RESUMO
In this study, the effects of 3,5,7,3',4'-pentamethoxyflavone (KP1), a major bioactive ingredient isolated from the Kaempferia parviflora rhizomes, on a neurite outgrowth in Neuro2a cells and its mechanism have been investigated. KP1 increased concentration-dependently the percentage of neurite-bearing cells. KP1 showed a remarkable capability to elicit neurite outgrowth in Neuro2a cells, as evidenced by morphological alterations and immunostaining using anti-class III ß-tubulin and anti-NeuN antibodies. KP1 also displayed a higher neurogenic activity than retinoic acid (RA), a promoter of neurite outgrowth in Neuro2a cells. KP1 treatment caused significant elevation in phosphorylation of extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (p38 MAPK) and glycogen synthase kinase-3ß (GSK-3ß). However, KP1-triggered neurite outgrowth was markedly inhibited by treatment with the ERK inhibitor U0126, whereas p38 MAPK inhibitor SB203580 and GSK-3ß inhibitor SB216763 did not influence KP1-induced neurite outgrowth. These results demonstrate that KP1 elicits neurite outgrowth and triggers cell differentiation of Neuro2a cells through ERK signal pathway.
Assuntos
Sistema de Sinalização das MAP Quinases , Crescimento Neuronal , Animais , Crescimento Neuronal/efeitos dos fármacos , Camundongos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Neuritos/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Flavonoides/farmacologia , Flavonas/farmacologia , Flavonas/química , Linhagem Celular Tumoral , Glicogênio Sintase Quinase 3 beta/metabolismo , Linhagem CelularRESUMO
In this study, we found that gene expression of the human ß-galactoside α2,6-sialyltransferase (hST6Gal I) was specifically increased during differentiation of human MG-63 osteoblastic cells by serum starvation (SS). In parallel, a distinct increase in binding to SNA, the α2,6-sialyl-specific lectin, was observed in serum-starved cells, as demonstrated by FACS analysis. 5'-Rapid amplification of cDNA ends analysis demonstrated that the increase of hST6Gal I transcript by SS is mediated by P1 promoter. To elucidate transcriptional regulation of hST6Gal I in SS-induced MG-63 cells, we functionally characterized the P1 promoter region of the hST6Gal I gene. The 5'-deletion analysis of P1 promoter region revealed that the 189 bp upstream region of transcription start site is critical for transcriptional activity of hST6Gal I gene in SS-induced MG-63 cells. This region contains the predicted binding sites for several transcription factors, including AREB6, FOXP1, SIX3, HNF1, YY2, and MOK2. The mutagenesis analysis for these sites and chromatin immunoprecipitation assay demonstrated that the YY2 binding site at -98 to -77 was essential for the SS-induced hST6Gal I gene expression during differentiation of MG-63 cells.
Assuntos
Antígenos CD/genética , Diferenciação Celular/genética , Osteoblastos/citologia , Sialiltransferases/genética , Transcrição Gênica , Proteínas de Ligação a DNA/genética , Proteínas do Olho/genética , Fatores de Transcrição Forkhead/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Regulação Enzimológica da Expressão Gênica/genética , Proteínas de Homeodomínio/genética , Humanos , Proteínas do Tecido Nervoso/genética , Osteoblastos/metabolismo , Regiões Promotoras Genéticas/genética , Proteínas Repressoras/genética , Fatores de Transcrição/genética , Sítio de Iniciação de Transcrição , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética , Proteína Homeobox SIX3RESUMO
Our recent report showed that curcumin, polyphenolic compound isolated from the herb Curcuma longa, upregulated the gene expression of human GD3 synthase (hST8Sia I) responsible for ganglioside GD3 synthesis with autophagy induction in human lung adenocarcinoma A549 cells. In this study, on the contrary to this finding, we demonstrated that curcumin downregulated the gene expression of human GM3 synthase (hST3Gal V) catalyzing ganglioside GM3 synthesis with autophagy induction in human colon carcinoma HCT116 cells. To clarify the mechanism leading to the downregulation of hST3Gal V gene expression in curcumin-treated HCT116 cells, we analyzed the curcumin-inducible promoter of the hST3Gal V gene by luciferase reporter assays. Promoter deletion analysis demonstrated that the -177 to -83 region, which includes putative binding sites for transcription factors NFY, CREB/ATF, SP1, EGR3, and MZF1, acts as the curcumin-responsive promoter of the hST3Gal V gene. Site-directed mutagenesis and chromatin immunoprecipitation analysis demonstrated that the CREB/ATF binding site at -143 is pivotal for curcumin-induced downregulation of hST3Gal V gene in HCT116 cells. The transcriptional activation of hST3Gal V in HCT116 cells was significantly repressed by an inhibitor of AMP-activated protein kinase (AMPK). These results suggest that AMPK signal pathway mediates hST3Gal V gene expression in HCT116 cells.
