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1.
Biosensors (Basel) ; 12(10)2022 Oct 17.
Artigo em Inglês | MEDLINE | ID: mdl-36291020

RESUMO

Alzheimer's disease (AD) is the most common form of dementia. The most convincing biomarkers in the blood for AD are currently ß-amyloid (Aß) and Tau protein because amyloid plaques and neurofibrillary tangles are pathological hallmarks in the brains of patients with AD. The development of assay technologies in diagnosing early-stage AD is very important. The study of human AD subjects is hindered by ethical and technical limitations. Thus, many studies have therefore turned to AD animal models, such as Drosophila melanogaster, to explore AD pathology. However, AD biomarkers such as Aß and p-Tau protein in Drosophilamelanogaster occur at extremely low levels and are difficult to detect precisely. In this study, we applied the immunomagnetic reduction (IMR) technology of nanoparticles for the detection of p-Tau expressions in hTauR406W flies, an AD Drosophila model. Furthermore, we used IMR technology as a biosensor in the therapeutic evaluation of Chinese herbal medicines in hTauR406W flies with Tau-induced toxicity. To uncover the pathogenic pathway and identify therapeutic interventions of Chinese herbal medicines in Tau-induced toxicity, we modeled tauopathy in the notum of hTauR406W flies. Our IMR data showed that the selected Chinese herbal medicines can significantly reduce p-Tau expressions in hTauR406W flies. Using evidence of notal bristle quantification and Western blotting analysis, we confirmed the validity of the IMR data. Thus, we suggest that IMR can serve as a new tool for measuring tauopathy and therapeutic evaluation of Chinese herbal medicine in an AD Drosophila model.


Assuntos
Doença de Alzheimer , Técnicas Biossensoriais , Medicamentos de Ervas Chinesas , Tauopatias , Animais , Humanos , Proteínas tau , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/patologia , Drosophila/metabolismo , Medicamentos de Ervas Chinesas/uso terapêutico , Drosophila melanogaster/metabolismo , Tauopatias/tratamento farmacológico , Peptídeos beta-Amiloides , Tecnologia
2.
Aging (Albany NY) ; 14(18): 7568-7586, 2022 09 26.
Artigo em Inglês | MEDLINE | ID: mdl-36170028

RESUMO

Decreased BDNF and impaired TRKB signaling contribute to neurodegeneration in Alzheimer's disease (AD). We have shown previously that coumarin derivative LM-031 enhanced CREB/BDNF/BCL2 pathway. In this study we explored if LM-031 analogs LMDS-1 to -4 may act as TRKB agonists to protect SH-SY5Y cells against Aß toxicity. By docking computation for binding with TRKB using 7,8-DHF as a control, all four LMDS compounds displayed potential of binding to domain d5 of TRKB. In addition, all four LMDS compounds exhibited anti-aggregation and neuroprotective efficacy on SH-SY5Y cells with induced Aß-GFP expression. Knock-down of TRKB significantly attenuated TRKB downstream signaling and the neurite outgrowth-promoting effects of these LMDS compounds. Among them, LMDS-1 and -2 were further examined for TRKB signaling. Treatment of ERK inhibitor U0126 or PI3K inhibitor wortmannin decreased p-CREB, BDNF and BCL2 in Aß-GFP cells, implicating the neuroprotective effects are via activating TRKB downstream ERK, PI3K-AKT and CREB signaling. LMDS-1 and -2 are blood-brain barrier permeable as shown by parallel artificial membrane permeability assay. Our results demonstrate how LMDS-1 and -2 are likely to work as TRKB agonists to exert neuroprotection in Aß cells, which may shed light on the potential application in therapeutics of AD.


Assuntos
Doença de Alzheimer , Glicoproteínas de Membrana/agonistas , Neuroblastoma , Fármacos Neuroprotetores , Receptor trkB/agonistas , Peptídeos beta-Amiloides/toxicidade , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Cumarínicos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Membranas Artificiais , Fármacos Neuroprotetores/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2 , Wortmanina
3.
Proc Natl Acad Sci U S A ; 119(32): e2204779119, 2022 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-35914128

RESUMO

Earlier work has shown that siRNA-mediated reduction of the SUPT4H or SUPT5H proteins, which interact to form the DSIF complex and facilitate transcript elongation by RNA polymerase II (RNAPII), can decrease expression of mutant gene alleles containing nucleotide repeat expansions differentially. Using luminescence and fluorescence assays, we identified chemical compounds that interfere with the SUPT4H-SUPT5H interaction and then investigated their effects on synthesis of mRNA and protein encoded by mutant alleles containing repeat expansions in the huntingtin gene (HTT), which causes the inherited neurodegenerative disorder, Huntington's Disease (HD). Here we report that such chemical interference can differentially affect expression of HTT mutant alleles, and that a prototypical chemical, 6-azauridine (6-AZA), that targets the SUPT4H-SUPT5H interaction can modify the biological response to mutant HTT gene expression. Selective and dose-dependent effects of 6-AZA on expression of HTT alleles containing nucleotide repeat expansions were seen in multiple types of cells cultured in vitro, and in a Drosophila melanogaster animal model for HD. Lowering of mutant HD protein and mitigation of the Drosophila "rough eye" phenotype associated with degeneration of photoreceptor neurons in vivo were observed. Our findings indicate that chemical interference with DSIF complex formation can decrease biochemical and phenotypic effects of nucleotide repeat expansions.


