Chizu Wada

A novel de novo mutation in the desmin gene causes desmin myopathy with toxic aggregates

Objective: To determine the cause and pathogenic mechanisms of a 21-year-old patient’s cardioskeletal myopathy. The patient’s muscle atrophy and weakness began in distal parts of limbs; cardiac and facial muscles were later involved. Background: Desmin myopathy is a skeletal myopathy often associated with cardiomyopathy, caused by mutations in the desmin gene and characterized by desmin accumulation in affected muscle fibers, a leading marker of myofibrillar myopathies. Two kinds of deletions and seven missense mutations in the desmin gene have been identified. Methods: Clinical examination, electron microscopy of muscle tissue, two-dimensional gel electrophoresis, DNA sequencing, restriction enzyme analysis, and gene transfection were performed. Results: Electron microscopy showed disruption of sarcomeres at Z discs and electron-dense aggregates in biopsied skeletal and heart muscle. Two-dimensional gel electrophoresis of the patient’s skeletal muscle proteins showed massive accumulation of desmin. The authors identified a novel desmin mutation, L385P in one allele in the carboxyl end of the rod domain 2B in the patient’s leukocytes and skeletal muscle; neither parent had the mutation. Serologic study and DNA markers confirmed the de novo mutation. A peptide harboring desmin rod domains 2A and 2B with L385P tagged with green fluorescent protein induced cytoplasmic aggregates, nuclear DNA condensation, and cell death. Conclusions: A novel de novo mutation, L385P, causes desmin myopathy. An expression study indicated the toxic effect of the L385P mutation.

Kinesin and cytoplasmic dynein in spinal spheroids with motor neuron disease

Kinesin and cytoplasmic dynein are two major molecular motors responsible for fast axonal transport. As visualized by immunohistochemistry with monoclonal antibodies, both motors were found to be distributed throughout the cell bodies, dendrites and axons of motor neurons in normal human spinal cords. Large axonal swellings, spheroids, in the spinal cords of patients with motor neuron disease showed massive accumulation of kinesin co-localized with highly phosphorylated neurofilaments. Of 114 spheroids in five spinal cords, 87% were stained heavily with the three anti-kinesin antibodies used in this study. Cytoplasmic dynein was scarce or absent in most of the spheroids. These findings suggest that kinesin selectively accumulates in the spheroids of motor neuron axons, causing disturbance of the machinery for anterograde fast axonal transport in motor neuron disease.

Kinesin accumulation in chick spinal axonal swellings with β,β′-iminodipropionitrile (IDPN) intoxication

Abstract Kinesin is a major molecular motor responsible for anterograde axonal transport. Chicks were injected with β , β ′-iminodipropionitrile (IDPN) to induce axonal swellings in spinal motor neurons and spinal sensory ganglion neurons. Cylindrical swollen axons were found in the anterior horn and anterior funiculus of the spinal cord, anterior root, and spinal ganglia. All of the axonal swellings were heavily stained with two anti-kinesin monoclonal antibodies. The swellings were mildly stained with an anti-cytoplasmic dynein and anti-tubulin antibodies, and weakly stained with an anti-tau antibody. These suggest the isolated disturbance of kinesin transport with neurofilament accumulation in IDPN intoxication.

Purkinje Cell Loss in the Cerebellar Flocculus in Patients with Ataxia with Ocular Motor Apraxia Type 1/Early-Onset Ataxia with Ocular Motor Apraxia and Hypoalbuminemia

We genetically screened patients with ataxia with ocular motor apraxia type 1 (AOA1)/early-onset ataxia with ocular motor apraxia and hypoalbuminemia (EAOH), with a Japanese variant form of Friedreich’s ataxia. Three patients were found to have a homozygous insertion mutation of the aprataxin gene (689insT). An elder sister of a patient in this series died of cerebral hemorrhage at the age of 45, and underwent autopsy. In her cerebellar cortex, the mean density of Purkinje cells in the flocculus had predominantly decreased to 6.7% of normal controls, whereas the Purkinje cells in the other areas of the cerebellar hemisphere had decreased to 78.2%. This suggests that the cerebellar flocculus is the primary affected lesion in AOA1/EAOH, which should be associated with ocular motor apraxia.

Pontine Atrophy in Spinocerebellar Ataxia Type 6

To investigate the clinical range of spinocerebellar ataxia type 6 (SCA6), we screened CAG repeat expansion in the voltage-dependent alpha 1A calcium channel gene (CACNL1A4) in 71 ataxic patients in 60 families; 54 patients in 43 families with hereditary ataxia and 17 sporadic patients. Thirteen patients with SCA6 were detected to have elongated CAG in CACNL1A4. Of these, 7 patients had been diagnosed as having hereditary cerebellar cortical atrophy, and 6 patients had been found to have sporadic occurrence. One patient showed distinct pontine atrophy with prominent horizontal or oblique gaze nystagmus which is an unusual feature in sporadic olivopontocerebellar atrophy. For the efficient screening of SCA6, we would propose testing CAG repeat expansion in CACNL1A4, in patients with one of two markers: (1) horizontal or oblique gaze nystagmus without other eye movement disorders, (2) pure cerebellar atrophy, even if occurrence is sporadic. We should note that the pontine atrophy could also be caused by CAG repeat expansion in CACNL1A4.

