RESUMO
NFAT plays a crucial role in the immune system by regulating the transcription of inducible genes during immune responses. In T cells, NFAT proteins govern various cellular events related to T cell development, activation, tolerance induction, and differentiation. We previously reported the NFAT1-dependent enhancer activity of conserved noncoding sequence (CNS)-9, a distal cis-acting element, in the regulation of IL-10 transcription in T cells. In this study, we developed a T cell-based reporter system to identify compounds that modulate the regulatory activity of CNS-9. Among the identified candidates, 6-methoxyflavone (6-MF) significantly inhibited the enhancer activity of CNS-9, thereby reducing IL-10 expression in T cells without affecting cell viability. 6-MF also downregulated the transcription of NFAT1 target genes such as IL-4, IL-13, and IFN-γ. Treatment of 6-MF inhibited the translocation of NFAT1 into the nucleus, which consequently interrupted NFAT1 binding to the target loci, without affecting the expression or dephosphorylation of NFAT1. Treatment of 6-MF to CD4(+) T cells or B cells isolated from mice with atopic dermatitis significantly reduced disease-associated cytokine production, as well as the levels of IgE. In addition, oral administration of 6-MF to atopic dermatitis mice ameliorated disease symptoms by reducing serum IgE levels and infiltrating lymphocytes. Conclusively, our results suggest that 6-MF can be a potential candidate for the development of an effective immunomodulator via the suppression of NFAT-mediated T cell activation.
Assuntos
Transporte Ativo do Núcleo Celular/imunologia , Flavonas/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Fatores de Transcrição NFATC/imunologia , Animais , Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Diferenciação Celular/imunologia , Linhagem Celular Tumoral , Núcleo Celular , Sequência Conservada/efeitos dos fármacos , Sequência Conservada/genética , Citocinas/biossíntese , Proteínas de Ligação a DNA/genética , Dermatite Atópica/tratamento farmacológico , Dermatite Atópica/imunologia , Células HEK293 , Humanos , Imunoglobulina E/sangue , Interferon gama/biossíntese , Interferon gama/genética , Interleucina-10/biossíntese , Interleucina-10/genética , Interleucina-13/biossíntese , Interleucina-13/genética , Interleucina-4/biossíntese , Interleucina-4/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fatores de Transcrição NFATC/antagonistas & inibidores , Fosforilação , Ligação Proteica/efeitos dos fármacos , RNA não Traduzido/efeitos dos fármacos , RNA não Traduzido/genética , Transcrição GênicaRESUMO
Ginger has long been used worldwide as a spice, seasoning, and wine and is also used as a traditional medicine. There have been no previous studies of the potential beneficial effects of the ginger constituent 12-dehydrogingerdione (12-DHGD). We investigated the anti-inflammatory effect of 12-DHGD on lipopolysaccharide (LPS)-stimulated Raw 264.7 cells. The cytotoxicity of 12-DHGD was measured using the MTT assay, and production of prostaglandin E2 (PGE2 ) and the inflammatory cytokines interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α was measured by ELISA. Production of nitric oxide (NO) was measured using Griess reagent and expression of cyclooxygenase-2 (COX-2) and inducible NO (iNOS) enzymes was assessed by reverse transcriptase-polymerase chain reaction. Treatment of Raw 264.7 cells with 12-DHGD significantly inhibited LPS-stimulated production of NO (at 12-DHGD concentrations of 150 and 200 ng/ml), IL-6 (at 50, 100, 150, and 200 ng/ml), and PGE2 (at 200 ng/ml). Consistent with the effects on NO and PGE2 production, 12-DHGD treatment also inhibited the LPS-stimulated increase in iNOS and COX-2 mRNA levels. However, 12-DHGD did not affect production of IL-1ß or TNF-α in response to LPS. 12-DHGD, a constituent of ginger, is a potent inhibitor of proinflammatory mediator production in Raw 264.7 macrophage cells.