Assuntos
Azauridina , Proteína Huntingtina , Doença de Huntington , Proteínas Mutantes , Mutação , Proteínas Nucleares , Fenótipo , Proteínas Repressoras , Fatores de Elongação da Transcrição , Alelos , Animais , Azauridina/farmacologia , Células Cultivadas , Expansão das Repetições de DNA , Modelos Animais de Doenças , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Humanos , Proteína Huntingtina/biossíntese , Proteína Huntingtina/genética , Proteína Huntingtina/metabolismo , Doença de Huntington/genética , Medições Luminescentes , Proteínas Mutantes/biossíntese , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Proteínas Nucleares/metabolismo , Células Fotorreceptoras de Invertebrados/efeitos dos fármacos , Proteínas Repressoras/metabolismo , Fatores de Elongação da Transcrição/metabolismo
4.
Plants (Basel) ; 11(4)2022 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-35214904

RESUMO

Alzheimer's disease (AD), a main cause of dementia, is the most common neurodegenerative disease that is related to the abnormal accumulation of amyloid ß (Aß) proteins. Yi-Gan-San (YGS), a traditional herbal medicine, has been used for the management of neurodegenerative disorders and for the treatment of neurosis, insomnia and dementia. The aim of this study was to examine antioxidant capacity and cytotoxicity of YGS treatment by using 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays in vitro. We explored neuroprotective effects of YGS treatment in alleviating Aß neurotoxicity of Drosophila melanogaster in vivo by comparing survival rate, climbing index, and Aß expressions through retinal green fluorescent protein (GFP) expression, highly sensitive immunomagnetic reduction (IMR) and Western blotting assays. In the in vitro study, our results showed that scavenging activities of free radical and SH-SY5Y nerve cell viability were increased significantly (p < 0.01-0.05). In the in vivo study, Aß42-expressing flies (Aß42-GFP flies) and their WT flies (mCD8-GFP flies) were used as an animal model to examine the neurotherapeutic effects of YGS treatment. Our results showed that, in comparison with those Aß42 flies under sham treatments, Aß42 flies under YGS treatments showed a greater survival rate, better climbing speed, and lower Aß42 aggregation in Drosophila brain tissue (p < 0.01). Our findings suggest that YGS should have a beneficial alternative therapy for AD and dementia via alleviating Aß neurotoxicity in the brain tissue.

5.
Cells ; 10(11)2021 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-34831318

RESUMO

Alzheimer's disease (AD) is a common neurodegenerative disease presenting with progressive memory and cognitive impairments. One of the pathogenic mechanisms of AD is attributed to the aggregation of misfolded amyloid ß (Aß), which induces neurotoxicity by reducing the expression of brain-derived neurotrophic factor (BDNF) and its high-affinity receptor tropomyosin-related kinase B (TRKB) and increasing oxidative stress, caspase-1, and acetylcholinesterase (AChE) activities. Here, we have found the potential of two novel synthetic coumarin derivatives, ZN014 and ZN015, for the inhibition of Aß and neuroprotection in SH-SY5Y neuroblastoma cell models for AD. In SH-SY5Y cells expressing the GFP-tagged Aß-folding reporter, both ZN compounds reduced Aß aggregation, oxidative stress, activities of caspase-1 and AChE, as well as increased neurite outgrowth. By activating TRKB-mediated extracellular signal-regulated kinase (ERK) and AKT serine/threonine kinase 1 (AKT) signaling, these two ZN compounds also upregulated the cAMP-response-element binding protein (CREB) and its downstream BDNF and anti-apoptotic B-cell lymphoma 2 (BCL2). Knockdown of TRKB attenuated the neuroprotective effects of ZN014 and ZN015. A parallel artificial membrane permeability assay showed that ZN014 and ZN015 could be characterized as blood-brain barrier permeable. Our results suggest ZN014 and ZN015 as novel therapeutic candidates for AD and demonstrate that ZN014 and ZN015 reduce Aß neurotoxicity via pleiotropic mechanisms.


Assuntos
Peptídeos beta-Amiloides/toxicidade , Cumarínicos/farmacologia , Proteínas de Fluorescência Verde/toxicidade , Fármacos Neuroprotetores/farmacologia , Acetilcolinesterase/metabolismo , Disponibilidade Biológica , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Caspase 1/metabolismo , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular/efeitos dos fármacos , Cumarínicos/química , Técnicas de Silenciamento de Genes , Humanos , Crescimento Neuronal/efeitos dos fármacos , Agregados Proteicos , Espécies Reativas de Oxigênio/metabolismo , Receptor trkB/metabolismo
6.
Oxid Med Cell Longev ; 2021: 3058861, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34812274

