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1.
Sci Rep ; 8(1): 7377, 2018 05 09.
Artigo em Inglês | MEDLINE | ID: mdl-29743517

RESUMO

The receptor tyrosine kinase, erythropoietin-producing hepatocellular A4 (EphA4), was recently identified as a molecular target for Alzheimer's disease (AD). We found that blockade of the interaction of the receptor and its ligands, ephrins, alleviates the disease phenotype in an AD transgenic mouse model, suggesting that targeting EphA4 is a potential approach for developing AD interventions. In this study, we identified five FDA-approved drugs-ergoloid, cyproheptadine, nilotinib, abiraterone, and retapamulin-as potential inhibitors of EphA4 by using an integrated approach combining virtual screening with biochemical and cellular assays. We initially screened a database of FDA-approved drugs using molecular docking against the ligand-binding domain of EphA4. Then, we selected 22 candidate drugs and examined their inhibitory activity towards EphA4. Among them, five drugs inhibited EphA4 clustering induced by ephrin-A in cultured primary neurons. Specifically, nilotinib, a kinase inhibitor, inhibited the binding of EphA4 and ephrin-A at micromolar scale in a dosage-dependent manner. Furthermore, nilotinib inhibited the activation of EphA4 and EphA4-dependent growth cone collapse in cultured hippocampal neurons, demonstrating that the drug exhibits EphA4 inhibitory activity in cellular context. As demonstrated in our combined computational and experimental approaches, repurposing of FDA-approved drugs to inhibit EphA4 may provide an alternative fast-track approach for identifying and developing new treatments for AD.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Avaliação Pré-Clínica de Medicamentos , Simulação de Acoplamento Molecular , Pirimidinas/farmacologia , Receptor EphA4/antagonistas & inibidores , Doença de Alzheimer/metabolismo , Androstenos/metabolismo , Androstenos/farmacologia , Animais , Compostos Bicíclicos Heterocíclicos com Pontes/metabolismo , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Ciproeptadina/metabolismo , Ciproeptadina/farmacologia , Modelos Animais de Doenças , Diterpenos/metabolismo , Diterpenos/farmacologia , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Ligantes , Camundongos , Camundongos Transgênicos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Ligação Proteica , Domínios Proteicos , Pirimidinas/metabolismo , Receptor EphA4/metabolismo
2.
J Agric Food Chem ; 65(31): 6608-6614, 2017 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-28707886

RESUMO

Two new diarylheptanoids, alpinin A (1) and alpinin B (2), together with 18 known diarylheptanoids (3-20), were isolated from the rhizomes of Alpinia officinarum. Their structures were elucidated by comprehensive spectroscopic analysis, including high-resolution mass spectrometry, infrared spectroscopy, and one- and two-dimensional nuclear magnetic resonance spectroscopy. Structurally, alpinin A is a new member of the small family of oxa-bridged diarylheptanoids and contains the characteristic 2,6-cis-configured tetrahydropyran motif (C1-C5 oxa bridge). The absolute configuration of alpinin A was confirmed by asymmetric total synthesis of the enantiomer (ent-1), corroborating the assignment of the molecular structure. The absolute configuration of alpinin B was determined on the basis of the analysis of the circular dichroism exciton chirality spectrum. We evaluated the inhibitory activity of all isolated diarylheptanoids against α-synuclein aggregation at 10 µM. Alpinins A and B significantly inhibited α-synuclein aggregation by 66 and 67%, respectively.


Assuntos
Alpinia/química , Diarileptanoides/química , Extratos Vegetais/química , Rizoma/química , alfa-Sinucleína/química , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Agregados Proteicos , Estereoisomerismo
3.
ACS Chem Neurosci ; 7(10): 1442-1451, 2016 10 19.
Artigo em Inglês | MEDLINE | ID: mdl-27467236

RESUMO

Neurite outgrowth is crucial during neuronal development and regeneration, and strategies that aim at promoting neuritogenesis are beneficial for reconstructing synaptic connections after neuronal degeneration and injury. Using a bivalent analogue strategy as a successful approach, the current study identifies a series of novel dimeric securinine analogues as potent neurite outgrowth enhancers. Compounds 13, 14, 17-19, and 21-23, with different lengths of carbon chain of N,N-dialkyl substituting diacid amide linker between two securinine molecules at C-15 position, exhibited notable positive effects on both neuronal differentiation and neurite extension of neuronal cells. Compound 14, one of the most active compounds, was used as a representative compound for mechanistic studies. Its action on neurite outgrowth was through phosphorylation/activation of multiple signaling molecules including Ca2+/calmodulin-dependent protein kinase II (CaMKII), extracellular signal-regulated kinase (ERK) and Akt. These findings collectively identify a new group of beneficial compounds for neuritogenesis, and may provide insights on drug discovery of neural repair and regeneration.