Assuntos
Anti-Inflamatórios/farmacologia , Guaiacol/análogos & derivados , Guaiacol/farmacologia , Macrófagos/efeitos dos fármacos , Zingiber officinale/química , Animais , Anti-Inflamatórios/química , Linhagem Celular , Sobrevivência Celular , Ciclo-Oxigenase 2/metabolismo , Dinoprostona/metabolismo , Guaiacol/química , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Lipopolissacarídeos , Macrófagos/enzimologia , Camundongos , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Plants have developed general and specific defense mechanisms for protection against various enemies. Among the general defenses, induced resistance has distinct characteristics, such as broad-spectrum resistance and long-lasting effectiveness. This study evaluated over 500 specific chemical compounds derived from native Korean plant species to determine whether they triggered induced resistance against Pectobacterium carotovorum supsp. carotovorum (Pcc) in tobacco (Nicotiana tabacum) and Pseudomonas syringae pv. tomato (Pst) in Arabidopsis thaliana. To select target compound(s) with direct and indirect (volatile) effects, a new Petri-dish-based in vitro disease assay system with four compartments was developed. The screening assay showed that capsaicin, fisetin hydrate, jaceosidin, and farnesiferol A reduced the disease severity significantly in tobacco. Of these four compounds, capsaicin and jaceosidin induced resistance against Pcc and Pst, which depended on both salicylic acid (SA) and jasmonic acid (JA) signaling, using Arabidopsis transgenic and mutant lines, including npr1 and NahG for SA signaling and jar1 for JA signaling. The upregulation of the PR2 and PDF1.2 genes after Pst challenge with capsaicin pre-treatment indicated that SA and JA signaling were primed. These results demonstrate that capsaicin and jaceosidin can be effective triggers of strong induced resistance against both necrotrophic and biotrophic plant pathogens.
Assuntos
Arabidopsis/microbiologia , Resistência à Doença/efeitos dos fármacos , Nicotiana/microbiologia , Pectobacterium carotovorum/fisiologia , Extratos Vegetais/farmacologia , Pseudomonas syringae/fisiologia , Arabidopsis/genética , Arabidopsis/metabolismo , Capsaicina/farmacologia , Ciclopentanos/metabolismo , Flavonoides/farmacologia , Flavonóis , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genes de Plantas , Interações Hospedeiro-Patógeno , Oxilipinas/metabolismo , Doenças das Plantas/microbiologia , Reguladores de Crescimento de Plantas/metabolismo , República da Coreia , Ácido Salicílico/metabolismo , Sesquiterpenos/farmacologia , Transdução de Sinais , Nicotiana/genética , Nicotiana/metabolismoRESUMO
Decursin (De), an active component of Angelica gigas, is known to exert anticancer and neuroprotective effects. However, its antiobesity and antidiabetic potential has not yet been investigated. This study evaluated the antiobesity effect of decursin, particularly focusing on its ability to inhibit adipocyte differentiation in 3T3-L1 cells. Decursin treatment resulted in the inhibition of adipocyte differentiation and the expression of fatty acid synthase. The study further investigated these antiobesity effects using mice fed a normal diet (ND), a high-fat diet (HFD) and a HFD plus decursin 200 mg/kg diet (HFD + De) for 7 weeks. Mice administered HFD plus decursin showed a drastic decrease in weight gain, triglyceride content, total cholesterol content and fat size compared with those that received the HFD alone; this was observed despite similar quantities of total food intake. Furthermore, decursin improved glucose tolerance in mice fed a HFD. Finally, administration of decursin along with the HFD significantly reduced the secretion of HFD-induced adipocytokines such as leptin, resistin, IL-6 and MCP-1. These results suggest that decursin might be useful for the treatment of obesity and diabetes.