RESUMO

Abnormal accumulations of misfolded Aß and tau proteins are major components of the hallmark plaques and neurofibrillary tangles in the brains of Alzheimer's disease (AD) patients. These abnormal protein deposits cause neurodegeneration through a number of proposed mechanisms, including downregulation of the cAMP-response-element (CRE) binding protein 1 (CREB) signaling pathway. Using CRE-GFP reporter cells, we investigated the effects of three coumarin-chalcone derivatives synthesized in our lab on CREB-mediated gene expression. Aß-GFP- and ΔK280 tauRD-DsRed-expressing SH-SY5Y cells were used to evaluate these agents for possible antiaggregative, antioxidative, and neuroprotective effects. Blood-brain barrier (BBB) penetration was assessed by pharmacokinetic studies in mice. Of the three tested compounds, (E)-3-(3-(4-(dimethylamino)phenyl)acryloyl)-4-hydroxy-2H-chromen-2-one (LM-021) was observed to increase CREB-mediated gene expression through protein kinase A (PKA), Ca2+/calmodulin-dependent protein kinase II (CaMKII), and extracellular signal-regulated kinase (ERK) in CRE-GFP reporter cells. LM-021 exhibited antiaggregative, antioxidative, and neuroprotective effects mediated by the upregulation of CREB phosphorylation and its downstream brain-derived neurotrophic factor and BCL2 apoptosis regulator genes in Aß-GFP- and ΔK280 tauRD-DsRed-expressing SH-SY5Y cells. Blockage of the PKA, CaMKII, or ERK pathway counteracted the beneficial effects of LM-021. LM-021 also exhibited good BBB penetration ability, with brain to plasma ratio of 5.3%, in in vivo pharmacokinetic assessment. Our results indicate that LM-021 works as a CREB enhancer to reduce Aß and tau aggregation and provide neuroprotection. These findings suggest the therapeutic potential of LM-021 in treating AD.


Assuntos
Doença de Alzheimer/prevenção & controle , Peptídeos beta-Amiloides/metabolismo , Chalconas/química , Cumarínicos/química , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Fármacos Neuroprotetores/farmacologia , Proteínas tau/metabolismo , Doença de Alzheimer/etiologia , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Animais , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Humanos , Masculino , Camundongos , Camundongos Endogâmicos ICR , Neuroblastoma/tratamento farmacológico , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Fármacos Neuroprotetores/química , Proteínas tau/genética
7.
Mol Brain ; 14(1): 152, 2021 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-34607601

RESUMO

The glutamatergic signaling pathway is involved in molecular learning and human cognitive ability. Specific single variants (SNVs, formerly single-nucleotide polymorphisms) in the genes encoding N-methyl-D-aspartate receptor subunits have been associated with neuropsychiatric disorders by altering glutamate transmission. However, these variants associated with cognition and mental activity have rarely been explored in healthy adolescents. In this study, we screened for SNVs in the glutamatergic signaling pathway to identify genetic variants associated with cognitive ability. We found that SNVs in the subunits of ionotropic glutamate receptors, including GRIA1, GRIN1, GRIN2B, GRIN2C, GRIN3A, GRIN3B, and calcium/calmodulin-dependent protein kinase IIα (CaMK2A) are associated with cognitive function. Plasma CaMK2A level was correlated positively with the cognitive ability of Taiwanese senior high school students. We demonstrated that elevating CaMK2A increased its autophosphorylation at T286 and increased the expression of its downstream targets, including GluA1 and phosphor- GluA1 in vivo. Additionally, methyl-CpG binding protein 2 (MeCP2), a downstream target of CaMK2A, was found to activate the expression of CaMK2A, suggesting that MeCP2 and CaMK2A can form a positive feedback loop. In summary, two members of the glutamatergic signaling pathway, CaMK2A and MeCP2, are implicated in the cognitive ability of adolescents; thus, altering the expression of CaMK2A may affect cognitive ability in youth.


Assuntos
Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/fisiologia , Cognição/fisiologia , Proteína 2 de Ligação a Metil-CpG/fisiologia , Psicologia do Adolescente , Receptores Ionotrópicos de Glutamato/genética , Transdução de Sinais/fisiologia , Adolescente , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/sangue , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Linhagem Celular Tumoral , Ativação Enzimática , Retroalimentação Fisiológica/fisiologia , Feminino , Ácido Glutâmico/fisiologia , Células HEK293 , Humanos , Masculino , Neuroblastoma , Fosforilação , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas , Processamento de Proteína Pós-Traducional , Receptores Ionotrópicos de Glutamato/fisiologia , Valores de Referência , Taiwan
8.
Front Aging Neurosci ; 13: 758895, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34975454