Assuntos
Azepinas/síntese química , Azepinas/farmacologia , Crescimento Celular/efeitos dos fármacos , Compostos Heterocíclicos de Anel em Ponte/síntese química , Compostos Heterocíclicos de Anel em Ponte/farmacologia , Neuritos/efeitos dos fármacos , Fármacos Neuroprotetores/síntese química , Fármacos Neuroprotetores/farmacologia , Animais , Azepinas/química , Western Blotting , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Linhagem Celular Tumoral , Desenho de Fármacos , Avaliação Pré-Clínica de Medicamentos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Compostos Heterocíclicos de Anel em Ponte/química , Imuno-Histoquímica , Lactonas/química , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/fisiologia , Camundongos , Estrutura Molecular , Neuritos/fisiologia , Fármacos Neuroprotetores/química , Fosforilação/efeitos dos fármacos , Piperidinas/química , Proteínas Proto-Oncogênicas c-akt/metabolismo
4.
Neurochem Int ; 89: 260-70, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26220901

RESUMO

Alzheimer's disease (AD) is a neurodegenerative illness associated with dementia and is most prevalent among the elderly population. Current medications can only treat symptoms. Alkaloids are structurally diverse and have been an important source of therapeutics for various brain disorders. Two US Food and Drug Administration (FDA)-approved acetylcholinesterase inhibitors for AD, galantamine and rivastigmine, are in fact alkaloids. In addition, clinical trials of four other extensively studied alkaloids-huperzine A, caffeine, nicotine, and indomethacin-have been conducted but do not convincingly demonstrate their clinical efficacy for AD. Interestingly, rhynchophylline, a known neuroprotective alkaloid, was recently discovered by in silico screening as an inhibitor of EphA4, a novel target for AD. Here, we review the pathophysiological mechanisms underlying AD, current treatment strategies, and therapeutic potential of several selected plant alkaloids in AD, highlighting their various drug targets and the key supportive preclinical and clinical studies. Future research should include more rigorous clinical studies of the most promising alkaloids, the further development of recently discovered candidate alkaloids, and the continual search for new alkaloids for relevant drug targets. It remains promising that an alkaloid drug candidate could significantly affect the progression of AD in addition to providing symptomatic relief.


Assuntos
Alcaloides/administração & dosagem , Alcaloides/metabolismo , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/metabolismo , Descoberta de Drogas/tendências , Fitoterapia/tendências , Alcaloides/isolamento & purificação , Animais , Descoberta de Drogas/métodos , Humanos , Fitoterapia/métodos
5.
Neuropsychopharmacology ; 40(8): 1877-87, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25649278

RESUMO

Compounds that have the ability to both strengthen synaptic function and facilitate neuroprotection are valuable cognitive enhancers that may improve health and quality of life, as well as retard age-related cognitive deterioration. Medicinal plants are an abundant source of potential cognitive enhancers. Here we report that anemoside A3 (AA3) isolated from Pulsatilla chinensis modulates synaptic connectivity in circuits central to memory enhancement. AA3 specifically modulates the function of AMPA-type glutamate receptors (AMPARs) by increasing serine phosphorylation within the GluA1 subunit, which is a modification required for the trafficking of GluA1-containing AMPARs to synapses. Furthermore, AA3 administration activates several synaptic signaling molecules and increases protein expressions of the neurotrophin brain-derived neurotrophic factor and monoamine neurotransmitters in the mouse hippocampus. In addition to acting through AMPARs, AA3 also acts as a non-competitive NMDA receptor (NMDAR) modulator with a neuroprotective capacity against ischemic brain injury and overexcitation in rats. These findings collectively suggest that AA3 possesses a unique ability to modulate the functions of both AMPARs and NMDARs. Concordantly, behavioral studies indicate that AA3 not only facilitates hippocampal long-term potentiation but also enhances spatial reference memory formation in mice. These multifaceted roles suggest that AA3 is an attractive candidate for further development as a cognitive enhancer capable of alleviating memory dysfunctions associated with aging and neurodegenerative diseases.


