森田 光哉

Objective: To examine the possibility that CAG/CTG expansions are implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS). Methods: We have used the repeat expansion detection (RED) method to screen for CAG/CTG expansions in fifty-six cases of familial ALS (FALS), fifty-one cases of sporadic ALS (SALS) and fifty-one controls, We then tested the possibility that any detected expanded alleles originated from two specific loci, ERDA1/Dir1 or CTG18.1 using either the polymerase chain reaction (PCR) or Southern blotting. Results: CAG/CTG alleles with expansions of 144 bp or more base pairs were detected in 28.6% of FALS, 33.3% of SALS and 21.6% of controls. These alleles originated from either ERDA1/Dir1 or CTG18.1. Conclusion: The pathogenesis of familial and sporadic ALS does not commonly involve CAG/CTG expansions.

We have identified a novel mutation in exon 4 of the Cu/Zn superoxide dismutase (superoxide dismutase 1: SOD1) gene (GAC to GTC), which resulted in an Asp(90) to Val substitution in a Japanese family with amyotrophic lateral sclerosis (ALS) inherited as an autosomal dominant trait. The patients in this family usually died in 2-3 years without sensory or urinary impairment. The SOD1 activity was lower in the proband as compared to the normal controls. The clinical characteristics of this family resemble those of some patients heterozygous for the Asp90Ala mutation, but both the clinical features and SOD1 activity of this family differ from those of patients homozygous for the ASP90Ala mutation, Eur J Neurol 5:389-392 (C) 1998 Lippincott-Raven Publishers.

We studied the relationship between the number of CAG repeat units in the MJD1 gene and clinical features of Machado-Joseph disease (MJD) in eight patients from two generations of a Japanese MJD family. Because of lack of characteristic clinical signs of MJD such as dystonia, bulging eyes or facial myokymia, clinical diagnosis of MJD in this family was difficult to make prior to molecular testing for the CAG repeat expansion in the MJD1 gene. All the patients exhibited maternal transmission of MJD, and the intergenerational change in the number of CAG repeat units in the MJD1 gene was very small (+0.5+/-0.3, mean+/-S.E.M., n=4) in spite of marked genetic anticipation (-17.0 years/generation). In the present family, the degree of anticipation per repeat unit in maternal transmissions was much larger than that in maternal transmissions in the other six MJD families. This indicates that some maternal factors other than the increase of the number of CAG repeat units, which is known to be the basis of anticipation, may play a role in genetic anticipation in this MJD family. (C) 1998 Elsevier Science B.V.

We have identified a novel two-base mutation in exon 1 of the Cu/Zn superoxide dismutase (SOD1) gene (TGC to TTT), which resulted in Cys(6) to Phe substitution in a Japanese family with amyotrophic lateral sclerosis (ALS). This is the first case of familial ALS-associated two-base change of the SOD1 gene. Similar to several mutations in exon 1 of the SOD1 gene such as Ala(4) to Val, Ala(4) to Thr and Val(14) to Met, affected members of the present family showed a rapid progression of motor dysfunction. Although Ala(4), Cys(6) and Val(7) reside in the middle of the first beta-strand of the SOD1, a family with a mutation of Va1(7) to Glu associates with slow progression of the disease. These findings suggest that clinical courses are variable with each mutation, even in the same exon.

Cytotoxic action of leukocytes may be involved in the pathogenesis of inflammatory heart muscle disorders. We investigated the expression of rat leukocyte chemotactic cytokines-cytokine induced neutrophil chemoattractant (CINC) and JE-in cultured neonatal rat cardiac myocytes; CINC belongs to the interleukin 8 (IL-8) family and JE is a homologue of human monocyte chemoattractant protein 1 (MCP-1), In Northern blot analysis, CINC and JE transcripts were not clearly observed in unstimulated rat cardiac myocytes, However, their expression was clearly observed after exposure to tumour necrosis factor-alpha (TNF-alpha; 100 U/ml) for 26 h, We then evaluated IL-8 and MCP-I mRNA expression in human endomyocardial biopsy specimens from seven patients with idiopathic dilated cardiomyopathy by polymerase chain reaction analysis, Both IL-8 and MCP-1 mRNA transcripts were recognized in all patients studied, These results show for the first time that leukocyte chemotactic cytokines, IL-8 and MCP-1, are expressed in myocardial tissue, which might contribute to the pathogenesis of inflammatory heart muscle disorders.

A glucocorticoid, dexamethasone, inhibited the production of a leukocyte chemotactic cytokine, interleukin 8 (IL-8), as well as mRNA expression by a glioblastoma cell line, T98G, stimulated with interleukin 1 (IL-1). Dexamethasone also inhibited IL-8 promoter-driven chloramphenicol acetyltransferase (CAT) activities induced by IL-1, suggesting that dexamethasone inhibited IL-8 production mainly at the transcriptional level. Moreover, CAT assay revealed that the nuclear factor-kappa B (NF-kappa B) binding site was the crucial cis-element required for conferring IL-1 responsiveness in conjunction with the CCAAT enhancer binding protein/nuclear factor-IL-6 (NF-IL6) and/or the AP-1 binding site(s). Mutation of either the AP-1 or NF-IL6 binding site did not abolish IL-8 gene repression by dexamethasone, suggesting that these sites were not targets for dexamethasone. Trimerized kappa B sequence in the IL-8 gene was enough for conferring the induction by IL-1 and inhibition by dexamethasone of CAT activity. Finally, dexamethasone diminished the IL-l-induced formation of NF-kappa B complexes, which were identified immunochemically to consist of p50 and p65, without reducing the amount of translocated factors. Collectively, dexamethasone interfered with the binding of the most essential transcription factor, NF-kappa B, to its cognate cis-element, thereby suppressing the transcription of IL-8 gene.

In order to elucidate the role of inflammatory cytokines in the central nervous system, we examined the production of two leukocyte chemoattractants, IL-8 and monocyte chemotactic and activating factor (MCAF) in brain tumor cell lines. The glioma cell lines tested exhibited high levels of IL-g and MCAF mRNA expression upon stimulation with IL-1 or TNF-alpha, while none of the neuroblastoma cell lines expressed these cytokine mRNA. Both IL-8 and MCAF mRNA expression depended on the dose of IL-la and TNF-a and appeared very rapidly, reaching maximal levels at 3-6 hr, with substantial production of these cytokines in the culture supernatants. When various immunosuppressive drugs were tested, glucocorticoids but not other immunosuppressive drugs markedly inhibited the IL-1 or TNF-a-induced IL-8 and MCAF mRNA accumulation, suggesting that glucocorticoid is a potent regulator of these inflammatory cytokine production in the neural tissues. In addition, reverse transcription-polymerase chain reaction (RT-PCR) revealed the expression of IL-8 and MCAF mRNA expression in resected brain tumor tissues including glioblastoma, astrocytoma grade 2, ependymoma and medulloblastoma, indicating that these inflammatory cytokines are expressed in vivo.