I-g Chen

Nuclear receptor NR4A2 IVS6 +18insG and brain derived neurotrophic factor (BDNF) V66M polymorphisms and risk of taiwanese Parkinson's disease

Both of environmental and genetic factors confer vulnerability to Parkinsons disease (PD). NR4A2 (Nurr1), a member of the steroid/thyroid hormone nuclear receptor superfamily, is essential for the neurogenesis and differentiation of dopaminergic neurons in the midbrain. Brain derived neurotrophic factor (BDNF) deficiency may play a role in the pathogenesis of PD, as the surviving dopaminergic nigrostriatal neurons have reduced levels of BDNF. This study examines whether BDNF V66M (c.196 G → A) or NR4A2 IVS6 +18insG polymorphism is associated with the risk of Taiwanese PD and the age of onset using a case‐control study. The genotype or allele frequency distribution of both BDNF V66M and NR4A2 IVS6 +18insG polymorphisms was not significantly different between the cases and the controls. Neither BDNF nor NR4A2 polymorphism influences PD onset age. Notably, after stratification by sex, female individuals carrying the NR4A2 2G/2G genotype demonstrated a trend toward significant decrease in risk of developing PD (OR = 0.49, 95% CI = 0.25–0.96, P = 0.039). These results suggest that the NR4A2 IVS6 +18insG polymorphism may play a minor role in PD susceptibility among Taiwanese women.

Spinocerebellar ataxia type 8 larger triplet expansion alters histone modification and induces RNA foci.

BackgroundSpinocerebellar ataxia type 8 (SCA8) involves the expression of an expanded CTG/CAG combined repeats (CR) from opposite strands producing CUG expansion transcripts (ataxin 8 opposite strand, ATXN8OS) and a polyglutamine expansion protein (ataxin 8, ATXN8). The pathogenesis of SCA8 is complex and the spectrum of clinical presentations is broad.ResultsUsing stably induced cell models expressing 0, 23, 88 and 157 CR, we study the role of ATXN8OS transcripts in SCA8 pathogenesis. In the absence of doxycycline, the stable ATXN8OS CR cell lines exhibit low levels of ATXN8OS expression and a repeat length-related increase in staurosporine sensitivity and in the number of annexin positive cells. A repeat length-dependent repression of ATXN8OS expression was also notable. Addition of doxycycline leads to 25~50 times more ATXN8OS RNA expression with a repeat length-dependent increase in fold of ATXN8OS RNA induction. ChIP-PCR assay using anti-dimethyl-histone H3-K9 and anti-acetyl-histone H3-K14 antibodies revealed increased H3-K9 dimethylation and reduced H3-K14 acetylation around the ATXN8OS cDNA gene in 157 CR line. The repeat length-dependent increase in induction fold is probably due to the increased RNA stability as demonstrated by monitoring ATXN8OS RNA decay in cells treated with the transcriptional inhibitor, actinomycin D. In cells stably expressing ATXN8OS, RNA FISH experiments further revealed ribonuclear foci formation in cells carrying expanded 88 and 157 CR.ConclusionThe present study demonstrates that the expanded CUG-repeat tracts are toxic to human cells and may affect ATXN8OS RNA expression and stability through epigenetic and post-transcriptional mechanisms.

ATP13A2 variability in Taiwanese Parkinson's disease

Mutations in ATP13A2 have been reported to associate with Parkinsons disease (PD). This study investigates the contribution of genetic variants in ATP13A2 to Taiwanese PD. ATP13A2 cDNA fragments from 65 early onset PD (onset <50 years) were sequenced. The identified variants were validated in a cohort of PD (n = 493) and ethnically matched controls (n = 585). A novel heterozygous G1014S, located at the conserved seventh transmembrane domain of ATP13A2 protein, was identified in an early onset PD patient, which was absent in 585 normal controls. Additionally, a reported heterozygous A746T was found in two PD patients and four controls. The clinical features and 99mTc‐TRODAT‐1 single photon emission computed tomography (SPECT) image of the patients carrying G1014S and A746T were similar to that of idiopathic PD. One normal control with A746T showed an asymmetric reduction of 99mT TRODAT‐1 uptake in the right striatum. Under oxidative stress or apoptotic stimulus, lymphoblastoid cells carrying either A764T or G1014S showed increased caspase 3 activity compared with the controls. The rates of decay for G1014S and A746T proteins were more or less reduced in cycloheximide chase experiment. In silico modeling of G1014S exhibited a more stable feature than wild‐type, and G1014S is mislocalized mainly in the intralysosomal space, which is coherent with the prediction of prohibiting N‐myristoylation and membrane association. We therefore hypothesize that rare variants of ATP13A2 may contribute to PD susceptibility in Taiwan. The role played by ATP13A2 variants in PD remains to be clarified.