RESUMO
In this study, a neurite outgrowth-inducing substance was isolated from the ethylacetate extract of the Polygonum multiflorum roots and identified as emodin by gas-liquid chromatography-mass spectrometry and (1)H NMR and (13)C NMR. Emodin displayed remarkable neurite outgrowth-inducing activity in Neuro2a cells, as demonstrated by morphological changes and immunocytochemistry for class III ß-tubulin. Emodin exhibited a stronger neutrophic activity than retinoic acid (RA) known as inducer of neurite outgrowth in Neuro2a cells. Emodin treatment resulted in marked increases in phosphorylation of Akt a direct downstream signaling molecule of phosphatidylinositol 3-kinase (PI3K), but upstream of glycogen synthase kinase-3ß (GSK-3ß) and cAMP response element-binding protein (CREB). These augmentations and neurite-bearing cells induced by emodin were remarkably reduced by the addition of PI3K inhibitor LY294002. These results demonstrate that emodin induces neuronal differentiation of Neuro2a cells via PI3K/Akt/GSK-3ß pathway.
Assuntos
Emodina/farmacologia , Quinase 3 da Glicogênio Sintase/metabolismo , Neuritos/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Emodina/isolamento & purificação , Glicogênio Sintase Quinase 3 beta , Camundongos , Neuritos/fisiologia , Extratos Vegetais/química , Polygonum/química , Transdução de SinaisRESUMO
The biochemical mechanisms of cell death by oleifolioside B (OB), a cycloartane-type triterpene glycoside isolated from Dendropanax morbifera Leveille, were investigated in A549 human lung carcinoma cells. Our data indicated that exposure to OB led to caspase activation and typical features of apoptosis; however, apoptotic cell death was not prevented by z-VAD-fmk, a pan-caspase inhibitor, demonstrating that OB-induced apoptosis was independent of caspase activation. Subsequently, we found that OB increased autophagy, as indicated by an increase in monodansylcadaverine fluorescent dye-labeled autophagosome formation and in the levels of the autophagic form of microtubule-associated protein 1 light chain 3 and Atg3, an autophagy-specific gene, which is associated with inhibiting phospho-nuclear factor erythroid 2-related factor 2 (Nrf2) expression. However, pretreatment with bafilomycin A1, an autophagy inhibitor, attenuated OB-induced apoptosis and dephosphorylation of Nrf2. The data suggest that OB-induced autophagy functions as a death mechanism in A549 cells and OB has potential as a novel anticancer agent capable of targeting apoptotic and autophagic cell death and the Nrf2 signaling pathway.
Assuntos
Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Saponinas/farmacologia , Clorometilcetonas de Aminoácidos/farmacologia , Antineoplásicos/farmacologia , Proteínas Relacionadas à Autofagia , Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/biossíntese , Caspase 3/metabolismo , Caspase 8/metabolismo , Caspase 9/metabolismo , Inibidores de Caspase/farmacologia , Linhagem Celular Tumoral , Inibidores Enzimáticos/farmacologia , Humanos , Proteínas Inibidoras de Apoptose/biossíntese , Macrolídeos/farmacologia , Proteínas Associadas aos Microtúbulos/biossíntese , Proteínas Associadas aos Microtúbulos/metabolismo , Fator 2 Relacionado a NF-E2/biossíntese , Fator 2 Relacionado a NF-E2/metabolismo , Extratos Vegetais/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Survivina , Enzimas de Conjugação de Ubiquitina/biossíntese , Enzimas de Conjugação de Ubiquitina/metabolismoRESUMO
Triptolide, a diterpene derived from Tripterygium wilfordii Hook f., a Chinese medicinal herb, has been reported to inhibit cell proliferation and induce apoptosis in various human cancer cells, but its anticancer effects on human osteosarcoma cells have not yet been elucidated. In this study, we investigated whether triptolide induces apoptosis in human osteosarcoma cells and the underlying molecular mechanisms. We firstly demonstrated that triptolide inhibited cell growth and induced apoptosis in U2OS cells. Western blot analysis showed that the levels of procaspase-8, -9, Bcl-2, Bid and mitochondrial cytochrome c were downregulated in triptolide-treated U2OS cells, whereas the levels of Fas, FasL, Bax, cytosolic cytochrome c, cleaved caspase-3 and cleaved PARP were upregulated. These results suggest that triptolide induces apoptosis in U2OS cells by activating both death receptor and mitochondrial apoptotic pathways.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Diterpenos/farmacologia , Osteossarcoma/patologia , Fenantrenos/farmacologia , Caspase 3/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Compostos de Epóxi/farmacologia , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Pontos de Checagem da Fase M do Ciclo Celular/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Poli(ADP-Ribose) Polimerases/metabolismo , Proteólise/efeitos dos fármacosRESUMO