RESUMO

Alzheimer's disease (AD) is a progressive neurodegenerative disease with memory loss and cognitive decline. Neurofibrillary tangles (NFTs) formed by hyperphosphorylated Tau protein are one of the pathological hallmarks of several neurodegenerative diseases including AD. Heat shock protein family B (small) member 1 (HSPB1) is a molecular chaperone that promotes the correct folding of other proteins in response to environmental stress. Nuclear factor erythroid 2-like 2 (NRF2), a redox-regulated transcription factor, is the master regulator of the cellular response to excess reactive oxygen species. Tropomyosin-related kinase B (TRKB) is a membrane-bound receptor that, upon binding brain-derived neurotrophic factor (BDNF), phosphorylates itself to initiate downstream signaling for neuronal survival and axonal growth. In this study, four natural flavones such as 7,8-dihydroxyflavone (7,8-DHF), wogonin, quercetin, and apigenin were evaluated for Tau aggregation inhibitory activity and neuroprotection in SH-SY5Y neuroblastoma. Among the tested flavones, 7,8-DHF, quercetin, and apigenin reduced Tau aggregation, oxidative stress, and caspase-1 activity as well as improved neurite outgrowth in SH-SY5Y cells expressing ΔK280 TauRD-DsRed folding reporter. Treatments with 7,8-DHF, quercetin, and apigenin rescued the reduced HSPB1 and NRF2 and activated TRKB-mediated extracellular signal-regulated kinase (ERK) signaling to upregulate cAMP-response element binding protein (CREB) and its downstream antiapoptotic BCL2 apoptosis regulator (BCL2). Knockdown of TRKB attenuated the neuroprotective effects of these three flavones. Our results suggest 7,8-DHF, quercetin, and apigenin targeting HSPB1, NRF2, and TRKB to reduce Tau aggregation and protect cells against Tau neurotoxicity and may provide new treatment strategies for AD.

9.
Psychopharmacology (Berl) ; 237(10): 3173-3190, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32748031

RESUMO

RATIONALE: The signaling pathways of tropomyosin-related kinase B (TrkB) receptor play a pivotal role in axonal sprouting, proliferation of dendritic arbor, synaptic plasticity, and neuronal differentiation. The levels of BDNF and TrkB receptor were reduced in patients with Alzheimer's disease (AD). OBJECTIVES: The activation of TrkB signaling pathways is a potential strategy for AD therapies. We intended to identify potential TrkB agonists to activate the neuroprotective signaling to alleviate the pathological features of AD mice. RESULTS: Both of the Aß-deteriorated hippocampal primary neurons and mouse models were generated and showed AD characteristics. We first investigated 12 potential TrkB agonists with primary hippocampal neurons of mice. Both 7,8-DHF and LMDS-1 were identified to have better effect than the other compounds on dendritic arborization of the neurons and were further applied to the Aß-injected mouse model. The short-term cognitive behavior and pathology in the mice were improved by LMDS-1. Further investigation indicated that LMDS-1 activated the TrkB through phosphorylation at Y516 rather than Y816. In addition, the ERK but not CaMKII or Akt was activated in the mouse hippocampus with LMDS-1 administration. LMDS-1 treatment also upregulated CREB and BDNF while downregulated the GSK3ß active form and tau phosphorylation. CONCLUSIONS: This study suggests that LMDS-1 upregulates the expression of BDNF and ameliorates the early-phase phenotypes of the AD-like mice through the pTrkB (Y516)-ERK-CREB pathway. In addition, LMDS-1 has better effect than 7,8-DHF in ameliorating the behavioral and pathological features of AD-like mice.


Assuntos
Doença de Alzheimer/induzido quimicamente , Doença de Alzheimer/prevenção & controle , Peptídeos beta-Amiloides/toxicidade , Fatores de Crescimento Neural/uso terapêutico , Fármacos Neuroprotetores/uso terapêutico , Fragmentos de Peptídeos/toxicidade , Receptor trkB/agonistas , Doença de Alzheimer/metabolismo , Animais , Células Cultivadas , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fatores de Crescimento Neural/farmacologia , Plasticidade Neuronal/efeitos dos fármacos , Plasticidade Neuronal/fisiologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Fármacos Neuroprotetores/farmacologia , Distribuição Aleatória , Receptor trkB/metabolismo
10.
Cells ; 9(5)2020 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-32429337

RESUMO

Parkinson's disease (PD) is a neurodegenerative disease characterized by selective dopaminergic (DAergic) neuronal degeneration in the substantia nigra (SN) and proteinaceous α-synuclein-positive Lewy bodies and Lewy neuritis. As a chemical chaperone to promote protein stability and an autophagy inducer to clear aggregate-prone proteins, a disaccharide trehalose has been reported to alleviate neurodegeneration in PD cells and mouse models. Its trehalase-indigestible analogs, lactulose and melibiose, also demonstrated potentials to reduce abnormal protein aggregation in spinocerebellar ataxia cell models. In this study, we showed the potential of lactulose and melibiose to inhibit α-synuclein aggregation using biochemical thioflavin T fluorescence, cryogenic transmission electron microscopy (cryo-TEM) and prokaryotic split Venus complementation assays. Lactulose and melibiose further reduced α-synuclein aggregation and associated oxidative stress, as well as protected cells against α-synuclein-induced neurotoxicity by up-regulating autophagy and nuclear factor, erythroid 2 like 2 (NRF2) pathway in DAergic neurons derived from SH-SY5Y cells over-expressing α-synuclein. Our findings strongly indicate the potential of lactulose and melibiose for mitigating PD neurodegeneration, offering new drug candidates for PD treatment.