Assuntos
Adipocinas/sangue , Tecido Adiposo/metabolismo , Angelica/química , Fármacos Antiobesidade/farmacologia , Benzopiranos/farmacologia , Butiratos/farmacologia , Hipoglicemiantes/farmacologia , Células 3T3-L1 , Adipocinas/antagonistas & inibidores , Adipocinas/metabolismo , Animais , Fármacos Antiobesidade/química , Fármacos Antiobesidade/isolamento & purificação , Benzopiranos/química , Benzopiranos/isolamento & purificação , Glicemia/metabolismo , Peso Corporal/efeitos dos fármacos , Butiratos/química , Butiratos/isolamento & purificação , Colesterol/análise , Dieta Hiperlipídica/efeitos adversos , Teste de Tolerância a Glucose , Hipoglicemiantes/química , Hipoglicemiantes/isolamento & purificação , Resistência à Insulina , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/induzido quimicamente , Obesidade/fisiopatologia , Aumento de Peso/efeitos dos fármacosRESUMO
Farnesylation of the activated RAS oncogene product by protein farnesyltransferase (FTase) is a critical step for its oncogenic function. Bioassay-guided purification of Ferula asafoetida (Umbelliferae) extract led to the isolation of the coumarin-derived sesquiterpene galbanic acid (1) as an active principal for FTase inhibitory activity, together with the four structurally related sesquiterpenes karatavicinol (2), umbelliprenin (3), farnesiferol B (4), and farnesiferol C (5). The 50 % inhibitory concentration (IC (50)) of 1 against FTase in an enzyme-based assay was calculated as 2.5 µM. Compound 1 also demonstrated potent inhibition of the proliferation of oncogenic RAS-transformed NIH3T3/Hras-F in a dose-dependent manner. The IC (50) value of 1 on the proliferation of oncogenic RAS-transformed NIH3T3/Hras-F cells was calculated as 16.2 µM, whereas its IC (50) value on control vector-transfected normal RAS-containing NIH3T3/ZIPneo cells was 58.5 µM.
Assuntos
Cumarínicos/farmacologia , Citostáticos/farmacologia , Farnesiltranstransferase/antagonistas & inibidores , Ferula/química , Animais , Encéfalo/enzimologia , Cumarínicos/química , Cumarínicos/isolamento & purificação , Citostáticos/química , Citostáticos/isolamento & purificação , Farnesiltranstransferase/isolamento & purificação , Concentração Inibidora 50 , Camundongos , Células NIH 3T3 , Prenilação/efeitos dos fármacos , RatosRESUMO
Three new guaiane-type sesquiterpene lactones (1-3), together with nine related sesquiterpenes (4-12), were isolated from the whole extract of Ixeris dentata (Asteraceae). The chemical structures of isolates 1-12 were established by spectroscopic analyses as 3 ß,8 ß-dihydroxy-guaia-10(14)-en-1 α,4 α,5 α,6 ß,7 α,11 ßH-12,6 α-olide (1), ixerin N 6'- O-acetate (2), ixerisoside A 6'- O-acetate (3), ixerin N ( 4), ixerisoside A (5), ixerin M (6), tectroside (7), 8-epidesacylcynaropicrin glucoside (8), 8-epiisolipidiol (9), 11 ßH-11,13-dihydrointegrifolin (10) 8 ß-hydroxy-4 ß,15-dihydrozaluzanin C (11), and integrifolin (12). Compounds 1-12 were evaluated for their inhibitory effect on the proliferation of the cultured human tumor cell lines MES-SA, MES-SA/DX5, HCT-15, and HCT15/CL02 in vitro.
Assuntos
Antineoplásicos Fitogênicos/isolamento & purificação , Asteraceae/química , Lactonas/isolamento & purificação , Extratos Vegetais/química , Sesquiterpenos/isolamento & purificação , Antineoplásicos Fitogênicos/farmacologia , Antineoplásicos Fitogênicos/uso terapêutico , Linhagem Celular Tumoral , Humanos , Lactonas/química , Lactonas/farmacologia , Estrutura Molecular , Neoplasias/tratamento farmacológico , Fitoterapia , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Sesquiterpenos/química , Sesquiterpenos/farmacologiaRESUMO
A new resveratrol oligomer (1) together with eight related components (2- 9) were isolated from the seed extract of Paeonia lactiflora (Paeoniaceae) as active principles responsible for the inhibition of beta-site APP-cleaving enzyme 1 (BACE-1) in vitro. The chemical structure of 1 was established as (-)-7a,8a- CIS- ε-viniferin with the aid of spectroscopic analyses including NOESY experiments. All isolated resveratrol oligomers (1- 9) demonstrated significant inhibition on baculovirus-expressed BACE-1 in a dose-dependent manner, which was assessed by the FRET assay using Rh-EVNLDAEFK as a substrate in vitro.