Assuntos
Cognição/efeitos dos fármacos , Hipocampo , Fármacos Neuroprotetores/farmacologia , Córtex Pré-Frontal/efeitos dos fármacos , Saponinas/farmacologia , Sinapses/efeitos dos fármacos , Triterpenos/farmacologia , Animais , Modelos Animais de Doenças , Reação de Fuga/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Comportamento Exploratório/efeitos dos fármacos , Hipocampo/citologia , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Técnicas In Vitro , Infarto da Artéria Cerebral Média/tratamento farmacológico , Infarto da Artéria Cerebral Média/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Aprendizagem em Labirinto/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , N-Metilaspartato/farmacologia , Rede Nervosa/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Navegação Espacial/efeitos dos fármacos
6.
Neurobiol Dis ; 73: 366-76, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25448764

RESUMO

Despite advances in promoting axonal regeneration after adult central nervous system injury, elicitation of a large number of lesion-passing axons reform active synaptic connections with natural target neurons remains limited. By deleting both Pten and Socs3 in retinal ganglion cells, we report that optic nerve axons after prechiasm lesion robustly reinnervate the hypothalamus, form new synapses with neurons in the suprachiasmatic nucleus (SCN), and re-integrate with the existing circuitry. Photic or electric stimulation of the retinal axons induces neuronal response in SCN. However both the innervation pattern and evoked responses are not completely restored by the regenerating axons, suggesting that combining with other strategies is necessary to overcome the defective rewiring. Our results support that boosting the intrinsic growth capacity in injured neurons promotes axonal reinnervation and rewiring.


Assuntos
Axônios/fisiologia , Hipotálamo/fisiologia , Regeneração Nervosa/fisiologia , Nervo Óptico/fisiologia , PTEN Fosfo-Hidrolase , Células Ganglionares da Retina/fisiologia , Proteínas Supressoras da Sinalização de Citocina , Núcleo Supraquiasmático/fisiologia , Sinapses/fisiologia , Animais , Axônios/patologia , Hipotálamo/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Nervo Óptico/patologia , PTEN Fosfo-Hidrolase/genética , Células Ganglionares da Retina/patologia , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina/genética , Núcleo Supraquiasmático/patologia , Sinapses/patologia
7.
PLoS One ; 8(5): e63580, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23696836

RESUMO

Oleanolic acid (3ß-hydroxy-olea-12-en-28-oic acid) is a natural pentacyclic triterpenoic acid found in many fruits, herbs and medicinal plants. In the past decade, increasing evidence has suggested that oleanolic acid exhibits inhibitory activities against different types of cancer including skin cancer and colon cancer, but not leukemia. We report here that a derivative of oleanolic acid, olean-12-eno[2,3-c] [1], [2], [5]oxadiazol-28-oic acid (designated OEOA) effectively blocks the proliferation of human leukemia cells. OEOA significantly reduces cell proliferation without inducing cell death in three types of leukemia cell lines, including K562, HEL and Jurket. Moreover, exposure of K562 cells to OEOA results in G1 cell cycle arrest, with a concomitant induction of cyclin-dependent kinase inhibitor p27 and downregulation of cyclins and Cdks that are essential for cell cycle progression. Interestingly, OEOA also enhances erythroid differentiation in K562 cells through suppressing the expression of Bcr-Abl and phosphorylation of Erk1/2. These findings identify a novel chemical entity for further development as therapeutics against leukemia.


Assuntos
Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Ácido Oleanólico/farmacologia , Western Blotting , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Proteínas de Fusão bcr-abl/metabolismo , Humanos , Leucemia/metabolismo , Ácido Oleanólico/análogos & derivados , Fosforilação/efeitos dos fármacos
8.
Planta Med ; 78(2): 115-21, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22083896

RESUMO

Two Chinese herb-derived small molecule telomerase activators, astragaloside IV (AG-IV) and cycloastragenol (CAG), have recently been shown to improve the proliferative response of CD8+ T lymphocytes from HIV-infected patients by upregulating telomerase activity. Here, we examined the signaling mechanism of AG-IV and CAG. Telomerase activity in human embryonic kidney HEK293 fibroblasts was increased upon treatment with increasing concentrations of AG-IV or CAG. Both compounds induced the phosphorylation of extracellular signal-regulated protein kinase (ERK) in a time- and dose-dependent manner in HEK293 cells and HEK-neo keratinocytes. AG-IV and CAG also stimulated ERK phosphorylation in other cell lines of lung, brain, mammary, endothelial, and hematopoietic origins. Use of selective inhibitors and dominant negative mutants revealed the involvement of c-Src, MEK (ERK kinase), and epidermal growth factor receptor in CAG-induced ERK phosphorylation. Our data indicate that AG-IV and CAG may exert their cellular effects through the activation of the Src/MEK/ERK pathway.