α-Synuclein promoter RsaI T-to-C polymorphism and the risk of Parkinson’s disease

Summary.Increased α-synuclein expression may be involved in the pathogenesis of Parkinson’s disease (PD). We investigated the association of Rep1 microsatellite and RsaI T-to-C substitution in the α-synuclein promoter region with the risk of PD by a case-control study. The RsaI C/C genotype and C allele were found less frequently in PD patients than in controls. A reduced risk of the Rep1-RsaI 0-C haplotype (OR = 0.57, 95% CI = 0.36–0.90) with PD was evident. The quantitative real-time PCR study showed that the α-synuclein mRNA expression was increased (although not significantly) in PD patients with RsaI T/T genotype or Rep1-RsaI 0-T haplotype as compared to T/C genotype or 0-C haplotype. Reporter constructs containing the RsaI C allele drove significantly lower transcriptional activity compared with the RsaI T allele in both IMR32 and 293 cells. The findings suggest that the RsaI T-to-C substitution may have a functional relevance to the susceptibility to PD.

Aqueous extract of Glycyrrhiza inflata inhibits aggregation by upregulating PPARGC1A and NFE2L2-ARE pathways in cell models of spinocerebellar ataxia 3.

Spinocerebellar ataxia (SCA) types 1, 2, 3, 6, 7, and 17 and dentatorubropallidoluysian atrophy, as well as Huntington disease, are a group of neurodegenerative disorders caused by a CAG triplet-repeat expansion encoding a long polyglutamine (polyQ) tract in the respective mutant proteins. The cytoplasmic and nuclear aggregate formation, a pathological hallmark of polyQ diseases, is probably the initial process triggering the subsequent pathological events. Compromised oxidative stress defense capacity and mitochondrial dysfunction have emerged as contributing factors to the pathogenesis of polyQ diseases. The roots of licorice (Glycyrrhiza species) have long been used as an herbal medicine. In this study, we demonstrate the aggregate-inhibitory effect of Glycyrrhiza inflata herb extract and its constituents licochalcone A and ammonium glycyrrhizinate (AMGZ) in both 293 and SH-SY5Y ATXN3/Q75 cells, SCA3 cell models. The reporter assay showed that G. inflata herb extract, licochalcone A, and AMGZ could enhance the promoter activity of peroxisome proliferator-activated receptor γ, coactivator 1α (PPARGC1A), a known regulator of mitochondrial biogenesis and antioxidative response genes. G. inflata extract, licochalcone A, and AMGZ upregulated PPARGC1A expression and its downstream target genes, SOD2 and CYCS, in the 293 ATXN3/Q75 cell model. The expression of nuclear factor erythroid 2-related factor 2 (NFE2L2), the principal transcription factor that binds to antioxidant-responsive elements (AREs) to promote ARE-dependent gene expression when the cells respond to oxidative stress, and its downstream genes, HMOX1, NQO1, GCLC, and GSTP1, was also increased by G. inflata herb extract, licochalcone A, and AMGZ. Knockdown of PPARGC1A increased aggregates in ATXN3/Q75 cells and also attenuated the aggregate-inhibiting effect of the tested compounds. G. inflata extract and its constituents significantly elevated GSH/GSSG ratio and reduced reactive oxidative species in ATXN3/Q75 cells. The study results suggest that the tested agents activate PPARGC1A activity and NFE2L2-ARE signaling to increase mitochondrial biogenesis, decrease oxidative stress, and reduce aggregate formation in SCA3 cellular models.

Expanded trinucleotide repeats in the TBP/SCA17 gene mapped to chromosome 6q27 are associated with schizophrenia

Schizophrenia has a complex and non-Mendelian mode of inheritance. Recently, trinucleotide repeat (TNR)-containing genes have been considered as the candidate genes predisposing to schizophrenia. The purpose of this study was to determine whether a genetic association could be observed between schizophrenia and the TNR polymorphisms within the KLHL1AS/SCA8, PPP2R2B/SCA12, and TBP/SCA17 genes. We studied 100 unrelated schizophrenia patients and 124 controls without evident neurodegenerative or psychiatric disorders. The overall allele frequency distributions of the KLHL1AS/SCA8 and PPP2R2B/SCA12 genes were not significantly different between the schizophrenic patients and the control subjects (P>0.05). The allele frequency distribution in the schizophrenic patients was significantly different from that in the controls at the TBP/SCA17 gene (P=0.0149), with an increased frequency of 36 repeats in the patients and two patients carrying 45 TNR expansions were identified. TBP/SCA17 is the TATA box binding protein gene mapped to chromosome 6q27. The study suggests that TNR expansions of the TBP/SCA17 gene may contribute to the genetic risk of schizophrenia in rare cases.