Assuntos
Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Ácido Aspártico Endopeptidases/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Paeonia/química , Extratos Vegetais/farmacologia , Estilbenos/farmacologia , Baculoviridae , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/química , Inibidores Enzimáticos/isolamento & purificação , Estrutura Molecular , Extratos Vegetais/química , Resveratrol , Sementes , Estilbenos/química , Estilbenos/isolamento & purificaçãoRESUMO
AIMS: This study was carried out to evaluate the hepatoprotective effect of Platycodon grandiflorum (PG) in ethanol (EtOH)-induced liver damage. METHODS AND RESULTS: PG treatment (both the total extract and saponin fraction) significantly blocked EtOH-induced oxidative stress through the preservation of activities of antioxidant enzymes in HepG2 cells. Furthermore, while the administration of EtOH to C57BL/6 mice for 6 weeks induced liver damage, along with a significant increase in plasma glutamic oxalacetic transaminase, glutamic pyruvic transaminase, hepatic triglyceride and thiobarbituric acid reactive substance levels, PG treatment significantly decreased glutamic oxalacetic transaminase, glutamic pyruvic transaminase, hepatic triglyceride and thiobarbituric acid reactive substance levels compared with the EtOH-treated control group (p < 0.05). Histological observation by hematoxylin-eosin and oil red O staining in the liver showed more effective inhibition of lipid accumulation in PG-treated groups, as compared to the EtOH-treated control group. Additionally, PG treatments appeared to enhance the activities of superoxide dismutase and catalase in the liver (p < 0.05). CONCLUSION: These results suggest that PG has a protective effect against EtOH-induced oxidative damage, possibly by inhibition of lipid accumulation and peroxidation through the enhancement of the antioxidant defense system. PG might be useful as a therapeutically potent natural ingredient for the prevention of chronic EtOH-induced oxidative stress and liver damage.
Assuntos
Doença Hepática Crônica Induzida por Substâncias e Drogas/tratamento farmacológico , Etanol/toxicidade , Fígado/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Platycodon/química , Alanina Transaminase/sangue , Animais , Antioxidantes/farmacologia , Doença Hepática Crônica Induzida por Substâncias e Drogas/patologia , Células Hep G2 , Humanos , Peroxidação de Lipídeos/efeitos dos fármacos , Fígado/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Substâncias Reativas com Ácido Tiobarbitúrico/análise , Triglicerídeos/sangueRESUMO
Here, we show the involvement of signaling pathways to induce the gene expression of bone morphogenetic protein (BMP) in the osteogenic activity of physcion-8-O-beta-D-glucopyranoside (physcion-Glu); it stimulated osteoblast differentiation in mouse osteoblast MC3T3-E1 subclone 4 cells and induced BMP-2 gene expression and activation of Akt and ERK/MAP kinases. Physcion-Glu-induced BMP-2 expression and mineralization were attenuated by LY294002, an inhibitor of PI3K that lies upstream of Akt and MAP kinases, suggesting that physcion-Glu induces osteoblast differentiation via PI3K-Akt/MAP kinase signaling pathways, which play important roles in inducing BMP-2 gene expression. Physcion-Glu also enhanced BMP-2-induced commitment of mouse bi-potential mesenchymal precursor C2C12 cells into osteoblasts while inducing the transcription of several osteogenic BMP isoforms, such as BMP-2, -4, -7, and -9. Osteogenic synergy between BMP-2 and physcion-Glu was supported by the fact that noggin inhibited BMP-2 and physcion-Glu-induced alkaline phosphatase expression and activity. Considering that physcion-Glu induced Runx2 activity and the nuclear translocation of p-Smad, physcion-Glu could act by enhancing the BMP signaling pathway that induces Smad activation and translocation to activate Runx2. In conclusion, physcion-Glu could enhance the commitment of mesenchymal progenitors into osteoblasts and their differentiation by activating signaling pathways to induce BMP gene expression.
Assuntos
Proteína Morfogenética Óssea 2/metabolismo , Emodina/análogos & derivados , Glucosídeos/química , Glucosídeos/farmacologia , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Osteoblastos/citologia , Transdução de Sinais/efeitos dos fármacos , Animais , Western Blotting , Proteína Morfogenética Óssea 2/genética , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Cromonas/farmacologia , Emodina/química , Camundongos , Morfolinas/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Reação em Cadeia da Polimerase , Regiões Promotoras Genéticas/genéticaRESUMO
Three new triterpenoid saponins, platyconic acid B lactone (1), deapio-platyconic acid B lactone (2), and deapio-platycodin D(2) (3), together with 17 known triterpenoid saponins, were isolated from a root extract of Platycodon grandiflorum. The structures of 1-3 were determined on the basis of spectroscopic data interpretation and chemical transformation. Saponins with a platycodigenin or polygalacic acid unit as a sapogenin demonstrated significant inhibitory effects on the proliferation of a small panel of cultured human tumor cells.
