Kenichi Yasui

The homozygous C677T mutation in the methylenetetrahydrofolate reductase gene is a genetic risk factor for migraine

Increased homocysteine levels are associated with various pathological conditions in humans, including stroke and cardiovascular disorders. Homocysteine acts as an excitatory amino acid in vivo and may influence the threshold of migraine headache. Frosst et al. [1995] reported an association between the homozygous C677T mutation in the 5,10-methylenetetrahydrofolate reductase (MTHFR) gene and serum homocysteine levels. This study was designed to determine the prevalence of the MTHFR mutation in Japanese patients with migraine and tension-type headache (TH). Seventy-four patients with migraine headaches (22 with aura and 52 without aura), 47 with THs, and 261 normal controls were recruited. Genotyping of MTHFR C677T polymorphism was performed by polymerase chain reaction-restriction fragment length polymorphism. We detected that the incidence of the homozygous transition (T/T) in migraine sufferers (20.3%) was significantly higher than that in controls (9.6%). Moreover, the frequency of the T/T genotype in individuals with migraine headaches with aura was remarkably high (40.9%). The MTHFR T allele was more frequent in the migraine group than in the control group. Our results support the conclusion that the MTHFR gene, causing mild hyperhomocysteinemia may be a genetic risk factor for migraine. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:762-764, 2000.

Mutations in COQ2 in familial and sporadic multiple-system atrophy the multiple-system atrophy research collaboration

BACKGROUND Multiple-system atrophy is an intractable neurodegenerative disease characterized by autonomic failure in addition to various combinations of parkinsonism, cerebellar ataxia, and pyramidal dysfunction. Although multiple-system atrophy is widely considered to be a nongenetic disorder, we previously identified multiplex families with this disease, which indicates the involvement of genetic components. METHODS In combination with linkage analysis, we performed whole-genome sequencing of a sample obtained from a member of a multiplex family in whom multiple-system atrophy had been diagnosed on autopsy. We also performed mutational analysis of samples from members of five other multiplex families and from a Japanese series (363 patients and two sets of controls, one of 520 persons and one of 2383 persons), a European series (223 patients and 315 controls), and a North American series (172 patients and 294 controls). On the basis of these analyses, we used a yeast complementation assay and measured enzyme activity of parahydroxybenzoate-polyprenyl transferase. This enzyme is encoded by the gene COQ2 and is essential for the biosynthesis of coenzyme Q10. Levels of coenzyme Q10 in lymphoblastoid cells and brain tissue were measured on high-performance liquid chromatography. RESULTS We identified a homozygous mutation (M78V-V343A/M78V-V343A) and compound heterozygous mutations (R337X/V343A) in COQ2 in two multiplex families. Furthermore, we found that a common variant (V343A) and multiple rare variants in COQ2, all of which are functionally impaired, are associated with sporadic multiple-system atrophy. The V343A variant was exclusively observed in the Japanese population. CONCLUSIONS Functionally impaired variants of COQ2 were associated with an increased risk of multiple-system atrophy in multiplex families and patients with sporadic disease, providing evidence of a role of impaired COQ2 activities in the pathogenesis of this disease. (Funded by the Japan Society for the Promotion of Science and others.).

Plasma homocysteine and MTHFR C677T genotype in levodopa-treated patients with PD

