Toshihide Kumamoto

Expression of three calpain isoform genes in human skeletal muscles

Calpain is thought to be involved in muscular degradation in progressive muscular dystrophy (PMD), especially Duchenne and Becker muscular dystrophies. To assess the expression of calpain genes in skeletal muscles of patients with myopathies, we examined mRNA levels of three calpain isoforms by the quantitative reverse transcriptase-polymerase chain reaction method in biopsied muscles from control, PMD and amyotrophic lateral sclerosis (ALS) patients. There was a statistically significant increase in calpain 1 and calpain 2 mRNA levels in PMD and ALS patients as compared to controls. In contrast, there was a decrease in expression of calpain 3 mRNA in PMD, but it was not statistically significant. Expression of calpain 1 and calpain 2 positively correlated with each other, but not with calpain 3. These results indicate that expression of calpain 1 and calpain 2, but not calpain 3, are upregulated in diseased human muscles, likely playing a regulatory role in the process of myofibrillar degradation at the transcriptional as well as posttranslational level.

Prevalence Studies of Multiple Sclerosis, Myasthenia Gravis, and Myopathies in Kumamoto District, Japan

The prevalence ratio per 100,000 population of each disease was multiple sclerosis (MS) 1.3, myasthenia gravis (MG) 6.7, and polymyositis (PM) 5.0 in Kumamoto city, Japan (population 0.56 million, and the survey date on June 30, 1982). The prevalence ratio of MS was similar to that of 25 years ago in Kumamoto city. The incidence of MS had been stable despite Westernization of life-style in Japan. The prevalence ratio of MG and PM had increased moderately, possibly due to prolongation of life with recent progress in therapy. The prevalence ratio of all types of myopathies in Kumamoto prefecture (population: 1.8 million) on July 31, 1983, was estimated as approximately 17.4 per 100,000, for progressive muscular dystrophy (PMD) 4.1 per 100,000 population, compared to 3.9 for MG, and 2.4 for PM. The relative frequency of PMD, MG and PM was 23.3, 22.0 and 13.9%, respectively. The data shows that the prevalence ratio of various myopathies has not changed over the last 20 years in selected cities of Japan but the relative frequency of the diseases have changed due to recognition of the disease and prolongation of long life due to developments in diagnosis and treatment.

Exonic trinucleotide repeats and expression of androgen receptor gene in spinal cord from X-linked spinal and bulbar muscular atrophy.

We studied exonic trinucleotide repeats and expression of androgen receptor (AR) gene in the spinal cord from an autopsied patient with X-linked spinal and bulbar muscular atrophy (SBMA). Forty-nine CAG triplet repeats were found in tissues from the spinal cord, cerebrum, cerebellum, cardiac muscle and bladder, while there were 20-24 CAG repeats in these tissues from control subjects, consisting of three patients with amyotrophic lateral sclerosis (ALS) and three patients with lung cancer. Thus, mitotic instability of the AR gene in SBMA may not occur at the level of somatic cells. To determine whether expression of the AR gene in the spinal cord of SBMA differs from that in control subjects, we used quantitative reverse transcriptase (RT)-PCR and Western blot. AR mRNA and protein were detected in the spinal cord from the patient with SBMA, but the levels of both AR mRNA and protein were less than those from the patients with ALS in whom the loss of motor neurons was similar to findings in the patient with SBMA. These findings suggest that structural alteration plus a reduced level of AR in the spinal cord are involved in the pathogenesis of SBMA, resulting in degeneration of motor neurons.

Progressive myoclonus epilepsy dentato-rubro-pallido- luysian atrophy (DRPLA) in childhood.

A 22-year-old female with progressive myoclonus epilepsy (PME) considered to be due to hereditary dentato-rubro-pallido-luysian atrophy (DRPLA) was reported. Some of her family members showed progressive myoclonus, seizures, dementia, ataxia and choreoathetosis, with variation of onset from childhood to adult life, which suggested that they had been suffering from DRPLA. CT scan and MRI studies, including some on family members, revealed cerebral and cerebellar atrophy accompanied by dilatation of the fourth ventricle, compatible with the findings in DRPLA reported previously. We emphasize that a detailed family history may be essential in dealing with a PME patient and that DRPLA should be considered in the differential diagnosis of the PME syndrome with onset in childhood, in Japan.