Impairment of proteasome and anti-oxidative pathways in the induced pluripotent stem cell model for sporadic Parkinson's disease

BACKGROUND Parkinsons disease (PD) is associated with the progressive degeneration of dopaminergic neurons with abnormal accumulation of α-synuclein mainly in the ventral midbrain. However, the lack of live human neurons from PD patients and their heterogeneous pathogenic nature limit mechanistic studies and therefore the development of drugs to modify the disease progression of PD. The evolution of induced pluripotent stem cell (iPSC) technology makes it possible to generate patient-specific neurons to explore the pathogenesis in individual PD patients. METHODS We generated PD-iPSCs from a sporadic early onset PD patient carrying a heterozygous deletion of exon 5 (Ex5del) in PARK2. The expression of α-synuclein and proteasome and anti-oxidative functions were examined in differentiated iPSC-derived neurons. RESULTS The neurons derived from our PD-iPSCs demonstrated abnormal α-synuclein accumulation and down-regulation of the proteasome and anti-oxidative pathways. Environmental triggers such as proteasome inhibitor MG132 and H2O2 markedly induced cell death, while the proteasome enhancer benzamil and anti-oxidative compound genipin significantly rescued these increased susceptibilities. CONCLUSIONS These results demonstrate that unique genetic-environmental interactions are involved in neuronal death in PD patients. Our findings also provide a new model to identify potential disease-modifying strategies and an insight into personalized medicine for patients with PD.

The aqueous extract of Glycyrrhiza inflata can upregulate unfolded protein response-mediated chaperones to reduce tau misfolding in cell models of Alzheimer’s disease

Background Alzheimer’s disease (AD) and several neurodegenerative disorders known as tauopathies are characterized by misfolding and aggregation of tau protein. Although several studies have suggested the potential of traditional Chinese medicine (TCM) as treatment for neurodegenerative diseases, the role of TCM in treating AD and tauopathies have not been well explored. Materials and methods Tau protein was coupled to the DsRed fluorophore by fusing a pro-aggregation mutant of repeat domain of tau (ΔK280 tauRD) with DsRed. The ΔK280 tauRD-DsRed fusion gene was then used to generate Tet-On 293 and SH-SY5Y cell clones as platforms to test the efficacy of 39 aqueous extracts of TCM in reducing tau misfolding and in neuroprotection. Results Seven TCM extracts demonstrated a significant reduction in tau misfolding and reactive oxidative species with low cytotoxicity in the ΔK280 tauRD-DsRed 293 cell model. Glycyrrhiza inflata and Panax ginseng also demonstrated the potential to improve neurite outgrowth in the ΔK280 tauRD-DsRed SH-SY5Y neuronal cell model. G. inflata further rescued the upregulation of ERN2 (pro-apoptotic) and downregulation of unfolded-protein-response-mediated chaperones ERP44, DNAJC3, and SERP1 in ΔK280 tauRD-DsRed 293 cells. Conclusion This in vitro study provides evidence that G. inflata may be a novel therapeutic for AD and tauopathies. Future applications of G. inflata on animal models of AD and tauopathies are warranted to corroborate its effect of reducing misfolding and potential disease modification.

Novel Lactulose and Melibiose Targeting Autophagy to Reduce PolyQ Aggregation in Cell Models of Spinocerebellar Ataxia 3.

Trehalose, a chemical chaperone and mTOR-independent autophagy enhancer, has shown promise in models of Huntingtons disease, Parkinsons disease and tauopathies. In this study, two trehalase analogs, lactulose and melibiose, were examined for their potentials in spinocerebellar ataxia treatment. Using a SCA3 ATXN3/Q75-GFP cell model, we found that the ATXN3/Q75 aggregation was significantly prohibited by lactulose and melibiose because of their abilities to up-regulate autophagy. Meanwhile, lactulose and melibiose reduced reactive oxygen species production in ATXN3/Q75 cells. Both of them further inhibited the ATXN3/Q75 aggregation in neuronally differentiated SH-SY5Y cells. These findings suggest the therapeutic applications of novel trehalose analogs in polyglutamine aggregation-associated neurodegenerative diseases.

ATXN8 -62 G/A promoter polymorphism and risk of Taiwanese Parkinson's disease

We recently reported a novel −62 G/A polymorphism within ataxin 8 (ATXN8) gene promoter region, with −62 G displaying significantly higher luciferase activity compared with −62 A. Phenotypic variability in spinocerebellar ataxia type 8 (SCA8) has been suggested, and large SCA8 repeats were found in patients with Parkinsons disease (PD). We aimed to investigate the association of ATXN8 −62 G/A polymorphism with the risk of Taiwanese PD, and identify the trans‐acting factor modulating the ATXN8 promoter activity.