb-girdle pattern. Further evaluation of the affected individuals revealed no apparent motor neuron dysfunction, cognitive impairment, or bone abnormality. The muscle pathology was compatible with IBM, lacking apparent neurogenic change and inflammation. Multiple immunohistochemical analyses revealed the cytoplasmic aggregation of hnRNPA1 in close association with autophagosomes and myonuclei. Furthermore, the aberrant accumulation was characterized by coaggregation with ubiquitin, sequestome-1/p62, valosin-containing protein/p97, and a variety of RNA-binding proteins (RBPs). Conclusions: The present study expands the clinical phenotype of hnRNPA1-linked multisystem proteinopathy. Mutations in hnRNPA1, and possibly hnRNPA2B1, will be responsible for isolated IBM with a pure muscular phenotype. Although the mechanisms underlying the selective skeletal muscle involvement remain to be elucidated, the immunohistochemical results suggest a broad sequestration of RBPs by the mutated hnRNPA1.

Aberrant astrocytic expression of chondroitin sulfate proteoglycan receptors in a rat model of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease. Progressive and systemic loss of motor neurons with gliosis in the central nervous system (CNS) is a neuropathological hallmark of ALS. Chondroitin sulfate proteoglycans (CSPGs) are the major components of the extracellular matrix of the mammalian CNS, and they inhibit axonal regeneration physically by participating to form the glial scar. Recently, protein tyrosine phosphatase sigma (PTPσ) and leukocyte common antigen‐related protein were discovered as CSPG receptors that play roles in inhibiting regeneration. Here we examined the expression of CSPG receptors in transgenic female rats overexpressing an ALS‐linked mutant cytosolic Cu/Zn superoxide dismutase gene (SOD1). In contrast to controls, multiple immunofluorescence analyses revealed aberrant expression of CSPG receptors dominantly in reactive astrocytes, while PTPσ expression in neurons decreased in the spinal ventral horns of ALS transgenic rats. The aberrant and progressive astrocytic expression of CSPG receptors and reactive astrocytes themselves may be therapeutic targets for reconstructing a regeneration‐supportive microenvironment under neurodegenerative conditions such as ALS.

Genotype-phenotype relationships in familial amyotrophic lateral sclerosis with FUS/TLS mutations in Japan.

Introduction: We investigated possible genotype–phenotype correlations in Japanese patients with familial amyotrophic lateral sclerosis (FALS) carrying fused in sarcoma/translated in liposarcoma (FUS/TLS) gene mutations. Methods: A consecutive series of 111 Japanese FALS pedigrees were screened for copper/zinc superoxide dismutase 1 (SOD1) and FUS/TLS gene mutations. Clinical data, including onset age, onset site, disease duration, and extramotor symptoms, were collected. Results: Nine different FUS/TLS mutations were found in 12 pedigrees. Most of the patients with FUS/TLS‐linked FALS demonstrated early onset in the brainstem/upper cervical region, and relatively short disease duration. A few mutations exhibited phenotypes that were distinct from typical cases. Frontotemporal dementia was present in 1 patient. Conclusions: This study revealed a characteristic phenotype in FUS/TLS‐linked FALS patients in Japan. FUS/TLS screening is recommended in patients with FALS with this phenotype. Muscle Nerve 54: 398–404, 2016

Prominent sensory involvement in a case of familial amyotrophic lateral sclerosis carrying the L8V SOD1 mutation

• We present a case of familial amyotrophic lateral sclerosis carrying the L8V SOD1 mutation with novel clinical manifestations.

A case of steroid-responsive MADSAM with late appearance of a partial conduction block in the forearm.