Single muscle fiber analysis of myoclonus epilepsy with ragged-red fibers

We examined two muscle biopsy specimens from a proband and her mother with myoclonus epilepsy with ragged‐red fibers (MERRF), both obtained at an interval of about 10 years, using histochemistry, in situ hybridization, and single‐fiber polymerase chain reaction. Total (wild‐type and mutant) mitochondrial DNAs (mtDNAs) were greatly increased in ragged‐red fibers (RRF) over non‐RRF in all muscle specimens analyzed. The proportion of mutant mtDNA was also significantly higher in RRF than in non‐RRF. By comparing the first and second muscle biopsied specimens in each patient, we found that while the proportion of RRF, cytochrome c oxidase deficient fibers, and mutant DNA in muscle changed over a 10‐year period, the proportion of wild‐type and mutant mtDNAs in RRF and in non‐RRF was similar between the two specimens. These results suggest that the ratio of wild‐type to mutant mtDNAs in RRF and non‐RRF in MERRF is at a steady state level in each muscle fiber, without replicative advantage of mutant mtDNA.

Elevated soluble intercellular adhesion molecules‐1 in inflammatory myopathy

Introduction – We evaluated the serum level of soluble intercellular adhesion molecule‐1 (sICAM‐1) in patients with polymyositis (PM) and dermatomyositis (DM) and investigated the correlation between the serum level of sICAM‐1 and clinical findings. Material and methods – We measured the serum level of sICAM‐1 using an enzyme‐linked immunosorbent assay in 19 untreated patients with inflammatory myopathy (14 patients with PM and 5 patients with DM), 20 patients with other neuromuscular disorders in which immunological mechanisms are unlikely to be involved, and 14 normal healthy controls. Results – The serum level of sICAM‐1 was significantly higher in patients with PM/DM compared with patients with other neurological disorders and control subjects. The sICAM‐1 level was, however, not correlated with the clinical characteristic including disease severity, the duration of illness, and the serum level of CK. Conclusion – These findings suggest that sICAM‐1 is involved in the inflammatory process of PM and DM.

Experimental Chloroquine Myopathy: Morphological and Biochemical Studies

Morphological and biochemical studies were performed on the soleus muscles of rats receiving a daily intraperitoneal injection of 50 mg chloroquine chloride (CQ) per kilogram of body weight. Light mic

Hereditary cerebellar ataxia with Leber's hereditary optic neuropathy mitochondrial DNA 11778 mutation

We investigated a family with optic atrophy which occurred in childhood or early adulthood plus late-onset cerebellar ataxia. The magnetic resonance imaging in the proband revealed cerebellar atrophy. The proband and her brother were homoplasmic for the most common mitochondrial DNA (mtDNA) 11778 mutation associated with Lebers hereditary optic neuropathy (LHON). This study showed further evidence that central nervous system lesions can occur in cases of LHON mtDNA mutation.

A novel transthyretin mutation at position 30 (Leu for Val) associated with familial amyloidotic polyneuropathy

A novel transthyretin (TTR) mutation associated with familial amyloidotic polyneuropathy was detected in a Japanese patient. Single-strand conformation polymorphism analysis and sequence analysis of polymerase chain reaction (PCR)-amplified exons of the patients TTR gene revealed a point mutation resulting in a substitution of leucine for valine at position 30. As the mutation creates a Cfr13I site, it was confirmed by PCR and restriction analysis. Our finding indicates the importance of position 30 in TTR-derived amyloid fibril formation.

Clinical and genetic studies of spinocerebellar ataxia type 2 in Japanese kindreds

Objectives – We report the results of clinical and genetic studies from 2 related Japanese kindreds with spinocerebellar ataxia type 2 (SCA2). Material and methods – Family A showed 19 patients through 4 generations, while family B showed 6 patients, including dizygotic twin brothers, through 3 generations. We performed clinical, radiological, neurophysiological, and genetic analyses in the family members. Results – Neurologic analysis of 13 affected patients revealed a mean age at onset of 43.5 years. The most common neurologic finding was cerebellar ataxia with deep sensory disturbance. Slow saccades was found only in the younger patients below age 35 years. Nerve conduction studies revealed subclinical sensory neuropathy. Brain MRI showed the presence of pontocerebellar atrophy. Genetic study using PCR revealed that all affected patients had an expanded CAG allele in the ataxin‐2 gene, which led to a final diagnosis of SCA2. Conclusion – SCA2 may be more clinically heterogeneous than previously thought. PCR is useful in differentiating SCA2 from other types of inherited ataxia.