Ikuko Mizuta

Multicenter Analysis of Glucocerebrosidase Mutations in Parkinson's Disease

BACKGROUND Recent studies indicate an increased frequency of mutations in the gene encoding glucocerebrosidase (GBA), a deficiency of which causes Gauchers disease, among patients with Parkinsons disease. We aimed to ascertain the frequency of GBA mutations in an ethnically diverse group of patients with Parkinsons disease. METHODS Sixteen centers participated in our international, collaborative study: five from the Americas, six from Europe, two from Israel, and three from Asia. Each center genotyped a standard DNA panel to permit comparison of the genotyping results across centers. Genotypes and phenotypic data from a total of 5691 patients with Parkinsons disease (780 Ashkenazi Jews) and 4898 controls (387 Ashkenazi Jews) were analyzed, with multivariate logistic-regression models and the Mantel-Haenszel procedure used to estimate odds ratios across centers. RESULTS All 16 centers could detect two GBA mutations, L444P and N370S. Among Ashkenazi Jewish subjects, either mutation was found in 15% of patients and 3% of controls, and among non-Ashkenazi Jewish subjects, either mutation was found in 3% of patients and less than 1% of controls. GBA was fully sequenced for 1883 non-Ashkenazi Jewish patients, and mutations were identified in 7%, showing that limited mutation screening can miss half the mutant alleles. The odds ratio for any GBA mutation in patients versus controls was 5.43 across centers. As compared with patients who did not carry a GBA mutation, those with a GBA mutation presented earlier with the disease, were more likely to have affected relatives, and were more likely to have atypical clinical manifestations. CONCLUSIONS Data collected from 16 centers demonstrate that there is a strong association between GBA mutations and Parkinsons disease.

Genome-wide association study identifies common variants at four loci as genetic risk factors for Parkinson's disease.

To identify susceptibility variants for Parkinsons disease (PD), we performed a genome-wide association study (GWAS) and two replication studies in a total of 2,011 cases and 18,381 controls from Japan. We identified a new susceptibility locus on 1q32 (P = 1.52 × 10−12) and designated this as PARK16, and we also identified BST1 on 4p15 as a second new risk locus (P = 3.94 × 10−9). We also detected strong associations at SNCA on 4q22 (P = 7.35 × 10−17) and LRRK2 on 12q12 (P = 2.72 × 10−8), both of which are implicated in autosomal dominant forms of parkinsonism. By comparing results of a GWAS performed on individuals of European ancestry, we identified PARK16, SNCA and LRRK2 as shared risk loci for PD and BST1 and MAPT as loci showing population differences. Our results identify two new PD susceptibility loci, show involvement of autosomal dominant parkinsonism loci in typical PD and suggest that population differences contribute to genetic heterogeneity in PD.

Mutations for Gaucher Disease Confer High Susceptibility to Parkinson Disease

BACKGROUND Increased frequency of pathogenic variants in GBA, the causative gene for Gaucher disease, has been suggested to be associated with Parkinson disease (PD). OBJECTIVES To conduct comprehensive resequencing of GBA to identify all sequence variants and to investigate the association of these variants with PD. DESIGN Case-control study. SETTING Multicenter university-based study. PARTICIPANTS Five hundred thirty-four patients with PD, 34 families in which multiple patients with PD are present, and 544 control subjects. MAIN OUTCOME MEASURES Disease status and GBA variations. RESULTS Comprehensive resequencing of GBA in 534 patients with PD and 544 controls revealed 27 sequence variants: 11 pathogenic variants associated with Gaucher disease, 11 nonsynonymous variants not associated with Gaucher disease, and 5 synonymous variants. Fifty patients with PD (9.4%) had 1 of the 11 pathogenic variants in the heterozygous state, whereas only 2 controls (0.37%) had such variants (odds ratio, 28.0). Among the pathogenic variants, R120W and L444P/RecNciI were highly prevalent, and each showed a significant association with PD. Furthermore, other rare pathogenic variants were found in 13 patients with PD but not in the controls, further confirming the role of these rare variants in the susceptibility to PD. Patients with PD carrying pathogenic variants were significantly younger than those not carrying them. In addition, concordance of PD states and pathogenic variants was observed in 8 multiplex families with PD. CONCLUSION Heterozygous pathogenic variants in GBA confer a high risk for sporadic PD, even for familial clustering, and are associated with significantly earlier age at onset of disease.