Triptolide (TPL) has been shown to inhibit cell proliferation and induce apoptosis in various human cancer cells; however, the precise mechanism of apoptosis induced by TPL in human melanoma cells has not yet been elucidated. In this study, we investigated the precise mechanism underlying cytocidal effects of TPL on human melanoma cells. Treatment of human melanoma cells with TPL significantly inhibited cell growth and induced apoptosis, as evidenced by flow cytometry and annexin V-fluorescein isothiocyanate analyses. TPL increased the levels of Fas and Fas-associated death domain (FADD) and induced cleavage of Bid by activation of caspase-8 and cytochrome c release from mitochondria to the cytosol, which resulted in activation of caspase-9 and caspase-3. Moreover, TPL-induced apoptosis in SK-MEL-2 cells was mediated through dephosphorylation of focal adhesion kinase (FAK) and its cleavage by caspase-8-mediated caspase-3 activation via upregulation of Fas expression. We also found that TPL mediated the dissociation of receptor-interacting protein (RIP) from FAK and enhanced the formation of RIP/Fas complex formation initiating cell death. In conclusion, our data firstly demonstrated that TPL induces apoptosis by both extrinsic and intrinsic apoptosis pathways in human melanoma cells and identified that RIP shuttles between Fas and FAK to mediate apoptosis.
RESUMO
An 80% ethanol extract of Hizikia fusiforme was obtained and followed by successive fractionation using the organic solvents n-hexane, ethyl acetate, and n-butanol to identify the antioxidative substance. The aqueous part of the nbutanol fractionation step, showing high antioxidative activity, was subjected to reverse-phase liquid chromatography. As a result, a substance purified from a BB-2 fraction showed high antioxidative activity. The m/z 419 [M+H] molecular ion peak in the fraction was observed by the analysis of the ESI-LC/MS spectrum. By the analysis of 1H NMR (500 MHz, DMSO-d6) and 13C NMR (125 MHz, DMSO-d6) spectra, a unique compound of the fraction was biochemically identified as a 5-hydroxy-3,6,7,8,3´,4´- hexamethoxyflavone (5HHMF). We also investigated the effect of 5HHMF on human gastric AGS carcinoma cells. Western blot analysis suggested that the flavone substantially increased the levels of the death receptor-associated apoptosis mediators Fas, Fas L, FADD, TRADD, and DR4 in a concentration-dependent manner. The levels of Fas, Fas L, TRADD, and DR4 in the cells treated with 5HHMF (5 microgram/ml) were approximately 26.4-, 12.8-, 6.7-, and 9.8- times higher than those of non-treated cells, respectively. Of note, the level of FADD protein in the cells exposed to 5HHMF (1 microgram/ml) increased approximately 9.6-times. In addition, the cleavage of caspase-3, -8, and -9 in cultured AGS cells treated with 5HHMF was significantly confirmed. Therefore, our results suggest that 5HHMF from H. fusiforme is involved in the induction of death receptor-associated apoptosis mediators in human gastric AGS carcinoma cells.
Assuntos
Apoptose/efeitos dos fármacos , Flavonas/farmacologia , Phaeophyceae/química , Extratos Vegetais/farmacologia , Neoplasias Gástricas/tratamento farmacológico , Western Blotting , Caspases/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Proteínas Adaptadoras de Sinalização de Receptores de Domínio de Morte/metabolismo , Flavonas/química , Flavonas/isolamento & purificação , Humanos , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Espécies Reativas de Oxigênio/metabolismo , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologiaRESUMO
Oleifolioside A, a new triterpenoid compound isolated from Dendropanax morbifera Leveille (D. morbifera), was shown in this study to have potent inhibitory effects on lipopolysaccharide (LPS-)stimulated nitric oxide (NO) and prostaglandin E(2) (PGE(2)) production in RAW 264.7 macrophages. Consistent with these findings, oleifolioside A was further shown to suppress the expression of LPS-stimulated inducible nitric oxide synthase (iNOS) and cyclooxigenase-2 (COX-2) in a dose-dependent manner at both the protein and mRNA levels and to significantly inhibit the DNA-binding activity and transcriptional activity of NF-κB in response to LPS. These results were found to be associated with the inhibition of the degradation and phosphorylation of IκB-α and subsequent translocation of the NF-κB p65 subunit to the nucleus. Inhibition of NF-κB activation by oleifolioside A was also shown to be mediated through the prevention of p38 MAPK and ERK1/2 phosphorylation. Taken together, our results suggest that oleifolioside A has the potential to be a novel anti-inflammatory agent capable of targeting both the NF-κB and MAPK signaling pathways.