To the Editor: The paper by Yasui et al.1 showing elevated plasma homocysteine levels in levodopa-treated patients with PD is interesting and confirms other reports2 and our unpublished findings (table). It also shows that patients with the TT-genotype of the C677T polymorphism of methylenetetrahydrofolate reductase (MTHFR) have especially high homocysteine levels. These findings have to be related to the metabolism of levodopa and methyl groups and may have importance in the future treatment of PD with levodopa. Homocysteine is the demethylated derivative of methionine and is only formed in numerous transmethylation reactions (e.g., catecholamine O-methyltransferase [COMT]) that use S-adenosylmethionine (SAM) as the methyl donor.3 Most administered levodopa is COMT-methylated to 3O-methyldopa and the methylation of 1000 mg of levodopa consumes 5 mMol of SAM methyls and produces 5 mMol of S-adenosylhomocysteine (SAH), which then is deadenosylated to homocysteine. In the study of Yasui et al., the average levodopa dose was 1500 mg. Therefore, it is most likely that levodopa treatment contributed to hyperhomocysteinemia in their patients with PD. In this context it is important to note that humans normally consume about 15 mMol/day of SAM methyls for methylation of endogenous compounds such as phospholipids, DNA, myeline, and proteins. About 10 mMol of these methyls are supplied from food, mainly as methionine and choline, and about 5 mMol are de novo synthesized in one-carbon (folate) metabolism and are transferred by a vitamin B12-dependent reaction to homocysteine, again forming methionine and SAM.4 As we can assume that levodopa-treated PD patients do not increase their intake of labile methyls, they must de novo synthesize all their extra methyls consumed for levodopa methylation. In the above 1000 mg example, the patient must increase his or her de novo synthesis of methyls by about 100%. If methyl neogenesis is hampered by poor folate intake, especially if the patient is a TT-homozygote for C677T/MTHFR polymorphism (about 12% of the Caucasian population), or has a low vitamin B12 level, production of SAM methyls may not be sufficient for both methylation of endogenous compounds and levodopa. This may lead to hypomethylation of essential neural components, and might explain the neuropsychiatric symptoms in folate and vitamin B12 deficiency.5 This mechanism may contribute to side effects of levodopa. If this is the case, early combination of levodopa with COMT inhibitors (tolcapone, entacapone) would prove to be beneficial. Moreover, studies on the relationship between complications to levodopa therapy in PD and homocysteine, folate, vitamin B12, and C677T/MTHFR genotypes are warranted.Plasma homocysteine and cysteine levels were measured in 90 patients with PD with the MTHFR C677T (T/T) genotype. The authors found that the levels of homocysteine-a possible risk factor for vascular disease-were elevated by 60% in levodopa-treated patients with PD, with the most marked elevation occurring in patients with the T/T genotype. Cysteine levels in subjects with PD did not differ from levels in control subjects. In the T/T genotype patients, homocysteine and folate levels were inversely correlated. Increased homocysteine might be related to levodopa, MTHFR genotype, and folate in PD.

Prevalence and clinical characteristics of restless legs syndrome in Japanese patients with Parkinson's disease

To explore the clinical significance of restless legs syndrome (RLS) in Parkinsons disease (PD) and the causal relationship between these two disorders, we made a comparison of both the prevalence of RLS and the severity of sleep disturbance manifested on the Pittsburg Sleep Quality Index (PSQI) between patients with PD (n = 165) and age‐ and sex‐matched control subjects (n = 131). The prevalence of RLS diagnosed by clinical interview was significantly higher in PD patients than in control subjects (12% vs. 2.3%). PSQI score was significantly higher in PD patients with RLS than in both patients without RLS and controls. However, PSQI score was not statistically different between the latter two groups. Among the PD patients with RLS, only 2 had a positive family history of RLS. Only 3 PD patients had requested treatment for the disorder. Our results emphasize the etiological link between RLS and PD in a Japanese cohort, and the existence of RLS is thought to be one of the most important factors aggravating sleep disturbance in PD, despite the low RLS severity.

Transcriptional activation of p62/A170/ZIP during the formation of the aggregates: possible mechanisms and the role in Lewy body formation in Parkinson's disease.

Formation of intracellular inclusion bodies due to defects in the protein degradation machinery is associated with the pathogenesis of neurodegenerative diseases. Sequestosomal protein p62/A170/ZIP, which is an oxidative stress-related protein and a ubiquitin-binding protein, is a component protein of Lewy bodies that are observed in patients with Parkinsons disease. The association of p62 with poly-ubiquitinated proteins may be an important step in the formation of intracellular protein aggregates like Lewy bodies. To study the role of p62 in the formation of protein aggregates in PC12 cells, we monitored the intracellular localizations of p62 and ubiquitinated proteins and the levels of both components during treatment with MG132, a proteasome inhibitor. In the early stage of aggregate formation, p62 did not always co-localize with ubiquitin. In contrast, these proteins were always co-localized in later stages. After the treatment of the cells with MG132, we found that the expression level of p62 increased due to the transcriptional activation of the gene and that higher molecular sizes of p62, corresponding to mono- and di-ubiquitinated formes, were also formed. Both the transcriptional inhibitor actinomycin D and an antisense oligonucleotide of p62 inhibited the MG132-mediated increase of p62, the sequestration of ubiquitinated proteins, and the enlargement of the aggregates. Furthermore, p62-positive aggregates were observed primarily in surviving cells. Together, these results suggest that p62 plays an important role in the protection of cells from the toxicity of misfolded proteins by enhancing aggregate formation especially in the later stages.