Assuntos
Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Platycodon/química , Saponinas/isolamento & purificação , Saponinas/farmacologia , Triterpenos/isolamento & purificação , Triterpenos/farmacologia , Antineoplásicos/química , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Estrutura Molecular , Raízes de Plantas/química , Saponinas/química , Triterpenos/química , Células Tumorais CultivadasRESUMO
Soil metagenome constitutes a reservoir for discovering novel enzymes from the unculturable microbial diversity. From three plant rhizosphere metagenomic libraries comprising a total of 142,900 members of recombinant plasmids, we obtained 14 recombinant fosmids that exhibited lipolytic activity. A selected recombinant plasmid, pFLP-2, which showed maximum lipolytic activity, was further analyzed. DNA sequence analysis of the subclone in pUC119, pELP-2, revealed an open reading frame of 1,191 bp encoding a 397-amino-acid protein. Purified EstD2 exhibited maximum enzymatic activity towards p-nitrophenyl butyrate, indicating that it is an esterase. Purified EstD2 showed optimal activity at 35 °C and at pH 8.0. The K(m) and K(cat) values were determined to be 79.4 µM and 120.5/s, respectively. The esterase exhibited an increase in enzymatic activity in the presence of 15% butanol and 15% methanol. Phylogenetic analysis revealed that the lipolytic protein EstD2 may be a member of a novel family of lipolytic enzymes. Several hypothetical protein homologs of EstD2 were found in the database. A hypothetical protein from Phenylobacterium zucineum HLK1, a close homolog of EstD2, displayed lipolytic activity when the corresponding gene was expressed in Escherichia coli. Our results suggest that the other hypothetical protein homologs of EstD2 might also be members of this novel family.
Assuntos
Butiratos/metabolismo , Esterases/isolamento & purificação , Esterases/metabolismo , Metagenoma , Rizosfera , Microbiologia do Solo , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biota , Caulobacteraceae/genética , Clonagem Molecular , DNA Bacteriano/genética , Escherichia coli/enzimologia , Escherichia coli/genética , Escherichia coli/metabolismo , Esterases/química , Expressão Gênica , Genoma Bacteriano , Biblioteca Genômica , Dados de Sequência Molecular , Filogenia , Plasmídeos/genética , República da Coreia , Alinhamento de Sequência , Análise de Sequência de DNA , Especificidade por SubstratoRESUMO
Platycodon grandiflorum (PG) (Korean name, Doraji; Chinese name, Jiegeng; and Japanese name, Kikyo) is a perennial plant in the Campanulaceae family that contains triterpenoid saponins, carbohydrates, and fibers. This study was carried out to investigate effects of root of PG on fatty liver inhibition in high fat diet (HFD)-fed C57BL/6 mice. C57BL/6 mice were divided into control, total extract of PG (T-PG, 500 mg/kg) and saponin fraction (S-PG, 50 mg/kg)-treated groups. Significant decreases in body weight, associated with fat mass reduction, were observed in PG-treated groups (p<0.05). Hepatic lipid content and score index calculated from morphometric observations on fatty liver were significantly decreased in the PG-treated groups (p<0.05). Moreover, activities of fatty acid synthase (FAS) and carnitine palmitoyl-transferase (CPT) were significantly suppressed and increased as compared with the control group, respectively (p<0.05). mRNA expressions of the sterol regulatory element binding protein (SREBP1c) and stearoyl-CoA desaturase (SCD1) gene were suppressed in the T-PG and S-PG groups (p<0.05). From these findings, we speculate that fatty liver inhibition effects of PG extract and its saponins appear to be conferred by hepatic lipogenesis and acceleration of energy expenditure, along with modulation of liver FAS and CPT activities in HFD-fed C57BL/6 mice.
