Mitochondrial DNA variants as genetic risk factors for Parkinson disease.

BACKGROUND AND PURPOSE: Investigation of the relationship between mitochondrial DNA (mtDNA) variants and Parkinson disease (PD) remains an issue awaiting more supportive evidence. Moreover, an affirming cellular model study is also lacking. METHODS: The index mtDNA variants and their defining mitochondrial haplogroup were determined in 725 PD patients and 744 non-PD controls. Full-length mtDNA sequences were also conducted in 110 cases harboring various haplogroups. Cybrid cellular models, composed by fusion of mitochondria-depleted rho-zero cells and donor mitochondria, were used for a rotenone-induced PD simulation study. RESULTS: Multivariate logistic regression analysis revealed that subjects harboring the mitochondrial haplogroup B5 have resistance against PD (odds ratio 0.50, 95% confidence interval 0.32-0.78; P = 0.002). Furthermore, a composite mtDNA variant group consisting of A10398G and G8584A at the coding region was found to have resistance against PD (odds ratio 0.50, 95% confidence interval 0.33-0.78; P = 0.001). In cellular studies, B4 and B5 cybrids were selected according to their higher resistance to rotenone, in comparison with cybrids harboring other haplogroups. The B5 cybrid, containing G8584A/A10398G variants, showed more resistance to rotenone than the B4 cybrid not harboring these variants. This is supported by findings of low reactive oxygen species generation and a low apoptosis rate in the B5 cybrid, whereas a higher expression of autophagy was observed in the B4 cybrid particularly under medium dosage and longer treatment time with rotenone. CONCLUSIONS: Our studies, offering positive results from clinical investigations and cybrid experiments, provide data supporting the role of variant mtDNA in the risk of PD.

Generating allergen-specific human IgEs for immunoassays by employing human ε gene knockin mice.

BACKGROUND: Antigen-specific human IgEs are important reagents in immunoassays to quantify antigen-specific IgEs in allergic patients, but they are not easy to prepare. METHODS: We constructed a knockin homozygous mouse strain, referred to as HεκKI strain, whose gene segment encoding γ1 constant region has been replaced by that encoding human ε constant region and gene segment encoding κ constant region replaced by that encoding human κ constant region. The mice were tested for their ability to produce antigen-specific chimeric human IgE (with mouse variable regions) upon the immunization with ovalbumin and papain. Subsequently, the spleen cells from the immunized mice were used as the source of B cells for the preparation of hybridomas, which secreted monoclonal human IgE antibodies specific for the antigens. RESULTS: The HεκKI mice expressed human IgE (ε, κ) in serum at levels 10- to 30-fold higher than those of mouse IgE. Upon immunization with an antigen, the mice yielded splenic B cells for preparing hybridomas that secrete chimeric human IgE specific for the antigen. Purified IgEs from those hybridomas could activate a basophilic cell line to undergo degranulation upon the stimulation with their respective antigens. CONCLUSIONS: We have developed a human ε gene and κ gene knockin mouse strain, which is useful for producing various antigen-specific chimeric human IgEs for potential use as standards in immunoassays.

Potent antitumor activity of Oct4 and hypoxia dual-regulated oncolytic adenovirus against bladder cancer.

Most solid tumors undergo hypoxia, leading to rapid cell division, metastasis and expansion of a cell population with hallmarks of cancer stem cells (CSCs). Tumor-selective replication of oncolytic adenoviruses may be hindered by oxygen deprivation in tumors. It is desirable to develop a potent oncolytic adenovirus, retaining its antitumor activity even in a hypoxic environment. We have previously generated an Oct4-dependent oncolytic adenovirus, namely Ad9OC, driven by nine copies of the Oct4 response element (ORE) for specifically killing Oct4-overexpressing bladder tumors. Here, we developed a novel Oct4 and hypoxia dual-regulated oncolytic adenovirus, designated AdLCY, driven by both hypoxia response element (HRE) and ORE. We showed that hypoxia-inducible factor (HIF)-2α and Oct4 were frequently overexpressed in hypoxic bladder cancer cells, and HIF-2α was involved in HRE-dependent and Oct4 transactivation. AdLCY exhibited higher cytolytic activities than Ad9OC against hypoxic bladder cancer cells, while sparing normal cells. AdLCY exerted potent antitumor effects in mice bearing human bladder tumor xenografts and syngeneic bladder tumors. It could target hypoxic CD44- and CD133-positive bladder tumor cells. Therefore, AdLCY may have therapeutic potential for targeting hypoxic bladder tumors and CSCs. As Oct4 is expressed in various cancers, AdLCY may be further explored as a broad-spectrum anticancer agent.