RESUMO
In this study, the effect of cordycepin (3'-deoxyadenosine), a major component of Cordyceps militaris, an ingredient of traditional Chinese medicine was investigated for the first time on apoptotsis in human neuroblastoma SK-N-BE(2)-C and melanoma SK-MEL-2 cells. Cordycepin significantly inhibited the proliferation of human neuroblastoma SK-N-BE(2)-C and human melanoma SK-MEL-2 cells with IC50 values of 120 microM and 80 microM, respectively. Cordycepin treatment at 120 microM and 80 microM, respectively, induced apoptosis in both cells and caused the increase of cell accumulation in a time-dependent manner at the apoptotic sub-G1 phase, as evidenced by the flow cytometry (FCM) and annexin V-fluorescein isothiocyanate (FITC) analyses. Western blot analysis revealed the induction of active caspase-3 and poly(ADP-ribose)polymerase (PARP) cleavage by cordycepin treatment. These results suggest that cordycepin is a potential candidate for cancer therapy of neuroblastoma and melanoma.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Desoxiadenosinas/farmacologia , Melanoma/patologia , Neuroblastoma/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , HumanosRESUMO
Apoptosis, the main type of programmed cell death, plays an essential role in a variety of biological events. Whereas "classical" apoptosis is dependent on caspase activation, caspase-independent death is increasingly recognized as an alternative pathway. To develop new anticancer agents, oleifolioside A was isolated from Dendropanax morbifera Leveille and the biochemical mechanisms of oleifolioside A-induced apoptosis in HeLa cells were investigated. Exposure to oleifolioside A resulted in caspase activation and typical features of apoptosis, although cell death was not prevented by caspase inhibition. Oleifolioside A treatment induced up-regulation of Bad, loss of mitochondrial membrane potential, nuclear relocation of mitochondrial factors, apoptosis-inducing factor (AIF), endonuclease G (EndoG), and apoptosis induction. This is the first report of anticancer activity of oleifolioside A, and nuclear translocation of AIF and EndoG in oleifolioside A-treated HeLa cells might represent an alternative death signaling pathway in the absence of caspase activity.
Assuntos
Fator de Indução de Apoptose/metabolismo , Apoptose/efeitos dos fármacos , Araliaceae/química , Núcleo Celular/metabolismo , Endodesoxirribonucleases/metabolismo , Extratos Vegetais/farmacologia , Saponinas/farmacologia , Neoplasias do Colo do Útero/metabolismo , Caspases/metabolismo , Núcleo Celular/enzimologia , Feminino , Células HeLa , Humanos , Transporte Proteico , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/enzimologia , Neoplasias do Colo do Útero/fisiopatologiaRESUMO
The expression of matrix metalloproteinase (MMPs)-9 is critical for cell migration and can lead to invasion and metastasis of cancer cells. In the present study, we examined the inhibitory effects of JNP3, a new compound which was isolated from traditional Chinese medicine, on cell invasion and MMP-9 activation in phorbol myristate acetate (PMA)-induced MCF-7 cells. Treatment with JNP3 significantly and selectively inhibited PMA-induced MMP-9 secretion, mRNA expression and protein levels, and these results led to reduction of cell invasion and migration in PMA-induced MCF-7 cells. The results of MMP-9 promoter assay and EMSA showed that JNP3 specifically inhibited PMA-induced MMP-9 gene expression by blocking NF-κB-dependent transcriptional activity. In addition, PMA-induced phosphorylation of ERK1/2 and JNK were suppressed by JNP3 treatment, whereas the phosphorylation of p38 MAPK was not affected by JNP3. These results suggest that JNP3 can be potential anti-cancer agents through specific inhibition of NF-κB-dependent MMP-9 gene expression.