Assuntos
Tecido Adiposo/efeitos dos fármacos , Fígado Gorduroso/prevenção & controle , Lipogênese/efeitos dos fármacos , Fígado/metabolismo , Extratos Vegetais/uso terapêutico , Platycodon/química , Saponinas/uso terapêutico , Animais , Peso Corporal/efeitos dos fármacos , Carnitina O-Palmitoiltransferase/metabolismo , Dieta Aterogênica , Gorduras na Dieta/administração & dosagem , Metabolismo Energético/efeitos dos fármacos , Ácido Graxo Sintases/metabolismo , Fígado Gorduroso/induzido quimicamente , Fígado Gorduroso/metabolismo , Regulação da Expressão Gênica , Lipogênese/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fitoterapia , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Raízes de Plantas , RNA Mensageiro/metabolismo , Saponinas/farmacologia , Estearoil-CoA Dessaturase/genética , Estearoil-CoA Dessaturase/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismoRESUMO
Natural compounds with bone-forming (or anabolic) activity have been recently focused on in bone research. The present study investigated the effect of undecylenic acid (UA) on osteoblast differentiation in mouse osteoblastic MC3T3-E1 subclone 4 cells and primary mouse calvarial cells. Low concentrations of UA (up to 5 microM) exhibited no cytotoxicity and significantly increased the expression and activity of alkaline phosphatase (early differentiation marker of osteoblast) and calcium deposition with the induction of expression of the osteocalcin gene in both cells. Interestingly, at low concentration of UA, the induction of NF-kappaB p65 translocation into nucleus and the up-regulation of AP-1 and NFATc1 transcript levels were also observed, suggesting that the stimulatory effect of UA on osteoblast differentiation could be mediated through the activation of transcription factors. Additionally, although the patterns of UA-induced activation of MAP kinases (JNK and p38) were not completely consistent with the increase of both ALP activity and calcium deposition by UA, MAP kinases might be partially involved in the biological function of UA during the early and late stages of osteoblast differentiation.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Osteoblastos/efeitos dos fármacos , Ácidos Undecilênicos/farmacologia , Animais , Linhagem Celular , Camundongos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Osteoblastos/citologia , Osteoblastos/enzimologia , Fatores de Transcrição/metabolismoRESUMO
The time-resolved fluorescence technique based on melanin-concentrating hormone (MCH) receptor subtype-1 (MCH-1 receptor) binding assay was adopted to carry out a bioassay-guided fractionation of the methanol extract of Morus alba leaves. This fractionation and purification led to the isolation of two compounds identified as pheophorbide a methyl ester and 13(2)(S)-hydroxypheophorbide a methyl ester. These active pheophorbides exhibited potent inhibitory activity in binding of europium-labeled MCH to the human recombinant MCH-1 receptor (IC(50) value; 4.03 and 0.33 microM, respectively). Besides binding activity, the pheophorbides inhibited MCH-mediated extracellular signal-regulated kinase (ERK) phosphorylation in Chinese hamster ovary cells expressing human MCH-1 receptor. These results suggest that pheophorbide a methyl ester and 13(2)(S)-hydroxypheophorbide a methyl ester act as modulators of MCH-1 receptor and MCH-mediated ERK signaling.
Assuntos
Clorofila/análogos & derivados , Morus/química , Extratos Vegetais/farmacologia , Receptores do Hormônio Hipofisário/metabolismo , Animais , Células CHO , Clorofila/farmacologia , Cricetinae , Cricetulus , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Estrutura Molecular , Fosforilação , Folhas de Planta/química , Proteínas Recombinantes/metabolismo , Transdução de SinaisRESUMO
Exendin-4, a long-acting glucagon-like peptide-1-receptor agonist, is known to enhance beta-cell function, but the active mechanism by which it modulates beta-cell mass still remains unclear. We investigated what the long-term effects of exendin-4 (300 pmol/kg body weight per day) on beta-cell function and mass would be in 90% pancreatectomized (Px) Sprague Dawley rats; half of whom were intraperitoneally injected with streptozotocin (STZ, 20 mg/kg body weight) and half of whom were not. Exendin-4 improved glucose tolerance by elevating serum insulin levels in both STZ-treated and untreated Px rats. At hyperglycemic clamp, STZ attenuated both first and second phase insulin secretion in STZ- and saline-treated Px rats, but exendin-4 incompletely reversed the attenuation. Since STZ mostly removed the remaining beta-cells by increasing apoptosis after Px, their regeneration was initiated through neogenesis, which was determined by the number of beta-cells budding from pancreatic duct layers and small clusters. Exendin-4 enhanced beta-cell proliferation and neogenesis in STZ-treated and -untreated Px rats and reduced beta-cell apoptosis partly by attenuating the expression of endoplasmic reticulum stress-response genes such as X-box-binding protein-1, activating transcription factor (ATF)-4, ATF6, and C/EBP-homologous protein. In conclusion, exendin-4 improved glycemic control by potentiating beta-cell function and increasing beta-cell mass by increasing beta-cell neogenesis and proliferation and by decreasing apoptosis in diabetic rats.
Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Retículo Endoplasmático/efeitos dos fármacos , Células Secretoras de Insulina/efeitos dos fármacos , Insulina/metabolismo , Ilhotas Pancreáticas/efeitos dos fármacos , Peptídeos/farmacologia , Estresse Fisiológico/efeitos dos fármacos , Peçonhas/farmacologia , Animais , Apoptose/efeitos dos fármacos , Glicemia/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Diabetes Mellitus Experimental/metabolismo , Retículo Endoplasmático/metabolismo , Ingestão de Energia/efeitos dos fármacos , Exenatida , Hipoglicemiantes/farmacologia , Hipoglicemiantes/uso terapêutico , Insulina/sangue , Secreção de Insulina , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/metabolismo , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/metabolismo , Masculino , Pancreatectomia , Peptídeos/uso terapêutico , Ratos , Ratos Sprague-Dawley , Fatores de Transcrição/metabolismo , Peçonhas/uso terapêuticoRESUMO
A new resveratrol dimer, (+)-vitisinol E (1) which demonstrated inhibitory activity on BACE-1 (beta-site APP-cleaving enzyme 1) in vitro, was isolated from the stembark extract of Vitis vinifera (Vitaceae) together with four known resveratrol oligomers, (+)-epsilon-viniferin (2), (+)-ampelopsin A (3), (+)-vitisin A (4) and (-)-vitisin B (5). The chemical structure of 1 was established by MR spectroscopic analyses, including HMBC. All isolated resveratrol derivatives (1-5) demonstrated significant inhibition on baculovirus-expressed BACE-1 in a dose-dependent manner, which was assessed with the FRET assay using Rh-EVNLDAEFK as a substrate in vitro.
Assuntos
Antioxidantes/farmacologia , Inibidores Enzimáticos/farmacologia , Extratos Vegetais/farmacologia , Estilbenos/farmacologia , Vitis/química , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Antioxidantes/isolamento & purificação , Ácido Aspártico Endopeptidases/antagonistas & inibidores , Baculoviridae , Inibidores Enzimáticos/isolamento & purificação , Estrutura Molecular , Casca de Planta , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Caules de Planta , Resveratrol , Estilbenos/isolamento & purificaçãoRESUMO
In this study, the effect of (-)-saucerneol, one of the lignans isolated from Saururus chinensis, on osteoclast differentiation and bone resorption was evaluated in two in vitro models for osteoclast differentiation, the receptor activator of nuclear factor kappaB (NF-kappaB) ligand (RANKL)-treated RAW264.7 cells and mouse BMMs treated with both RANKL and macrophage-colony stimulating factor. (-)-Saucerneol significantly inhibited the RANKL-induced activity of tartrate-resistance acid phosphatase (TRAP, an early marker of osteoclast formation) and formation of osteoclasts in a dose-dependent manner. Interestingly, (-)-saucerneol was shown to inhibit the RANKL-induced activation of extracellular signal-regulated kinase in both in vitro models. In addition, (-)-saucerneol inhibited the bone resorptive activity and the expression of transcription factors and genes essential for osteoclast formation and bone resorption as well. In conclusion, (-)-saucerneol has a potential to inhibit the osteoclast differentiation via preventing the activation of ERK signaling pathway. In addition, its activity to inhibit the bone resorption activities of osteoclasts could result from its potential to inhibit RANKL-induced expression levels of transcription factors and genes essential for bone resorption.