Assuntos
Autofagia/efeitos dos fármacos , Lactulose/farmacologia , Melibiose/farmacologia , Neurônios/metabolismo , Agregados Proteicos , Regulação para Cima , alfa-Sinucleína/metabolismo , Sequência de Aminoácidos , Antioxidantes/farmacologia , Benzotiazóis/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Fluorescência , Proteínas de Fluorescência Verde/metabolismo , Humanos , Lactulose/química , Melibiose/química , Crescimento Neuronal/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Agregados Proteicos/efeitos dos fármacos , Trealose/química , Trealose/farmacologia , Regulação para Cima/efeitos dos fármacos , alfa-Sinucleína/química
11.
Genes Brain Behav ; 18(5): e12559, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30806012

RESUMO

Several reports have shown that methyl CpG-binding protein 2 (MeCP2), brain-derived neurotrophic factor (BDNF), phospho-cAMP response element-binding protein (p-CREB) and microRNAs may be important in regulating academic performance because of their roles in neuropsychiatry and cognitive diseases. The first goal of this study was to explore the associations among MeCP2, BDNF, CREB and academic performance. This study also examined the pathway responsible for the effects of MeCP2, BDNF, p-CREB and microRNAs on academic performance. Scores from the basic competency test, an annual national competitive entrance examination, were used to evaluate academic performance. Subjects' plasma RNA was extracted and analyzed. This study determined that participants in the higher academic performance group had a significant difference in MECP2 mRNA expression compared with the lower academic performance group. We then used neuronal human derived neuroblastoma cell line (SH-SY5Y) cells with inducible MeCP2 expression from a second copy of the gene as a gain-of-function model and found that MeCP2 overexpression positively affected p-CREB and BDNF expression initially. After negative feedback, the p-CREB and BDNF levels subsequently decreased. In the neuronal phenotype examination, we found a significant reduction in total outgrowth and branches in MeCP2-induced cells compared with noninduced cells. This work describes pathways that may be responsible for the effects of MeCP2, BDNF, p-CREB and microRNAs on academic performance. These results may shed light on the development of promising clinical treatment strategies in the area of neuropsychological adjustment.


Assuntos
Desempenho Acadêmico , Epigênese Genética , Adolescente , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Linhagem Celular Tumoral , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Feminino , Mutação com Ganho de Função , Humanos , Masculino , Proteína 2 de Ligação a Metil-CpG/genética , Proteína 2 de Ligação a Metil-CpG/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo
12.
Psychopharmacology (Berl) ; 236(2): 763-773, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30426182

RESUMO

RATIONALE: Hyperglycemia accelerates the progression of Alzheimer's disease (AD), and GSK3ß plays a potential link between AD and hyperglycemia. Therefore, a direct or indirect GSK3ß inhibition may have potential to delay the progression of AD. Our previous biochemical assay identified AM404 as a GSK3ß inhibitor at high dose (IC50 = 5.353 µM); however, other study suggests that AM404 impaired synaptic plasticity of hippocampus at high dose (10 mg/kg; i.p.). Therefore, the dose and duration of treatment are crucial for the effects of AM404. OBJECTIVE: The effects and molecular mechanisms of AM404 at low dose were evaluated from in vitro to in vivo models. METHODS: AM404 (0.1-0.5 µM) was tested on tau hyperphosphorylated mouse hippocampal primary cultures treated with Wortmannin (WT) and GF109203X (GFX). Hyperglycemic triple transgenic AD (3×Tg-AD) mice at 6 months old were intraperitoneally injected with AM404 (0.25 mg/kg) for 4 weeks. The spatial learning and memory of mice were measured using the Morris water maze. Mouse brain and serum samples were collected for pathological analyses. RESULTS: AM404 (0.5 µM) exhibited significantly augmented neuroprotection toward tau hyperphosphorylation in primary cultures. The chronic systemic administration of AM404 (0.25 mg/kg) attenuated cognitive deficits in hyperglycemic 3×Tg-AD mice. Moreover, chronic low dose of AM404 significantly attenuated Aß production, tau protein phosphorylation, and inflammation associated with an increase of pS473Akt and pS9-GSK3ß. Therefore, AM404 at low dose, not only increased neuroprotection, but also ameliorated cognitive deficit, could be partly by regulating the Akt/GSK3ß signaling, which may contribute to downregulation of Aß, tau hyperphosphorylation, and inflammation in hyperglycemic 3×Tg-AD mice. CONCLUSIONS: These results highlight that chronic administration of AM404 at low dose may be through the Akt/GSK3ß pathway to ameliorate the impairment in hyperglycemic 3×Tg-AD mice.