Multifocal acquired demyelinating sensory and motor neuropathy (MADSAM) [1] is defined as an asymmetrical presentation of chronic inflammatory demyelinating polyneuropathy (CIDP). Nerve conduction studies (NCS) in MADSAM show characteristic demyelinating changes such as conduction block and temporal dispersion. We report a case of MADSAM with only F-waves absent three months after clinical onset of the disease and partial motor conduction blocks in the left forearm for the first time seven months after onset. A 63-year-old male was admitted to our hospital in January 2012 because of progressive muscle weakness of the right leg from three months before, dysesthesia of the left hand and the right foot from two months before, and bilateral weakness in the arms from one month before. Neurological examination on admission revealed asymmetrical weakness in all four limbs, with generalized areflexia and superficial sensory disturbance in the left hand and both feet. The left medial, right femoral, and left deep peroneal nerves were affected clinically, and bilateral effects were also observed in the ulnar, radial, and sciatic nerves. Blood cell counts and biochemistry including autoantibodies were normal. Cerebrospinal fluid examination showed an elevated protein concentration (92 mg/dl) without pleocytosis (2/ll). NCS showed no demyelination such as temporal dispersion or conduction block in the median nerve between the wrist and axilla (Fig. 1a), the ulnar nerve between the wrist and above the elbow (Fig. 1b), or the peroneal and tibial nerves between the ankle and knee, but F-waves were absent in the bilateral median and the left ulnar and tibial nerves. No compound muscle action potential (CMAP) was evoked with bilateral stimulation at Erb’s point. Whole spine MRI showed no remarkable abnormal findings. In May 2012, the weakness and dysesthesia in the left leg had increased gradually and the patient could not walk unassisted. NCS revealed partial motor conduction blocks in the left median (Fig. 1c) and ulnar (Fig. 1d) nerves in the forearm in addition to absent F-waves in the bilateral median and the left ulnar and tibial nerves. The patient was diagnosed with MADSAM based on the European Federation of Neurological Societies and the Peripheral Nerve Society (EFNS/PNS) criteria [1]. Intravenous immunoglobulin (IVIG) treatment (400 mg/ kg/day for five days) administered three times was ineffective, but his symptoms improved dramatically after intravenous corticosteroid pulse therapy (methylprednisolone 1 g/day for three days) followed by oral corticosteroids (prednisolone 60 mg/day) (Fig. 2). In July 2012, F-waves appeared partially in the left median nerve. In March 2013, the patient was treated with oral prednisolone 7 mg/day and could walk with a cane by himself. Hyperproteinorrhachia, the absent F-waves, and the lack of abnormal demyelinating findings in the distal limbs below the axilla or knee suggest that demyelination might S. Yoshida A. Kikuchi (&) M. Tateyama O. Tano A. Nishiyama T. Akaishi M. Kato M. Aoki Department of Neurology, Tohoku University Graduate School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8574, Japan e-mail: akikuchi@med.tohoku.ac.jp

TARDBP p.G376D mutation, found in rapid progressive familial ALS, induces mislocalization of TDP-43

Amyotrophic lateral sclerosis (ALS) is a fatal motor neuron disease which causes the progressive weakness and atrophy of limbs and respiratory muscles, resulting in death within 3–5 years of onset. Targeted next-generation sequencing has enabled a comprehensive analysis of ALS disease-causing genes [1]. However, a method to confirm the pathogenicity of variants of unknown significance (VUS) remains to be elucidated. The previously reported variants require validation if they are not isolated in the designated family members. For example, we have reported the first Asian familial ALS case with TARDBP p.G376D mutation [1], which was previously reported as a familial ALS-linked mutation in European familial cases [2,3]. In the present study, we focused on TARDBP and the detailed clinical profile of the patient. We also examined TARDBP p.G376D pathogenicity using overexpression cell models.

Antagonizing bone morphogenetic protein 4 attenuates disease progression in a rat model of amyotrophic lateral sclerosis

Abstract Amyotrophic lateral sclerosis (ALS) is an adult‐onset, fatal neurodegenerative syndrome characterized by the systemic loss of motor neurons with prominent astrocytosis and microgliosis in the spinal cord and brain. Astrocytes play an essential role in maintaining extracellular microenvironments that surround motor neurons, and are activated by various insults. Growing evidence points to a non‐cell autonomous neurotoxicity caused by chronic and sustained astrocytic activation in patients with neurodegenerative diseases, including ALS. However, the mechanisms that underlie the harmful effects of astrocytosis in patients with ALS remain unresolved. We focused on bone morphogenetic proteins as a major soluble factor that promotes astrocytogenesis and its activation in the adult spinal cord. In a transgenic rat model with ALS‐linked mutant Cu/Zn superoxide dismutase gene, BMP4 was progressively up‐regulated in reactive astrocytes of the spinal ventral horns, whereas the BMP‐antagonist noggin was decreased in association with neuronal degeneration. Continuous intrathecal noggin supplementation after disease onset significantly ameliorated motor dysfunction symptoms, neurogenic muscle atrophy, and extended survival of symptomatic ALS model rats, despite lack of deterrence against neuronal death itself. The exogenous noggin inhibited astrocytic hypertrophy, astrocytogenesis, and neuroinflammation by inactivating both Smad1/5/8 and p38 mitogen‐activated protein kinase pathways. Moreover, intrathecal infusion of a Bmp4‐targeted antisense oligonucleotides and provided selective Bmp4 knockdown in vivo, which suppressed astrocyte and microglia activation, reproducing the aforementioned results by noggin treatment. Collectively, we clarified the involvement of BMP4 in the processes of excessive gliosis that exacerbate the disease progression of the ALS model rats. Our study demonstrated that BMP4, with its downstream signaling, might be a novel therapeutic target for disease‐modifying therapies in ALS. HighlightsBMP4 increased in astrocytes of a rat model of amyotrophic lateral sclerosis (ALS).Inhibiting the BMP4 attenuated the disease progression via suppressing gliosis.The gliosis‐promoting BMP4 may become a therapeutic target in ALS.