Assuntos
Astrágalo/química , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fibroblastos/efeitos dos fármacos , Extratos Vegetais/farmacologia , Sapogeninas/farmacologia , Saponinas/farmacologia , Telomerase/metabolismo , Triterpenos/farmacologia , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Mama/efeitos dos fármacos , Mama/metabolismo , Linhagem Celular , Relação Dose-Resposta a Droga , Células Endoteliais/efeitos dos fármacos , Receptores ErbB/metabolismo , Humanos , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Fosforilação , Quinases da Família src/metabolismo
9.
J Neurosci ; 30(43): 14366-70, 2010 Oct 27.
Artigo em Inglês | MEDLINE | ID: mdl-20980593

RESUMO

Precise regulation of cyclin-dependent kinase 5 (Cdk5), a member of the cyclin-dependent kinase family, is critical for proper neuronal development and functions. Cdk5 is activated through its association with the neuron-specific activator p35 or p39. Nonetheless, how its kinase activity is regulated in neurons is not well understood. In this study, we found that Cdk5 activity is regulated by S-nitrosylation, a post-translational modification of protein that affects a plethora of neuronal functions. S-nitrosylation of Cdk5 occurs at Cys83, which is one of the critical amino acids within the ATP-binding pocket of the kinase. Upon S-nitrosylation, Cdk5 exhibits reduced kinase activity, whereas mutation of Cys83 to Ala on Cdk5 renders the kinase refractory to such inhibition. Importantly, S-nitrosylated Cdk5 can be detected in the mouse brain, and blocking the S-nitrosylation of Cdk5 in cultured hippocampal neurons enhances dendritic growth and branching. Together, our findings reveal an important role of S-nitrosylation in regulating Cdk5 kinase activity and dendrite growth in neurons during development.


Assuntos
Quinase 5 Dependente de Ciclina/genética , Quinase 5 Dependente de Ciclina/fisiologia , Dendritos/fisiologia , Neurônios/fisiologia , Processamento de Proteína Pós-Traducional/genética , Processamento de Proteína Pós-Traducional/fisiologia , Animais , Biotina , Química Encefálica/fisiologia , Células Cultivadas , Cisteína/fisiologia , DNA Complementar/genética , Hipocampo/citologia , Hipocampo/crescimento & desenvolvimento , Humanos , Camundongos , Compostos Nitrosos/química , Proteínas Recombinantes de Fusão , Transfecção
10.
Planta Med ; 76(10): 998-1003, 2010 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-20143293

RESUMO

Two new secoiridoid glucosides, namely iso-oleonuezhenide (1) and methyloleoside 7-ethyl ester (2), along with five known ones, oleonuezhenide (3), nuezhenide (4), oleuropein (5), G13 (6), and jaspolyside methyl ester (7), were isolated from the fruits of Ligustrum lucidum. Their structures were assigned based on 1H-NMR, 13C-NMR, and 2D-NMR analyses, in combination with HR-MS experiments and the comparison with literature data of related compounds, as well as on chemical experiments. We have examined the ability of these compounds to activate ERK and CREB in cultured cortical neurons. Our studies demonstrate that compound 1 induces ERK and CREB phosphorylation in primary cortical neurons in a dose- and temporal-dependent manner, suggesting its bioactivity on neurons.


Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Glucosídeos Iridoides/farmacologia , Ligustrum/química , Neurônios/efeitos dos fármacos , Extratos Vegetais/farmacologia , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Técnicas de Cultura de Células , Relação Dose-Resposta a Droga , Frutas , Glucosídeos Iridoides/química , Glucosídeos Iridoides/isolamento & purificação , Estrutura Molecular , Neurônios/metabolismo , Fosforilação , Extratos Vegetais/química , Ratos , Ratos Sprague-Dawley
11.
Planta Med ; 73(12): 1298-303, 2007 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-17893832