Riluzole stimulates nerve growth factor, brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor synthesis in cultured mouse astrocytes

Riluzole is an antiexcitotoxic agent used for the treatment of amyotrophic lateral sclerosis, and reported to have neuroprotective effects in animal models of Parkinsons disease, Huntingtons disease and brain ischemia. We investigated the effects of riluzole on synthesis of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) in cultured mouse astrocytes. The protein and mRNA levels were measured by enzyme-linked immunosorbent assay and semiquantitative reverse transcription-polymerase chain reaction, respectively. Treatment with riluzole at 100 microg/ml (426 microM) for 24 h increased the contents of NGF, BDNF, and GDNF in the culture medium 109-fold, 2.0-fold and 3.1-fold over the control, respectively. The drug-induced relative mRNA levels of NGF, BDNF, and GDNF were 7.3-fold at 2 h, 2.1-fold at 4 h, and 1.9-fold at 2 h, respectively. These results indicate that riluzole stimulates synthesis of NGF, BDNF and GDNF in cultured astrocytes. Riluzole might exert neuroprotective effects, at least in part, via stimulation of neurotrophic factors.

Influence of interleukin-1β gene polymorphisms on age-at-onset of sporadic Parkinson's disease

We studied genetic polymorphisms in the promoter region (position -511) and exon 5 (position +3953) of the interleukin (IL)-1β gene in 122 Japanese patients with Parkinsons disease (PD) and 112 controls. We also examined polymorphisms in the IL–1α and the IL–1 receptor antagonist genes. No significant difference was found in these genetic markers between PD patients and controls. However, PD patients with homozygotes for allele 1 at position -511 of the IL–1β gene (IL–1B-511*1), a low producer of IL–1β, were significantly earlier in the disease onset than those with the IL–1B-511*2 homozygotes, a high producer of IL–1β. This suggests that IL–1β might play a role, possibly a protective effect for dopaminergic neurons, in PD. Further population and functional studies are necessary to clarify the role of IL–1β in PD patients.

Tumor necrosis factor gene polymorphisms in patients with sporadic Parkinson's disease

We studied promoter region polymorphisms in the tumor necrosis factor (TNF) gene at position -1031, -863, and -857, in 172 Japanese patients with sporadic Parkinsons disease (PD). The frequency of the -1031C allele, a high producer of TNF, increased significantly in early onset PD patients compared with controls. In addition, PD patients with the -1031C allele showed a significantly earlier onset than those without -1031C allele, after stratification of the data by an interleukin-1beta gene polymorphism. Our findings suggest that TNF might have a toxic effect in PD.

Effects of dopamine agonists bromocriptine, pergolide, cabergoline, and SKF-38393 on GDNF, NGF, and BDNF synthesis in cultured mouse astrocytes

We examined the stimulatory effects of the dopamine agonists bromocriptine, pergolide, cabergoline, and SKF-38393 on the synthesis and secretion of neurotrophic factors (nerve growth factor, NGF; brain-derived neurotrophic factor, BDNF; and glial cell line-derived neurotrophic factor, GDNF) in cultured mouse astrocytes, and clarified the role of dopamine D1 and D2 receptors in these effects. Bromocriptine, a D2 agonist, elevated NGF levels in the culture medium 6.8-fold vs. control, and significantly decreased GDNF and BDNF levels, at 24 h. Both pergolide, a D1/D2 agonist, and cabergoline, a D2/weak D1 agonist, rapidly elevated NGF and GDNF levels at 4-6 h, respectively to 21- and 1.5-fold, respectively, and 84- and 9-fold, respectively, of control levels at 24 h. SKF-38393, a D1 agonist, elevated NGF and GDNF levels to 20- and 2.8-fold of controls, respectively, at 24 h. Relative levels of NGF and GDNF mRNA detected by Northern blot analysis or semiquantitative reverse transcriptase-polymerase chain reaction confirmed that increases in levels of the 2 proteins in culture medium were due to overexpression as opposed to leakage from cells. Cabergoline rapidly increased GDNF mRNA expression at 4 h, producing a potent and long-lasting increase in GDNF levels. Bromocriptine significantly suppressed GDNF synthesis. These findings suggest that stimulation of dopamine D1 receptors may be required for GDNF synthesis and secretion, and that concurrent stimulation of dopamine D1 and D2 receptors may augment synthesis and secretion of NGF and GDNF. These dopamine agonists may play a role in neuronal survival by stimulating NGF and GDNF synthesis in the brain, and as drugs are good candidates as NGF and GDNF inducers.