Clinical and genetic analysis of Taiwanese patients with hereditary spastic paraplegia type 5.

BACKGROUND AND PURPOSE: Spastic paraplegia type 5 (SPG5) is an autosomal recessive (AR) hereditary spastic paraplegia (HSP) associated with pure or complicated phenotypes. This study aimed to screen SPG5 in Taiwanese HSP patients. METHODS: Sequencing of the SPG5 gene, CYP7B1, was performed in a cohort of 25 ethnic Han Taiwanese patients with AR or sporadic HSP. Clinical information and magnetic resonance imaging (MRI) were analyzed in confirmed SPG5 patients. RESULTS: One (33%) AR kindred and four (18%) sporadic cases had CYP7B1 mutations. All of the SPG5 cases carried the mutation c.334 C>T (R112X). Haplotype analysis suggested a 'founder effect' in ethnic Hans for this mutation. The phenotype was either pure or complicated by cerebellar ataxia. For the primary HSP phenotype, there were profound dorsal column sensory deficits in all patients. Spine MRI showed thoraco-lumbar cord atrophy in some patients. CONCLUSIONS: Spastic paraplegia type 5 is a common cause of AR and sporadic HSPs that has a higher frequency in Taiwanese than in other ethnic groups. It is associated with a CYP7B1 founder mutation and its phenotype is characterized by pronounced dorsal column sensory loss, with cerebellar ataxia in some patients.

(G2019S) LRRK2 activates MKK4-JNK pathway and causes degeneration of SN dopaminergic neurons in a transgenic mouse model of PD.

(G2019S) mutation of leucine-rich repeat kinase 2 (LRRK2) is the most common genetic cause of both familial and sporadic Parkinson's disease (PD) cases. Twelve- to sixteen-month-old (G2019S) LRRK2 transgenic mice prepared by us displayed progressive degeneration of substantia nigra pars compacta (SNpc) dopaminergic neurons and parkinsonism phenotypes of motor dysfunction. LRRK2 is a member of mixed lineage kinase subfamily of mitogen-activated protein kinase kinase kinases (MAPKKKs). We hypothesized that (G2019S) mutation augmented LRRK2 kinase activity, leading to overphosphorylation of downstream MAPK kinase (MKK) and resulting in activation of neuronal death signal pathway. Consistent with our hypothesis, (G2019S) LRRK2 expressed in HEK 293 cells exhibited an augmented kinase activity of phosphorylating MAPK kinase 4 (MKK4) at Ser(257), and protein expression of active phospho-MKK4(Ser257) was upregulated in the SN of (G2019S) LRRK2 transgenic mice. Protein level of active phospho-JNK(Thr183/Tyr185) and phospho-c-Jun(Ser63), downstream targets of phospho-MKK4(Ser257), was increased in the SN of (G2019S) LRRK2 mice. Upregulated mRNA expression of pro-apoptotic Bim and FasL, target genes of phospho-c-Jun(Ser63), and formation of active caspase-9, caspase-8 and caspase-3 were also observed in the SN of (G2019S) LRRK2 transgenic mice. Our results suggest that mutant (G2019S) LRRK2 activates MKK4-JNK-c-Jun pathway in the SN and causes the resulting degeneration of SNpc dopaminergic neurons in PD transgenic mice.

Clinical characteristics of essential tremor in Taiwan: an exploratory-comparative study.