RESUMO

In this study five new limonoids, toosendone [24,25,26,27-tetra-nor-6alpha-acetoxy-21,22-epoxy-7alpha-tigloyl-1alpha,3alpha,28-trihydroxyapotirucalla-(apoeupha)-14,20,22-trien-12-one, 1] and 12-ethoxynimbolinins A-D (2-5), together with five known limonoids, 1-acetyltrichilinin (6), 1-cinnamoyltrichilinin (7), trichilinin B (8), 1,7-di-O-acetyl-14,15-deoxyhavanensin (9) and 12-O-methylnimbolinin B (10),were isolated from the fruits of Melia toosendan. Their structures and relative configurations were established based on spectroscopic analysis. Compound 4 exhibited significant antibacterial activity against the oral pathogen, Porphyromonas gingivalis ATCC 33 277, with an MIC value of 15.6 microg/mL. Compounds 7 and 8 were also active against P. gingivalis ATCC 33 277, with MIC values of 31.3 and 31.5 microg/mL respectively.


Assuntos
Antibacterianos/farmacologia , Limoninas/farmacologia , Melia/química , Porphyromonas gingivalis/efeitos dos fármacos , Streptococcus mutans/efeitos dos fármacos , Antibacterianos/química , Antibacterianos/isolamento & purificação , Frutas/química , Humanos , Limoninas/química , Limoninas/isolamento & purificação , Testes de Sensibilidade Microbiana , Estrutura Molecular , Boca/microbiologia , Plantas Medicinais/química
12.
Planta Med ; 73(1): 84-90, 2007 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17315313

RESUMO

There were five new diterpenoids, 18-beta-D-3',4'-diacetoxyxylopyranosyl-ent-kaur-16-ene (1), 18-beta-L-3',5'-diacetoxyarabinofuranosyl-ent-kaur-16-ene (2), 18-beta-D-3',6'-diacetoxyglucopyranosyl-ent-kaur-16-ene (3), ent-isopimar-8(14),15-dien-19-oic acid (4), and 5alpha-hydroxy-ent-rosa-15-en-18-oic acid (5), isolated from the whole herb of Sagittaria pygmaea. Their structures and relative configurations were established based on spectroscopic studies, chemical methods, and X-ray crystallographic analysis. Compound 2 exhibited significant antibacterial activity against the oral pathogens, Streptococcus mutans ATCC 25 175 and Actinomyces viscosus ATCC 27 044, with MIC values against both pathogens of 15.6 microg/mL. Compound 3 was active against only A. viscosus ATCC 27 044 with an MIC value of 62.5 microg/mL. Compounds 4 and 5 were active against S. mutans ATCC 25 175 and A. viscosus ATCC 27 044, with MIC values against both pathogens of 125.0 microg/mL.


Assuntos
Aggregatibacter actinomycetemcomitans/efeitos dos fármacos , Antibacterianos/farmacologia , Bactérias Gram-Positivas/efeitos dos fármacos , Fitoterapia , Extratos Vegetais/farmacologia , Sagittaria , Actinomyces viscosus/efeitos dos fármacos , Antibacterianos/administração & dosagem , Antibacterianos/química , Antibacterianos/uso terapêutico , Diterpenos/administração & dosagem , Diterpenos/química , Diterpenos/farmacologia , Diterpenos/uso terapêutico , Humanos , Testes de Sensibilidade Microbiana , Extratos Vegetais/administração & dosagem , Extratos Vegetais/química , Extratos Vegetais/uso terapêutico , Streptococcus mutans/efeitos dos fármacos
13.
J Biol Chem ; 281(15): 9852-8, 2006 Apr 14.
Artigo em Inglês | MEDLINE | ID: mdl-16461345

RESUMO

Pctaire1, a member of the cyclin-dependent kinase (Cdk)-related family, has recently been shown to be phosphorylated and regulated by Cdk5/p35. Although Pctaire1 is expressed in both neuronal and non-neuronal cells, its precise functions remain elusive. We performed a yeast two-hybrid screen to identify proteins that interact with Pctaire1. N-Ethylmaleimide-sensitive fusion protein (NSF), a crucial factor in vesicular transport and membrane fusion, was identified as one of the Pctaire1 interacting proteins. We demonstrate that the D2 domain of NSF, which is required for the oligomerization of NSF subunits, binds directly to and is phosphorylated by Pctaire1 on serine 569. Mutation of this phosphorylation site on NSF (S569A) augments its ability to oligomerize. Moreover, inhibition of Pctaire1 activity by transfecting its kinase-dead (KD) mutant into COS-7 cells enhances the self-association of NSF. Interestingly, Pctaire1 associates with NSF and synaptic vesicle-associated proteins in adult rat brain. To investigate whether Pctaire1 phosphorylation of NSF is involved in regulation of Ca(2+)-dependent exocytosis, we examined the effect of expressing Pctaire1 or NSF phosphorylation mutants on the regulated secretion of growth hormone from PC12 cells. Interestingly, expression of either Pctaire1-KD or NSF-S569A in PC12 cells significantly increases high K(+)-stimulated growth hormone release. Taken together, our findings provide the first demonstration that Pctaire1 phosphorylation of NSF regulates the ability of NSF to oligomerize, implicating an unexpected role of this kinase in modulating exocytosis. These findings open a new avenue of research in studying the functional roles of Pctaire1 in the nervous system.