Assuntos
Reabsorção Óssea/metabolismo , Furanos/farmacologia , Lignanas/farmacologia , Osteoclastos/efeitos dos fármacos , Fosfatase Ácida/antagonistas & inibidores , Animais , Linhagem Celular , Furanos/isolamento & purificação , Expressão Gênica/efeitos dos fármacos , Lignanas/isolamento & purificação , Fator Estimulador de Colônias de Macrófagos/farmacologia , Camundongos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Ligante RANK/farmacologia , Fatores de Transcrição/metabolismoRESUMO
Thee present study analysed the quantification of rutin in raw buckwheat extract (RBE) and germinated buckwheat extract (GBE) by high performance liquid chromatography (HPLC), and examined changes in body weight, systolic blood pressure (SBP) and nitrotyrosine (a marker for peroxynitrite formation) immunoreactivity in aortic endothelial cells in spontaneously hypertensive rats (SHRs) and normotensive Wistar-Kyoto (WKY) rats after treatment with RBE and GBE for 5 weeks. In the HPLC study, RBE and GBE contained a mean content of rutin of 1.52 +/- 0.21 and 2.92 +/- 0.88 mg/g, respectively. In the 600 mg/kg GBE-treated group, SBP was lower than that in the 600 mg/kg RBE-treated group. The treatment with RBE and/or GBE significantly reduced oxidative damage in aortic endothelial cells by lowering nitrotyrosine immunoreactivity. These results suggest that GBE has an antihypertensive effect and may protect arterial endothelial cells from oxidative stress.
Assuntos
Anti-Hipertensivos/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Extratos Vegetais/farmacologia , Rutina/farmacologia , Tirosina/análogos & derivados , Animais , Aorta/citologia , Peso Corporal , Cromatografia Líquida de Alta Pressão , Células Endoteliais/metabolismo , Fagopyrum/química , Estrutura Molecular , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Tirosina/metabolismoRESUMO
The identification of anabolic agents that directly stimulate bone formation has recently attracted greater interest. Here, baicalein was identified as a natural compound that stimulates the differentiation of mouse osteoblastic MC3T3-E1 subclone 4 cells. Baicalein induced the activation of NF-kappaB in the initiation stage of osteoblast differentiation, and it activated the MAP kinase/NF-kappaB signaling pathway and induced the expression of osteoblast differentiation markers in the early stage. In the late stage, baicalein stimulated the calcium deposition with the activation of MAP kinases and AP-1 family members such as Fra-1 and Fra-2. Another transcription factor, NFATc1, was slightly induced by baicalein in the late stage. Thus, baicalein could stimulate the osteoblast differentiation via the activation of complexly coordinated signaling pathways that include MAP kinases and transcription factors such as NF-kappaB, AP-1, and NFATc1.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Flavanonas/farmacologia , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Osteoblastos/citologia , Osteoblastos/enzimologia , Fatores de Transcrição/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Biomarcadores/metabolismo , Calcificação Fisiológica/efeitos dos fármacos , Diferenciação Celular/genética , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Flavanonas/química , Regulação da Expressão Gênica/efeitos dos fármacos , Camundongos , NF-kappa B/metabolismo , Fatores de Transcrição NFATC/genética , Fatores de Transcrição NFATC/metabolismo , Osteoblastos/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fator de Transcrição AP-1/genética , Fator de Transcrição AP-1/metabolismoRESUMO
Inhibition of adipocyte differentiation is one approach among the anti-obesity strategies. This study demonstrates that vitisin A, a resveratrol tetramer, inhibits adipocyte differentiation most effectively of 18 stilbenes tested. Fat accumulation and PPARgamma expression were decreased by vitisin A in a dose-dependent manner. Vitisin A significantly inhibited preadipocyte proliferation and consequent differentiation within the first 2 days of treatment, indicating that the anti-adipogenic effect of vitisin A was derived from anti-proliferation. Based on cell cycle analysis, vitisin A blocked the cell cycle at the G1-S phase transition, causing cells to remain in the preadipocyte state. Vitisin A increased p21 expression, while the Rb phosphorylation level was reduced. Therefore, vitisin A seems to induce G1 arrest through p21- and consequent Rb-dependent suppression of transcription. On the other hand, ERK and Akt signaling pathways were not involved in the anti-mitotic regulation by vitisin A. Taken together, these results suggest that vitisin A inhibits adipocyte differentiation through preadipocyte cell cycle arrest.