Assuntos
Ácidos Araquidônicos/administração & dosagem , Disfunção Cognitiva/tratamento farmacológico , Hiperglicemia/tratamento farmacológico , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/genética , Peptídeos beta-Amiloides/metabolismo , Animais , Disfunção Cognitiva/genética , Disfunção Cognitiva/metabolismo , Relação Dose-Resposta a Droga , Esquema de Medicação , Feminino , Glicogênio Sintase Quinase 3 beta , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Humanos , Hiperglicemia/genética , Hiperglicemia/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Camundongos Transgênicos , Gravidez , Proteínas tau/genética , Proteínas tau/metabolismo
13.
Neurotoxicology ; 67: 259-269, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29936316

RESUMO

Spinocerebellar ataxia type 17 (SCA17) is caused by the expansion of translated CAG repeat in the TATA box binding protein (TBP) gene encoding a long polyglutamine (polyQ) tract in the TBP protein, which leads to intracellular accumulation of aggregated TBP and cell death. The molecular chaperones act in preventing protein aggregation to ameliorate downstream harmful events. In this study, we used Tet-On cells with inducible SCA17 TBP/Q79-GFP expression to test five in-house NC009 indole compounds for neuroprotection. We found that both aggregation and polyQ-induced reactive oxygen species can be significantly prohibited by the tested NC009 compounds in Tet-On TBP/Q79 293 cells. Among the five indole compounds, NC009-1 up-regulated expression of heat shock protein family B (small) member 1 (HSPB1) chaperone to reduce polyQ aggregation and promote neurite outgrowth in neuronal differentiated TBP/Q79 SH-SY5Y cells. The increased HSPB1 thus ameliorated the increased BH3 interacting domain death agonist (BID), cytochrome c (CYCS) release, and caspase 3 (CASP3) activation which result in apoptosis. Knock down of HSPB1 attenuated the effects of NC009-1 on TBP/Q79 SH-SY5Y cells, suggesting that HSPB1 might be one of the major pathways involved for NC009-1 effects. NC009-1 further reduced polyQ aggregation in Purkinje cells and ameliorated behavioral deficits in SCA17 TBP/Q109 transgenic mice. Our results suggest that NC009-1 has a neuroprotective effect on SCA17 cell and mouse models to support its therapeutic potential in SCA17 treatment.


Assuntos
Proteínas de Choque Térmico/metabolismo , Indóis/uso terapêutico , Transtornos Mentais/tratamento farmacológico , Transtornos Mentais/metabolismo , Proteínas de Neoplasias/metabolismo , Crescimento Neuronal/efeitos dos fármacos , Proteína de Ligação a TATA-Box/metabolismo , Animais , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Proteínas de Choque Térmico/agonistas , Humanos , Indóis/química , Indóis/farmacologia , Camundongos , Camundongos Transgênicos , Chaperonas Moleculares , Proteínas de Neoplasias/agonistas , Crescimento Neuronal/fisiologia , Proteína de Ligação a TATA-Box/genética
14.
Eur J Pharm Sci ; 89: 11-9, 2016 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-27094783

RESUMO

Glycogen synthase kinase 3ß (GSK-3ß) is widely known as a critical target protein for treating Alzheimer's disease (AD). We utilized virtual screening to search databases for compounds with the potential to be used in drugs targeting GSK-3ß kinase, and kinase as well as cell assays to investigate top-scored, selected compounds. Virtual screening of >1.1 million compounds in the ZINC and in-house databases was conducted using an optimized computational protocol in the docking program GOLD. Of the top-ranked compounds, 16 underwent a luminescent kinase assay and a cell assay using HEK293 cells expressing DsRed-tagged ΔK280 in the repeat domain of tau (tauRD). The compounds VB-003 (a potent GSK-3ß inhibitor) and VB-008 (AM404, an anandamide transport inhibitor), with determined IC50 values of 0.25 and 5.4µM, respectively, were identified as reducing tau aggregation. Both compounds increased expression of phospho-GSK-3ß (Ser9) and reduced endogenous tau phosphorylation at the sites of Ser202, Thr231, and Ser396. In the ∆K280 tauRD-DsRed SH-SY5Y cells, VB-008, but not VB-003, enhanced HSPB1 and GRP78 expression, increased ∆K280 tauRD-DsRed solubility, and promoted neurite outgrowth. Thus VB-008 performed best to the end of the present study. The identified compound VB-008 may guide the identification and synthesis of potential inhibitors analogous to this compound.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Inibidores Enzimáticos/farmacologia , Glicogênio Sintase Quinase 3 beta/antagonistas & inibidores , Doença de Alzheimer/metabolismo , Linhagem Celular , Chaperona BiP do Retículo Endoplasmático , Células HEK293 , Humanos , Neuritos/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Proteínas tau/metabolismo
15.
Brain Res ; 1639: 132-48, 2016 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-26972528

RESUMO

Spinocerebellar ataxia type 17 (SCA17), an autosomal dominant cerebellar ataxia, is a devastating, incurable disease caused by the polyglutamine (polyQ) expansion of transcription factor TATA binding protein (TBP). The polyQ expansion causes misfolding and aggregation of the mutant TBP, further leading to cytotoxicity and cell death. The well-recognized prodromal phase in many forms of neurodegeneration suggests a prolonged period of partial neuronal dysfunction prior to cell loss that may be amenable to therapeutic intervention. The objective of this study was to assess the effects and molecular mechanisms of granulocyte-colony stimulating factor (G-CSF) therapy during the pre-symptomatic stage in SCA17 mice. Treatment with G-CSF at the pre-symptomatic stage improved the motor coordination of SCA17 mice and reduced the cell loss, insoluble mutant TBP protein, and vacuole formation in the Purkinje neurons of these mice. The neuroprotective effects of G-CSF may be produced by increases in Hsp70, Beclin-1, LC3-II and the p-ERK survival pathway. Upregulation of chaperone and autophagy levels further enhances the clearance of mutant protein aggregation, slowing the progression of pathology in SCA17 mice. Therefore, we showed that the early intervention of G-CSF has a neuroprotective effect, delaying the progression of SCA17 in mutant mice via increases in the levels of chaperone expression and autophagy.