Assuntos
Quinases Ciclina-Dependentes/fisiologia , Proteínas Sensíveis a N-Etilmaleimida/fisiologia , Proteínas Serina-Treonina Quinases/fisiologia , Animais , Western Blotting , Células COS , Cálcio/metabolismo , Sistema Nervoso Central/metabolismo , Chlorocebus aethiops , Clonagem Molecular , Quinases Ciclina-Dependentes/química , DNA Complementar/metabolismo , Dimerização , Relação Dose-Resposta a Droga , Exocitose , Humanos , Imunoprecipitação , Proteínas Sensíveis a N-Etilmaleimida/química , Proteínas Sensíveis a N-Etilmaleimida/metabolismo , Células PC12 , Fosforilação , Plasmídeos/metabolismo , Ligação Proteica , Proteínas Serina-Treonina Quinases/química , Estrutura Terciária de Proteína , Ratos , Serina/química , Transfecção , Técnicas do Sistema de Duplo-Híbrido , beta-Galactosidase/metabolismo
14.
J Nat Prod ; 69(2): 255-60, 2006 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-16499326

RESUMO

Seven new ent-rosane diterpenoids, sagittines A-G (1-7), together with one new labdane diterpene, 13-epi-manoyl oxide-19-O-alpha-l-2',5'-diacetoxyarabinofuranoside (8), were isolated from the whole plant of Sagittaria sagittifolia. The structures and relative configurations of 1-8 were characterized using spectroscopic means, chemical methods, and X-ray crystallography. Compounds 1-4 exhibited antibacterial activity against the oral pathogens Streptococcus mutans ATCC 25175 and Actinomyces naeslundiis ATCC 12104, with MIC values between 62.5 and 125 microg/mL. Compound 5 was active against only A. naeslundiis ATCC 12104, with an MIC value of 62.5 microg/mL.


Assuntos
Antibacterianos/isolamento & purificação , Antibacterianos/farmacologia , Arabinose/análogos & derivados , Diterpenos/isolamento & purificação , Diterpenos/farmacologia , Medicamentos de Ervas Chinesas/isolamento & purificação , Medicamentos de Ervas Chinesas/farmacologia , Plantas Medicinais/química , Sagittaria/química , Actinomyces/efeitos dos fármacos , Antibacterianos/química , Arabinose/química , Arabinose/isolamento & purificação , Arabinose/farmacologia , Cristalografia por Raios X , Diterpenos/química , Medicamentos de Ervas Chinesas/química , Testes de Sensibilidade Microbiana , Conformação Molecular , Estrutura Molecular , Streptococcus mutans/efeitos dos fármacos
15.
J Biol Chem ; 280(50): 41744-52, 2005 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-16223723

RESUMO

Neurotrophin signaling plays important roles in regulating the survival, differentiation, and maintenance of neurons in the nervous system. Binding of neurotrophins to their cognate receptors Trks induces transactivation and phosphorylation of the receptor at several tyrosine residues. These phosphorylated tyrosine residues then serve as crucial docking sites for adaptor proteins containing a Src homology 2 or phosphotyrosine binding domain, which upon association with the receptor initiates multiple signaling events to mediate the action of neurotrophins. Here we report the identification of a Src homology 2 domain-containing molecule, SLAM-associated protein (SAP), as an interacting protein of TrkB in a yeast two-hybrid screen. SAP was initially identified as an adaptor molecule in SLAM family receptor signaling for regulating interferon-gamma secretion. In the current study, we found that SAP interacted with TrkA, TrkB, and TrkC receptors in vitro and in vivo. Binding of SAP required Trk receptor activation and phosphorylation at the tyrosine 674 residue, which is located in the activation loop of the kinase domain. Overexpression of SAP with Trk attenuated tyrosine phosphorylation of the receptors and reduced the binding of SH2B and Shc to TrkB. Moreover, overexpression of SAP in PC12 cells suppressed the nerve growth factor-dependent activation of extracellular signal-regulated kinases 1/2 and phospholipase Cgamma, in addition to inhibiting neurite outgrowth. In summary, our findings demonstrated that SAP may serve as a negative regulator of Trk receptor activation and downstream signaling.


Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Receptor trkA/metabolismo , Sequência de Aminoácidos , Animais , Southern Blotting , Western Blotting , Encéfalo/metabolismo , Linhagem Celular , DNA Complementar/metabolismo , Proteínas Fúngicas/metabolismo , Humanos , Imuno-Histoquímica , Interferon gama/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Camundongos , Microscopia de Fluorescência , Modelos Genéticos , Dados de Sequência Molecular , Mutação , Fatores de Crescimento Neural/metabolismo , Neurônios/metabolismo , Células PC12 , Fosforilação , Ligação Proteica , Estrutura Terciária de Proteína , Ratos , Proteínas Recombinantes de Fusão/química , Transdução de Sinais , Proteína Associada à Molécula de Sinalização da Ativação Linfocitária , Técnicas do Sistema de Duplo-Híbrido , Tirosina/química , Domínios de Homologia de src
16.
Planta Med ; 71(6): 580-4, 2005 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15971136

RESUMO

A silicon-based DNA microarray was designed and fabricated for the identification of toxic traditional Chinese medicinal plants. Species-specific oligonucleotide probes were derived from the 5S ribosomal RNA gene of Aconitum carmichaeli, A. kusnezoffi, Alocasia macrorrhiza, Croton tiglium, Datura inoxia, D. metel, D. tatula, Dysosma pleiantha, Dy. versipellis, Euphorbia kansui, Hyoscyamus niger, Pinellia cordata, P. pedatisecta, P. ternata, Rhododendron molle, Strychnos nux-vomica, Typhonium divaricatum and T. giganteum and the leucine transfer RNA gene of Aconitum pendulum and Stellera chamaejasme. The probes were immobilized via dithiol linkage on a silicon chip. Genomic target sequences were amplified and fluorescently labeled by asymmetric polymerase chain reaction. Multiple toxic plant species were identified by parallel genotyping. Chip-based authentication of medicinal plants may be useful as inexpensive and rapid tool for quality control and safety monitoring of herbal pharmaceuticals and neutraceuticals.


Assuntos
Medicamentos de Ervas Chinesas/química , Fitoterapia , Plantas Medicinais/genética , Sequência de Bases , Primers do DNA , DNA de Plantas/análise , Humanos , Dados de Sequência Molecular , Análise de Sequência com Séries de Oligonucleotídeos , RNA Ribossômico/genética
17.
Planta Med ; 71(4): 349-54, 2005 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15856412

RESUMO

Three new diterpenoids, yuexiandajisu D (1), E (2) and F were isolated from the roots of Euphorbia ebracteolata, along with eight known diterpenoids, jolkinolide B (4), jolkinolide A, ent-11alpha-hydroxyabieta-8(14),13(15)-dien-16,12alpha-olide (6), ent-(13S)-hydroxyatis-16-ene-3,14-dione, ent-3beta,(13S)-dihydroxyatis-16-en-14-one, ent-3-oxokaurane-16alpha,17-diol, ent-16alpha,17-dihydroxyatisan-3-one and ent-atisane-3beta,16alpha,17-triol. The structures of all compounds were deduced using spectroscopic methods and confirmed for 1 and 2 by single-crystal X-ray diffraction. A biogenetic pathway for the formation of 1 and 2 is proposed briefly. Cytotoxic activities were evaluated against ANA-1, B 16 and Jurkat tumor cells. Jolkinolide B (4) displayed modest activity on ANA-1, B 16 and Jurkat tumor cells with IC50 values 4.46 x 10(-2), 4.48 x 10(-2), 6.47 x 10(-2) microM, and ent-11alpha-hydroxyabieta-8(14),13(15)-dien-16,12alpha-olide (6) showed significant activity against ANA-1 and Jurkat cells with IC50 values 7.12 x 10(-3) and 1.79 x 10(-2) microM. Compound 1 was found to be slightly active against ANA-1 cells with an IC50 value 2.88 x 10(-1)microM. Structure-activity relationships of isolated compounds are also discussed.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Euphorbia , Fitoterapia , Extratos Vegetais/farmacologia , Animais , Antineoplásicos Fitogênicos/administração & dosagem , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/uso terapêutico , Linhagem Celular Tumoral , Diterpenos/administração & dosagem , Diterpenos/química , Diterpenos/farmacologia , Diterpenos/uso terapêutico , Humanos , Concentração Inibidora 50 , Camundongos , Extratos Vegetais/administração & dosagem , Extratos Vegetais/química , Extratos Vegetais/uso terapêutico , Raízes de Plantas , Relação Estrutura-Atividade , Difração de Raios X
18.
J Nat Prod ; 67(9): 1548-51, 2004 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-15387657