BACKGROUND: There are few large-scale clinical analyses of essential tremor (ET) in Asia. We studied the detailed clinical profile with emphasizing the age of onset, tremor location, specific tremor patterns, and rate of progression (ROP) to delineate the characteristics of Taiwanese ET patients and found the difference between the Taiwanese and the Caucasians ET patients. METHODS: All ET patients fulfilled the Movement Disorders Society diagnosis criteria were investigated with a standardized assessment protocol, which including clinical evaluation, uniform severity scoring, self-reported questionnaires, accelerometry, surface electromyography, and videotaped tremor examination. RESULTS: Of 219 patients recruited from July 2008 to October 2009, 153 completed the study protocol. Their mean age was 58.9 years and 47% were women, and 33.3% had family history (FH). There was bimodal distribution in age of tremor onset in patients without but not in those with FH. Head tremor (HT) was present in 48 of 153 (31%) patients. Patients with HT showed slower tremor frequency and less ROP than those without HT. Sixty-seven (44%) patients presented with intention tremor (IT). Male gender and voice tremor were predictive factors of IT occurrence. CONCLUSIONS: Comparing with the Caucasians, Taiwanese ET patients have different patterns of onset-age distribution and lack of female predominance in ET with HT. However, patients with IT and without HT also progressed more rapid as found in the Caucasian.

Contribution of glucocerebrosidase mutation in a large cohort of sporadic Parkinson's disease in Taiwan.

BACKGROUND AND PURPOSE: The association between glucocerebrosidase (GBA) mutations and Parkinson's disease (PD) is attracting increased attention worldwide. In patients of Chinese ethnicity, other than the common L444P mutation, a few mutations have been reported. However, the contribution of GBA to PD can be answered only by a thorough investigation of its mutations in a unique large population. METHODS: We enrolled 1747 participants: 967 PD patients and 780 healthy individuals. We screened entire GBA coding regions and exon-intron boundaries in 30 randomly chosen PD patients, followed by testing five variants (L444P, D409H, R120W, L174P, and Q497R) in all participants. The G2385R and R1628P in LRRK2 had been previously studied in almost all participants. RESULTS: In total, 36 patients (3.72%) carried a heterozygous mutant GBA allele (27 L444P, 7 RecNciI, and 2 D409H). Only two controls (0.26%) carried heterozygous GBA mutation (1 L444P and 1 RecNciI). In PD group, the mean age at onset in carriers was younger than in non-carriers. The difference in percentage of mutation frequencies between patients and controls was highly significant for the L444P mutation (P < 0.0001). One L444P carrier was also associated with LRRK2 G2385R variant, but no atypical Parkinsonism was observed. CONCLUSIONS: The present study ascertains that L444P mutation in GBA gene may contribute to an earlier onset of development of PD in Han/Chinese population. Following LRRK2 variants, GBA is the second most frequent mutations indicated for sporadic PD development in the Han/Chinese population. These GBA carriers are associated with an earlier onset of Parkinsonism.

Comparison of photodegradative efficiencies and mechanisms of Victoria Blue R assisted by Nafion-coated and fluorinated TiO2 photocatalysts.

The purposes of this research were to study the effects of two modified photocatalysts, Nafion-coated TiO(2) and fluorinated TiO(2), and photocatalytic degradation of Victoria Blue R in aqueous solution. Photocatalytic degradation of Victoria Blue R was accelerated by the modified photocatalysts. Bulk and surface characterizations of the resulting powders were carried out. Attachment of the anions to the TiO(2) surface using the Nafion-coated-TiO(2) possibly results in increased adsorption of the cationic dye, and the degradation rate is larger for the cationic dye. It was found that Victoria Blue R on the two illuminated TiO(2) surfaces underwent very different changes. To obtain a better understanding on the mechanistic details of this modified-TiO(2)-assisted photodegradation of the Victoria Blue R dye with UV irradiation, a large number of intermediates of the process were separated, identified, and characterized by a high-performance liquid chromatography-mass spectrometry technique. Several probable photodegradation pathways were proposed and discussed.