Assuntos
Fator Estimulador de Colônias de Granulócitos/farmacologia , Fármacos Neuroprotetores/farmacologia , Sintomas Prodrômicos , Ataxias Espinocerebelares/tratamento farmacológico , Animais , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Autofagia/efeitos dos fármacos , Autofagia/fisiologia , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Camundongos Transgênicos , Proteínas Associadas aos Microtúbulos/metabolismo , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Células de Purkinje/efeitos dos fármacos , Células de Purkinje/patologia , Células de Purkinje/fisiologia , Ataxias Espinocerebelares/patologia , Ataxias Espinocerebelares/fisiopatologia
16.
PLoS One ; 9(12): e115809, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25549101

RESUMO

Spinocerebellar ataxia type 17 (SCA17) involves the expression of a polyglutamine (polyQ) expanded TATA-binding protein (TBP), a general transcription initiation factor. TBP interacts with other protein factors, including high mobility group box 1 (HMGB1), to regulate gene expression. Previously, our proteomic analysis of soluble proteins prepared from mutant TBP (TBP/Q61) expressing cells revealed a reduced concentration of HMGB1. Here, we show that HMGB1 can be incorporated into mutant TBP aggregates, which leads to reduced soluble HMGB1 levels in TBP/Q(61∼79) expressing cells. HMGB1 overexpression reduced mutant TBP aggregation. HMGB1 cDNA and siRNA co-transfection, as well as an HSPA5 immunoblot and luciferase reporter assay demonstrated the important role of HMGB1 in the regulation of HSPA5 transcription. In starvation-stressed TBP/Q36 and TBP/Q79 cells, increased reactive oxygen species generation accelerated the cytoplasmic translocation of HMGB1, which accompanied autophagy activation. However, TBP/Q79 cells displayed a decrease in autophagy activation as a result of the reduction in the cytoplasmic HMGB1 level. In neuronal SH-SY5Y cells with induced TBP/Q(61∼79) expression, HMGB1 expression was reduced and accompanied by a significant reduction in the total outgrowth and branches in the TBP/Q(61∼79) expressing cells compared with the non-induced cells. The decreased soluble HMGB1 and impaired starvation-induced autophagy in cells suggest that HMGB1 may be a critical modulator of polyQ disease pathology and may represent a target for drug development.


Assuntos
Regulação da Expressão Gênica , Proteína HMGB1/fisiologia , Ataxias Espinocerebelares/genética , Autofagia/genética , Citoplasma/metabolismo , Chaperona BiP do Retículo Endoplasmático , Células HEK293 , Proteína HMGB1/genética , Proteína HMGB1/metabolismo , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Estresse Oxidativo , Proteína de Ligação a TATA-Box/genética , Proteína de Ligação a TATA-Box/metabolismo , Proteína de Ligação a TATA-Box/fisiologia
17.
Drug Des Devel Ther ; 8: 1929-39, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25342886

RESUMO

In spinocerebellar ataxia type 17 (SCA17), the expansion of a translated CAG repeat in the TATA box binding protein (TBP) gene results in a long polyglutamine (polyQ) tract in the TBP protein, leading to intracellular accumulation of aggregated TBP and cell death. The molecular chaperones act in preventing protein aggregation to ameliorate downstream harmful events. In this study, we used Tet-On SH-SY5Y cells with inducible SCA17 TBP/Q79-green fluorescent protein (GFP) expression to test indole and synthetic derivative NC001-8 for neuroprotection. We found that indole and NC001-8 up-regulated chaperone expression to reduce polyQ aggregation in neuronal differentiated TBP/Q79 cells. The effects on promoting neurite outgrowth and on reduction of aggregation on Purkinje cells were also confirmed with cerebellar primary and slice cultures of SCA17 transgenic mice. Our results demonstrate how indole and derivative NC001-8 reduce polyQ aggregation to support their therapeutic potentials in SCA17 treatment.


Assuntos
Antineoplásicos/farmacologia , Indóis/farmacologia , Chaperonas Moleculares/metabolismo , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Peptídeos/metabolismo , Agregação Patológica de Proteínas/tratamento farmacológico , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Técnicas In Vitro , Indóis/síntese química , Indóis/química , Camundongos , Chaperonas Moleculares/biossíntese , Neurônios/citologia , Neurônios/metabolismo , Neurônios/patologia , Fármacos Neuroprotetores/síntese química , Fármacos Neuroprotetores/química , Técnicas de Cultura de Órgãos , Peptídeos/química , Agregação Patológica de Proteínas/patologia , Agregação Patológica de Proteínas/prevenção & controle , Ligação Proteica/efeitos dos fármacos , Relação Estrutura-Atividade , Células Tumorais Cultivadas
18.
Hum Mol Genet ; 23(25): 6878-93, 2014 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-25104854