RESUMO

Three new macrocyclic diterpenes, kansuinins F (1), G (2), and H (3), together with four known jatrophane diterpenes, kansuinins D (4), E (5), and A (6) and 3beta,5alpha,7beta,15beta-tetraacetoxy-9alpha-nicotinoyloxyjatropha-6(17)-11E-dien-14-one, were isolated from the roots of Euphorbia kansui. Compounds 1 and 2 were assigned as 6(17)-en-11,12-epoxy-14-one-type jatrophane diterpenes, and compound 3 as a 6(17)-en-11,14-epoxy-12-one jatrophane diterpene. The structures of compounds 1-3 and the relative configurations of compounds 4 and 5 were determined by spectral data analysis. Kansuinin E (5) exhibited a specific survival effect on fibroblasts that expressed TrkA, a high-affinity receptor for nerve growth factor.


Assuntos
Diterpenos/isolamento & purificação , Euphorbia/química , Plantas Medicinais/química , Diterpenos/química , Diterpenos/farmacologia , Fibroblastos/metabolismo , Medicina Tradicional Chinesa , Estrutura Molecular , Fator de Crescimento Neural/efeitos dos fármacos , Raízes de Plantas/química , Receptor trkA/efeitos dos fármacos
19.
Neurosignals ; 13(5): 248-57, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15305092

RESUMO

With a history of several thousand years, traditional Chinese medicine has been well documented to be effective in the treatment of various disorders. We have investigated the activities of potential neuroactive compounds in traditional Chinese medicine such as Melia toosendan using an in vitro model system, rat pheochromocytoma PC12 cells. We report here that treatment of PC12 cells with a crude extract of the fruits of M. toosendan reduces cell growth in a dose-dependent manner without detectable cytotoxicity. Upon treatment with M. toosendan, PC12 cells exhibit robust neurite outgrowth, to a greater extent than that observed with nerve growth factor. Results obtained with specific kinase inhibitors and protein kinase A-deficient PC12 cells indicate that the actions of M. toosendan are mediated by the activation of protein kinase A and extracellular signal-regulated kinases.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Medicamentos de Ervas Chinesas/farmacologia , Melia/química , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Animais , Western Blotting/métodos , Células COS/efeitos dos fármacos , Diferenciação Celular/fisiologia , Divisão Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Chlorocebus aethiops , Meios de Cultura Livres de Soro/farmacologia , Relação Dose-Resposta a Droga , Interações Medicamentosas , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , L-Lactato Desidrogenase , Fator de Crescimento Neural/farmacologia , Neuritos/efeitos dos fármacos , Células PC12 , Ratos , Receptor trkA/metabolismo , Sais de Tetrazólio , Tiazóis , Fatores de Tempo
20.
Anal Chem ; 74(13): 3168-73, 2002 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-12141679

RESUMO

A novel method for the fast identification of genetic material utilizing a micro-DNA amplification and analysis device (micro-DAAD) consisting of multiple PCR microreactors with integrated DNA microarrays was developed. The device was fabricated in Si-technology and used for the genotyping of Chinese medicinal plants on the basis of differences in the noncoding region of the 5S-rRNA gene. Successful amplification of the genetic material and the consecutive analysis of the fluorescent-labeled amplicons in the micro-DAAD by the integrated oligonucleotide probes were demonstrated. Parallel analysis was performed by loading the four PCR reactors of the micro-DAAD with different samples of 3-microL volume. Temperature sensors and heating elements of the micro-DAAD enable precise temperature control and fast cycling, allowing the rapid completion of a combined amplification and analysis (hybridization) experiment.


Assuntos
Análise de Sequência com Séries de Oligonucleotídeos/instrumentação , Reação em Cadeia da Polimerase Via Transcriptase Reversa/instrumentação , Genótipo , Microcomputadores , Microscopia Confocal , Sondas de Oligonucleotídeos , Espectrometria de Fluorescência
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