CSF orexin levels of Parkinson's disease, dementia with Lewy bodies, progressive supranuclear palsy and corticobasal degeneration

Excessive daytime sleepiness has been widely accepted as a common problem not only in Parkinsons disease (PD) but also in other related disorders. Lowered excretion of orexin A (hypocretin 1) into the cerebrospinal fluid (CSF) is known to play a pathological role in narcolepsy and secondary hypersomnia due to hypothalamic dysfunction. Although the levels of CSF orexin in PD have been previously examined, the results have been controversial, and no systematic investigation of CSF orexin excretion has been conducted on PD related disorders. In this study, orexin was measured in CSF collected by lumbar puncture in 62 patients with PD, 13 patients with dementia with Lewy bodies (DLB), 16 patients with progressive supranuclear palsy (PSP), and 7 patients with corticobasal degeneration (CBD). Levels of CSF orexin (mean+/-SD pg/ml) were 302+/-38 in PD, 297+/-48 in DLB, 258+/-37 in PSP, 246+/-90 in CBD. The occurrence of low orexin levels (<or=110pg/ml) was rare in both PD and DLB, and orexin levels were significantly lower in the PSP and CBD groups compared to PD (PSP: p<0.001, CBD: p<0.05). Orexin levels were inversely correlated with duration of morbidity in PSP but not in the other conditions studied. These findings suggest that loss of orexin neurons or impaired orexin neurotransmission might exist as a part of the neurodegeneration associated with advanced PSP with long duration of morbidity.

Association of the insertion/deletion polymorphism of the angiotensin I-converting enzyme gene in patients of migraine with aura

Recently, several angiotensin I-converting enzyme (ACE) inhibitors and an angiotensin II receptor blocker were demonstrated to have a clinically important prophylactic effect in migraine. ACE is one of the key enzymes in the rennin-angiotensin-aldosterone system, which modulates vascular tension and blood pressure. In humans, serum ACE levels are strongly genetically determined. Individuals who were homozygous for the deletion (D) allele showed increased ACE activity levels. To investigate the role of ACE polymorphism in headache, we analyzed the ACE insertion (I)/deletion (D) genotypes of 54 patients suffering from migraine with aura (MwA), 122 from migraine without aura, 78 from tension-type headache (TH), and 248 non-headache healthy controls. The ACE D allele were significantly more frequent in the MwA than controls (p<0.01). The incidence of the D/D genotype in MwA (25.9%) was significantly higher than that in controls (12.5%; p<0.01; odds ratio=5.26, 95% confidence interval: 1.69-16.34, adjusted for age and gender). No differences in the remaining groups were found. Our results support the conclusion that the D allele and the D/D genotype in the ACE gene is a genetic risk factor for Japanese MwA. There seems to be a possible relationship between ACE activity and the pathogenesis of migraine.

Hypertrophy of IMC of carotid artery in Parkinson's disease is associated with L-DOPA, homocysteine, and MTHFR genotype.

In recent years, an intense interest has developed in the association between Parkinsons disease (PD) and hyperhomocysteinemia. Homocysteine (Hcy) is a neuronal excitotoxic amino acid, and is well known as a risk factor for vascular diseases. Some reports suggest that the administration of L-DOPA may promote hyperhomocysteinemia and idiopathic atherosclerosis. In this study, we report that a mild hypertrophy of the intima-media complex (IMC) of the carotid artery, which has been established as a marker for systemic atherosclerosis, is observed in PD patients compared with normal subjects. PD patients that were treated with L-DOPA for long durations showed a hypertrophic IMC, while the patients that were not treated with L-DOPA did not show any hypertrophic changes in the IMC. These hypertrophic changes were observed primarily in patients with a Hoehn-Yahr stage of 3-5. PD patients with hypertrophic IMC of the carotid artery also exhibited elevated plasma levels of Hcy associated with the C677T genotype of 5,10-methylenetetrahydrofolate reductase (MTHFR). Moreover, a prolonged duration of treatment with L-DOPA in patients with MTHFR T/T genotype enhanced the hypertrophy of IMC, compared with patients with the C/C or C/T genotype. These results suggest that hyperhomocysteinemia promoted by the C677T genotype of MTHFR and prolonged treatment with L-DOPA enhances atherosclerosis in PD patients and affects their general condition.