Analysis of the UCHL1 genetic variant in Parkinson's disease among Chinese.

The inverse association of the functional ubiquitin carboxy-terminal hydrolase L1 (UCHL1) S18Y variant with Parkinson's disease (PD) among Caucasian populations has been debated. We conducted a large-scale analysis to investigate the age-of-onset effect of the UCHL1 variant in PD among ethnic Chinese. Individual data sets from 5 centers comprising a total of 4088 study subjects were analyzed. In the univariate analysis, only data from 1 center showed a trend towards a protective effect among young subjects. However, in the combined analysis, no significant association between the UCHL1 variant and PD was detected (A allele frequency 0.531 vs. 0.528, p=0.87, OR 1.01, 95% CI 0.92-1.1). Among subjects less than 60 years old, the OR is 0.99 (95% CI 0.84-1.16, p=0.88). A multivariate logistic regression analysis showed that family history, UCHL1 variant and the interaction of UCHL1 variant and age at onset (p=0.816) were not significantly associated with PD.

A longitudinal study of Taiwanese sialidosis type 1: an insight into the concept of cherry-red spot myoclonus syndrome.

BACKGROUND AND PURPOSE: Sialidosis type 1 (ST-1) is a neurodegenerative disorder with limited long-term follow-up report. This study is to document the chronological profile of ST-1. METHODS: We perform serial analysis of 17 Taiwanese patients with ST-1 focusing on evolution of clinical features, electrophysiological findings, genetic studies, and neuroimage examinations. RESULTS: All patients had a mutation at 554A-->G in exon 3 of the NEU1 gene causing Ser182Gly substitution. Fifteen patients were homozygous. Two patients were heterozygous with novel mutations, 956C-->T causing Ala319Val in one and 163C-->T causing Gln55stop codon in the other. The neuraminidase activity was markedly decreased in all 11 available patients. Only three patients (17.6%) manifested the macular cherry-red spot. The majority of patients (82.3%) developed full-blown manifestation of myoclonus, ataxia, and seizures within 5 years. Abnormal somatosensory evoked potentials with giant cortical waves were found in all patients. Prolonged P100 peak latency of the visual evoked potentials (VEPs) were found in 16 patients (94.1%) in the early stage even without visual symptoms. CONCLUSION: ST-1 in Taiwanese population illustrates distinct characteristics of phenotype with infrequent cherry-red spot. We suggest to screen the NEU1 mutations in patients presenting action myoclonus with abnormal VEPs, even without macular cherry-red spots.

UV light induced photodegradation of malachite green on TiO2 nanoparticles.

The photodegradation of malachite green (MG), a cationic triphenylmethane dye, is examined both under different pH values and amounts of TiO(2). After 15W UV-365nm irradiation for 4h, ca. 99.9% of MG was degraded with addition of 0.5gL(-1) TiO(2) to solutions containing 50mgL(-1) of the MG dye. The HPLC-PDA-ESI-MS technique was used to obtain a better understanding on the mechanistic details of this TiO(2)-assisted photodegradation of the MG dye with UV irradiation. Five intermediates of the process were separated, identified, and characterized for the first time. The results indicated that the N-de-methylation degradation of MG dye took place in a stepwise manner to yield mono-, di-, tri-, and tetra-N-de-methylated MG species generated during the processes. Under acidic conditions, the results indicated that the photodegradation mechanism is favorable to cleavage of the whole conjugated chromophore structure of the MG dye. Under basic conditions, the results showed that the photodegradation mechanism is favorable to a formation of a series of N-de-methylated intermediates of the MG dye.

The LRRK2 Gly2385Arg variant is associated with Parkinson's disease: genetic and functional evidence.