TNF, TNF receptor type 1, and allograft inflammatory factor-1 gene polymorphisms in Japanese patients with type 1 diabetes.

The human leukocyte antigen (HLA) class III region, located on chromosome 6p21, has been regarded as one of the susceptible loci for type 1 diabetes. Because it contains many genes related to inflammatory and immune responses, including tumor necrosis factor (TNF), lymphotoxin-alpha (LT-alpha), and allograft inflammatory factor 1 (AIF-1) genes, it is unclear which gene within the class III region is responsible for the susceptibility to the disease. We sequenced the AIF-1 gene region and detected three novel polymorphisms, all of which were diallelic and localized at introns. Then, we investigated AIF-1, TNF, and LT-alpha gene polymorphisms in 165 patients with type 1 diabetes, consisting of 90 patients with young-onset type 1 diabetes, 75 patients with adult-onset type 1 diabetes, and 200 control patients. We also analyzed TNF receptors type 1 (TNFR1) and type 2 (TNFR2) gene polymorphisms, located on chromosome 12p13 and 1p36, respectively. Although there were significant differences between type 1 diabetes patients and controls in the distributions of TNF promoter polymorphisms at position -1031 and -857, and LT-alpha gene NcoI polymorphism, none of them was independently associated with the disease after two-locus analysis with HLA class II alleles. We detected the significantly increased frequency of the -383C allele, located in the TNFR-1 promoter region, in both young-onset and adult-onset diabetes patients compared with controls. In addition, the -383C allele was found to be associated with higher expression of the TNFR1 gene than that of -383A allele in in vitro expression. These results suggest that the TNFR1 gene region might be a susceptible locus to type 1 diabetes in Japanese.

Nerve ultrasound depicts peripheral nerve enlargement in patients with genetically distinct Charcot-Marie-Tooth disease

Objective To elucidate the ultrasound (US) features of peripheral nerves including nerve roots in patients with different types of Charcot-Marie-Tooth disease (CMT), and the association between US findings, clinical features and parameters of nerve conduction studies (NCS) in CMT1A. Methods US of median, sural and great auricular nerves and the C6 nerve root was performed in patients with CMT1A (n=20), MPZ-associated CMT (n=3), NEFL-associated CMT (n=4), EGR2-associated CMT (n=1), ARHGEF10-associated CMT (n=1) and in controls (n=30). In patients with CMT1A, we analysed the correlations between US findings and the following parameters: age, CMT Neuropathy Score (CMTNS) and NCS indices of the median nerve. Results Cross-sectional areas (CSAs) of all the nerves were significantly increased in patients with CMT1A compared with that in controls. In MPZ-associated CMT, increased CSAs were found in the median nerve at wrist and in the great auricular nerve, whereas it was not increased in patients with NEFL-associated CMT. In patients with CMT1A, there was a positive correlation between CMTNS and the CSAs in the median nerves or great auricular nerves. In median nerves in patients with CMT1A, we found a negative correlation between the nerve conduction velocity and the CSA. Conclusions Nerve US may aid in differentiating among the subtypes of CMT in combination with NCS. In CMT1A, the median nerve CSA correlates with the disease severity and peripheral nerve function.

Fibroblast growth factor 20 gene and Parkinson's disease in the Japanese population.

A genetic association between the fibroblast growth factor 20 (FGF20) gene and Parkinsons disease has been found by the pedigree disequilibrium test. This association, however, was not replicated by a case–control association study. In order to clarify the association between the FGF20 gene and Parkinsons disease, we attempted to replicate this association by a case–control association study using a large number of Japanese samples (1388 patients and 1891 controls). rs1721100 exhibited a significant difference in allele C versus G (P=0.0089), and in genotype CC+CG versus GG (P=0.0053). Haplotype association analysis showed that haplotype 2 was the protective haplotype for Parkinsons disease (permutation-P=0.0075). These results suggest that the FGF20 gene is a susceptibility gene for Parkinsons disease in the Japanese population.