Levodopa‐induced hyperhomocysteinaemia in Parkinson's disease

Elevated plasma homocysteine (Hcy) has been observed in Parkinson’s disease (PD) in recent years (1, 2). It has been suggested that hyperhomocysteinaemia in PD patients may be associated with levodopa administration and with the methylenetetrahydrofolate reductase (MTHFR) C677T genotype (1). Hcy is well known as one of the risk factors for vascular diseases (3, 4). Furthermore, it may act as an excitotoxic amino acid in neuronal cells, and it is therefore important to manage Hcy levels during treatment for PD. To determine the effect of levodopa administration on secondary hyperhomocysteinaemia in different MTHFR genotypes, Hcy levels in de novo PD patients before and after levodopa treatment were investigated. This study included 20 de novo PD patients, all of whom were Japanese and had given their informed consent to this study, whose plasma Hcy concentrations were measured before levodopa treatment. All 20 PD patients were then administered several doses of levodopa, and plasma Hcy concentrations were measured again. Levodopa was administered as levodopa+carbidopa, at a daily dose of 235 115 mg (mean SD, range: 100–600). The duration of levodopa+carbidopa administration was 110 181 days (mean SD, range: 7–360). Plasma Hcy concentrations were measured by high performance liquid chromatography and the MTHFR genotype was determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), as described previously (1, 5). Hcyconcentrationsbefore levodopaadministration in the 20 de novo PD patients (11.0 4.5 nmol/ml) did not differ significantly compared with control subjects (10.2 5.3 nmol/ml). However, Hcy concentrations were significantly elevated after levodopa administration (18.8 13.5 nmol/ml, P < 0.001 vs control subjects, ANOVA). In all but one patient, Hcy concentrations were elevated after levodopa administration. In order to investigate the association between the increase in Hcy concentrations following levodopa treatment and MTHFR C677T genotype, patients were classified into three groups according to their MTHFR genotypes. Six patients were C/C (wild), eight were C/T, and six were the T/T genotype. It is noteworthy that plasma Hcy concentrations greatly increased following levodopa treatment in patients with the T/T genotype, while those with the C/C or C/T genotypes showed only a slight increase. Hcy concentrations increased from 10.9 1.6 to 14.6 2.4 nmol/ml in the C/C genotype group, from 10.3 4.0 to 14.1 4.2 nmol/ml in the C/T group, and from 11.9 7.1 to 29.3 21.8 nmol/ml in the T/T group. Post-treatment Hcy concentrations in T/T genotype patients were higher than in the other genotype groups (P < 0.03, ANOVA). The relative increase in Hcy concentrations following levodopa administration was 35.0 21.3% (mean SD) for the C/C genotype, 44.8 36.7% for the C/T genotype, and 156.2 108.5% for the T/T genotype. The percentage increase in Hcy concentrations was higher in patients with the T/T genotype than in the other genotype groups

Static stabilometry in patients with migraine and tension-type headache during a headache-free period

Abstract The vestibulospinal system was evaluated using a stabilometric method in patients with migraine and episodic tension‐type headache during headache‐free periods. Migraine patients often complain of dizziness or vertigo during headache attacks and some exhibit these symptoms between attacks. Computerized static stabilometry is a reliable and non‐invasive technique to evaluate the equilibrium function in various diseases. The subjects consisted of 21 patients with migraine, 12 patients with episodic tension‐type headache and, age‐ and sex‐matched controls. We performed two sets of static stabilometric measurements with eyes open (EO) and eyes closed (EC) for 30 s. The averages of two sessions of the following six stabilometric parameters were used for the analysis: locus length (LNG), environmental area (ENV‐AREA), rectangle area (REC‐AREA), locus length per second, locus length per environ area (L/E), and root mean square area. Romberg quotients (EC/EO) of these six parameters were also analyzed. The mean values of LNG, ENV‐AREA and REC‐AREA in the EC session in the migraine group were significantly greater than those in the controls (P < 0.05, Mann–Whitney rank sum test). Romberg quotients of all stabilometric parameters except the L/E in the migraine group were significantly greater than in the controls. Patients with episodic tension‐type headache did not show any differences in the stabilometric study from the controls. The present findings suggest that patients with migraine show a significant increase of the body sway during the EC session, which indicates an underlying dysfunction in the vestibulospinal system.