Evidence of LRRK2 haplotypes associated with Parkinson's disease (PD) risk was recently found in the Chinese population from Singapore, and a common LRRK2 missense variant, Gly2385Arg, was independently detected as a putative risk factor for PD in the Chinese population from Taiwan. To test the association between the Gly2385Arg variant in a large case-control sample of Chinese ethnicity from Singapore, and to perform functional studies of the wild type and Gly2385Arg LRRK2 protein in human cell lines. In a case-control study involving 989 Chinese subjects, the frequency of the heterozygous Gly2385Arg genotype was higher in PD compared to controls (7.3 vs. 3.6%, odds ratio = 2.1, 95% CI: 1.1-3.9, P = 0.014); these values yield an estimated population attributable risk (PAR) of approximately 4%. In a multivariate logistic regression analysis with the disease group (PD vs. controls) as the dependent variable and the genotype as an independent factor with adjustments made for the effect of age and gender, the heterozygous Gly2385Arg genotype remained associated with an increased risk of PD compared to wild type genotype (odds ratio = 2.67, 95% CI: 1.43-4.99, P = 0.002). The glycine at position 2385 is a candidate site for N-myristoylation, and the Gly2385Arg variant replaces the hydrophobic glycine with the hydrophilic arginine, and increases the net positive charge of the LRRK2 WD40 domain. In transfection studies, we demonstrated that both the wild type and Gly2385Arg variant LRRK2 protein localize to the cytoplasm and form aggregates. However, under condition of oxidative stress, the Gly2385Arg variant was more toxic and associated with a higher rate of apoptosis. Our study lends support to the contention that the Gly2385Arg is a common risk factor for PD in the Chinese population. Our bioinformatics and in-vitro studies also suggest that the Gly2385Arg variant is biologically relevant and it might act through pro-apoptotic mechanisms.

The natural history of neurological manganism over 18 years.

We investigated the clinical features and progression of four patients with chronic manganese intoxication, 18 years after cessation of exposure. Because the results were to be compared with previous observations, we employed the same scoring system. The clinical manifestations were foot dystonia, wide based gait, rigidity, and difficulty in walking backwards. Resting tremor was rarely seen, but tongue tremor was found in 2 patients. The asymmetry initially present in 2 patients persisted 18 years later. Measurements had previously revealed rapid progression in the initial 10 years. We found a plateau over the following decade.

Genetic and DAT imaging studies of familial parkinsonism in a Taiwanese cohort.

We here summarize the results of genetic investigations on a series of 82 parkinsonian patients from 60 families in Taiwan. We found 13 parkin patients in 7 families (12%), 2 PINK1 sibs from 1 family, and 1 LRRK2 patient from 1 family with I2012T mutation. We also identified SCA2 in 8 patients from 5 families (8%) and SCA3 in 3 patients from 1 family, all presenting with parkinsonian phenotype. In the available patients with parkin, PINK1, SCA2 and SCA3, the dopamine transporter (DAT) scan revealed that the reduction of uptake was primarily observed in the bilateral putamen, basically sharing a similar pattern with that in idiopathic Parkinson's disease. We concluded that the genetic causes contributed to about 25% of our series of familial parkinsonism. The parkin mutations and SCA2 were the most frequent genetic causes in our series with Chinese ethnicity. The results of DAT scan indicated that bilateral putamen was essentially involved in various genetically-caused familial parkinsonism.

PARK6-linked autosomal recessive early-onset parkinsonism in Asian populations.

The authors performed linkage analysis in 39 families with autosomal recessive early-onset PD (AR-EOPD) negative for parkin and DJ-1 mutations. Eight families including three Japanese, two Taiwanese, one Turkish, one Israeli, and one Philippine showed evidence of linkage with PARK6 with multipoint log of the odds (lod) score of 9.88 at D1S2732. The results indicate worldwide distribution of PARK6-linked parkinsonism.

Two novel mutations of the glycine receptor gene in a Taiwanese hyperekplexia family.

The authors report a Taiwanese family with autosomal recessive hyperekplexia. Two novel mutations, W96C (from the paternal allele) and R344X (from the maternal allele), which are located in exon 4 and exon 7 of the GLRA1 gene, were identified in this family. A series of electrophysiologic investigations were conducted in one of the probands, and the results suggest that the "startle center" is located subcortically.