RESUMO

Spinocerebellar ataxia type 17 (SCA17) is an autosomal dominant cerebellar ataxia caused by the expansion of polyglutamine (polyQ) within the TATA box-binding protein (TBP). Previous studies have shown that polyQ-expanded TBP forms neurotoxic aggregates and alters downstream genes. However, how expanded polyQ tracts affect the function of TBP and the link between dysfunctional TBP and SCA17 is not clearly understood. In this study, we generated novel Drosophila models for SCA17 that recapitulate pathological features such as aggregate formation, mobility defects and premature death. In addition to forming neurotoxic aggregates, we determined that polyQ-expanded TBP reduces its own intrinsic DNA-binding and transcription abilities. Dysfunctional TBP also disrupts normal TBP function. Furthermore, heterozygous dTbp amorph mutant flies exhibited SCA17-like phenotypes and flies expressing polyQ-expanded TBP exhibited enhanced retinal degeneration, suggesting that loss of TBP function may contribute to SCA17 pathogenesis. We further determined that the downregulation of TBP activity enhances retinal degeneration in SCA3 and Huntington's disease fly models, indicating that the deactivation of TBP is likely to play a common role in polyQ-induced neurodegeneration.


Assuntos
Drosophila melanogaster/genética , Doença de Huntington/genética , Doença de Machado-Joseph/genética , Degeneração Retiniana/genética , Ataxias Espinocerebelares/genética , Proteína de Ligação a TATA-Box/genética , Animais , Modelos Animais de Doenças , Drosophila melanogaster/metabolismo , Regulação da Expressão Gênica , Heterozigoto , Humanos , Doença de Huntington/metabolismo , Doença de Huntington/patologia , Longevidade/genética , Doença de Machado-Joseph/metabolismo , Doença de Machado-Joseph/patologia , Peptídeos/química , Fenótipo , Agregados Proteicos , Ligação Proteica , Degeneração Retiniana/metabolismo , Degeneração Retiniana/patologia , Transdução de Sinais , Ataxias Espinocerebelares/metabolismo , Ataxias Espinocerebelares/patologia , Proteína de Ligação a TATA-Box/química , Proteína de Ligação a TATA-Box/metabolismo
19.
J Biomed Opt ; 19(1): 011014, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24002232

RESUMO

Prior studies have established the necessity of an angiotensin-converting enzyme-related (ACER) gene for heart morphogenesis of Drosophila. Nevertheless, the physiology of ACER has yet to be comprehensively understood. Herein, we employed RNA interference to down-regulate the expression of ACER in Drosophila's heart and swept source optical coherence tomography to assess whether ACER is required for cardiac functions in living adult flies. Several contractile parameters of Drosophila heart, including the heart rate (HR), end-diastolic diameter (EDD), end-systolic diameter (ESD), percent fractional shortening (%FS), and stress-induced cardiac performance, are shown, which are age dependent. These age-dependent cardiac functions declined significantly when ACER was down-regulated. Moreover, the lifespans of ACER knock-down flies were significantly shorter than those of wild-type control flies. Thus, we posit that ACER, the Drosophila ortholog of mammalian angiotensin-converting enzyme 2 (ACE2), is essential for both heart physiology and longevity of animals. Since mammalian ACE2 controls many cardiovascular physiological features and is implicated in cardiomyopathies, our findings that ACER plays conserved roles in genetically tractable animals will pave the way for uncovering the genetic pathway that controls the renin-angiotensin system.


Assuntos
Drosophila melanogaster/fisiologia , Peptidil Dipeptidase A/fisiologia , Tomografia de Coerência Óptica/métodos , Animais , Drosophila melanogaster/enzimologia , Drosophila melanogaster/genética , Coração/fisiopatologia , Longevidade/fisiologia , Masculino , Contração Miocárdica/fisiologia , Peptidil Dipeptidase A/genética , Peptidil Dipeptidase A/metabolismo
20.
PLoS One ; 8(9): e73885, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24040107

RESUMO

Spinocerebellar ataxia type 8 (SCA8) involves the expansion of CTG/CAG repeats from the overlapping ataxin 8 opposite strand (ATXN8OS) and ataxin 8 (ATXN8) genes located on chromosome 13q21. Although being transcribed, spliced and polyadenylated in the CTG orientation, ATXN8OS does not itself appear to be protein coding, as only small open reading frames (ORFs) were noted. In the present study we investigated the translation of a novel 102 amino acids containing-ORF in the ATXN8OS RNA. Expression of chimeric construct with an in-frame ORF-EGFP gene demonstrated that ATXN8OS RNA is translatable. Using antiserum raised against ORF, ATXN8OS ORF expression was detected in various human cells including lymphoblastoid, embryonic kidney 293, neuroblastoma IMR-32, SK-N-SH, SH-SY5Y cells and human muscle tissue. The biological role of the ATXN8OS ORF and its connection to SCA8 remains to be determined.


Assuntos
RNA Longo não Codificante/genética , RNA/genética , RNA/metabolismo , Linhagem Celular , Expressão Gênica , Ordem dos Genes , Humanos , Fases de Leitura Aberta , RNA Longo não Codificante/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Sequências Repetitivas de Ácido Nucleico
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