Massive accumulation of M and H subunits of neurofilament proteins in spinal motor neurons of neurofilament deficient Japanese quail, Quv

Quiver (Quv) is a non-sense mutation of neurofilament protein L subunit (NF-L) that causes neurofilament deficiency with preserved microtubules in Japanese quail. Anti-NF-M and anti-NF-H mAbs stained cell bodies of motor neurons in Quv embryo spinal cords much more intense than those in control spinal cords. Volume of motor neurons in Quv spinal cords increased to 2.3 times of control motor neurons. Immunoblot of Quv spinal cords revealed a relative increase in non- and hypo-phosphorylated NF-M and NF-H, and a decrease in the total amount of NFs. Quv sciatic nerves showed faintly reacted phosphorylated NF-M and NF-H. These results suggest that deficiency of assembled neurofilament results in decreased axonal transport of NFs and accumulation of NFs in cell bodies of spinal motor neurons.

Acute encephalopathy associated with ingestion of a mushroom, Pleurocybella porrigens (angel's wing), in a patient with chronic renal failure

The authors report an autopsy case of acute encephalopathy in which generalized convulsion and coma occurred after ingestion of Pleurocybella porrigens (angels wing mushroom). The patient was a 65‐year‐old man who had undergone hemodialysis for 3 months due to chronic renal failure. Pathologic examination of the brain revealed extensive postinfarction‐like cystic necrosis in the bilateral putamens and multiple spotty necroses in the deep cerebral and cerebellar cortices. In 2004, similar acute encephalopathy related to ingestion of the mushroom was endemic in Japan, the pathogenesis of which remains to be elucidated.

Long-term follow up of thymus in patients with myasthenia gravis

We examined transversely the thymus of 33 myasthenia gravis (MG) patients followed up for more than 5 years and found three thymomas. One was found 21 years after thymoma resection (Masaoka I, WHO Type B2 thymoma) and extended thymectomy. The other two were non-thymomatous at onset, and they were not treated with extended thymectomy. Therapeutic guidelines should mention the importance of follow-up in MG thymus.

Approaches to Cough Peak Flow Measurement With Duchenne Muscular Dystrophy

BACKGROUND: Manually-assisted coughing and mechanical insufflation-exsufflation (MI-E) are commonly used in patients with Duchenne muscular dystrophy (DMD). Few studies have compared cough peak flow (CPF) with manually-assisted coughing to other methods, such as MI-E + manually-assisted coughing. In addition, few studies have reported the reliability of the measured CPF values. This study aimed to compare CPF with different cough-assistance methods and to examine the reliability of CPF data. METHODS: The study included 12 subjects with DMD (mean age, 34 ± 8 y). CPF, CPF + manually-assisted coughing (assisted CPF), maximum insufflation capacity (MIC) + CPF (MIC-CPF), MIC + manually-assisted coughing (MIC+assisted CPF), MI-E (MI-E–CPF), and MI-E + assisted CPF were measured. A spirometer was used to compare CPF measurements obtained with each of the cough-assist techniques. The reliability of the measured CPF values was analyzed using Bland-Altman analysis. RESULTS: CPF was 59 ± 34 L/min, assisted CPF was 113 ± 32 L/min, MIC-CPF was 170 ± 30 L/min, MIC+assisted CPF was 224 ± 62 L/min, MI-E–CPF was 199 ± 40 L/min, and MI-E + assisted CPF was 240 ± 38 L/min. A fixed and proportional bias was found in the CPF measurements made with the peak flow meter and the spirometer. The average 95% CI in the difference between peak flow meter, MI-E, and CPF obtained using the spirometer were −7.45 to −1.95 and −1.45 to 4.95, respectively. Test for correlation was r = 0.54 (P < .001) for CPF (peak flow meter) and CPF (spirometer) and r = 0.17 (P = .17) in CPF (MI-E) and CPF (spirometer), respectively. CONCLUSION: MI-E + assisted CPF was the highest. The CPF measured with the peak flow meter suggested underestimation.

Neurofilament proteins in spinal motor neurons and cultured cells in Quv, NF-L deficient quail

Quv is a non-sense mutation of neurofilament L subunit (NF-L) and lacks neurofilaments in nervous system. Accumulation of NF-H and NF-M immunoreactivity was found in swollen spinal motor neurons in 14-d embryo (E14). Numerous rough endoplasmic reticulum and G&j apparatus were found but no 10 nm filaments was observed in the cell bodies of motor neurons. CelI cuture of spinal cords in El4 Quv showed NF-H and NF-M immunoreactivity in cell bodies and axons but no 10 nm filaments. Relatively abundant microtubules were present in axons compared with control axons. These indicate the inability to constract 10 nm filaments in Quv embryo and cultured neurons. Normal size of cell bodies and the presence of immunoreactivity of NFs in axons of cultured spinal neurons suggest the difference in the transport of NFs between in uivo and in culture in Quv nervous system.