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Inibidores de Metaloproteinases de Matriz , NF-kappa B/antagonistas & inibidores , Triterpenos/farmacologia , Linhagem Celular Tumoral , Regulação para Baixo , Feminino , Expressão Gênica/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , MAP Quinase Quinase 4/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Metaloproteinase 9 da Matriz/genética , Invasividade Neoplásica , Fosforilação/efeitos dos fármacos , Acetato de Tetradecanoilforbol/farmacologia , Triterpenos/químicaRESUMO
We assessed the effects of chloroform extract of fermented Viola mandshurica (CEFV) on melanogenesis B16 melanoma cells. CEFV treatment significantly decreased melanin content and tyrosinase activity in dose-dependent manners. To elucidate the mechanism of the inhibitory effects of CEFV on melanogenesis, we performed RT-PCR and Western blotting for melanogenesis-related genes such as tyrosinase, tyrosinase-related protein-1 (TRP-1), TRP-2, and microphthalmia-associated transcription factor (MITF). CEFV strongly inhibited mRNA as well as the protein expression of tyrosinase and MITF, but had no significant effect on TRP-1 or TRP-2 expressions. It markedly decreased the phosphorylation of cAMP responsive element binding protein (CREB), and induced the duration of extracellular signal-regulated kinase (ERK) activation, leading to reduction of MITF expression and subsequently that of tyrosinase. Therefore, we suggest that CEFV induces downregulation of melanogenesis through decreased CREB phosphorylation and ERK activation.
Assuntos
Fermentação , Melaninas/biossíntese , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Extratos Vegetais/farmacologia , Viola/química , Viola/metabolismo , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Regulação para Baixo/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Melanoma Experimental/enzimologia , Melanoma Experimental/genética , Camundongos , Fator de Transcrição Associado à Microftalmia/metabolismo , Monofenol Mono-Oxigenase/metabolismo , Fosforilação/efeitos dos fármacos , Extratos Vegetais/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
Investigation of collagenase and gelatinase inhibitory natural components afforded two isoflavonoids. Two isoflavonoids, tectorigenin-7-O-ß-D-glucoside (1) and luteolin-7-O-ß-D-glucuronopyranoside (2), were isolated from ethyl acetate fraction of Viola patrinii fermentation extracts (VPFE). Of these, compounds 1 and 2 exhibited collagenase inhibitory activity (IC(50)) at a concentration of less than 1.5 µM, and compound 2 showed gelatinases A and B inhibitory activity (IC(50)) at 0.3 µM and 0.8 µM, respectively.
Assuntos
Fermentação , Gelatinases/antagonistas & inibidores , Inibidores de Metaloproteinases de Matriz , Extratos Vegetais/química , Inibidores de Proteases/química , Inibidores de Proteases/isolamento & purificação , Viola/microbiologia , Biocatálise/efeitos dos fármacos , Cromatografia/métodos , Colagenases/metabolismo , Flavonoides/química , Flavonoides/isolamento & purificação , Flavonoides/farmacologia , Gelatinases/metabolismo , Concentração Inibidora 50 , Isoflavonas/química , Isoflavonas/isolamento & purificação , Isoflavonas/farmacologia , Luteolina/química , Luteolina/isolamento & purificação , Luteolina/farmacologia , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Estrutura Molecular , Inibidores de Proteases/farmacologia , Viola/metabolismoRESUMO
Our previous study has demonstrated that the methanol extract of Hyul-Tong-Ryung (HM) specifically suppresses the phorbol 12-myristate 13-acetate (PMA)-induced matrix metalloproteinase-9 (MMP-9) production through the inhibition of MMP-9 mRNA expression in MCF-7 human breast carcinoma cells. However, the molecular mechanisms involved in transcriptional suppression of MMP-9 by HM in PMA-induced MCF-7 cells are not known. In this study, we aimed to elucidate the molecular mechanisms involved in the inhibition of MMP-9 expression by HM in PMA-induced MCF-7 cells. The results of promoter assay and EMSA showed that HM specifically inhibits MMP-9 gene expression by blocking PMA-stimulated activation of activator protein-1 (AP-1). In addition, PMA-stimulated phosphorylation of extracellular signal regulated kinase 1/2 (ERK 1/2) was suppressed by HM treatment, whereas the phosphorylation of either c-Jun N-terminal kinase (JNK) or p38 mitogen-activated protein kinase (MAPK) was not affected. HM could inhibit the PMA-induced MMP-9 expression through suppression of the transcriptional activity of MMP-9 gene in MCF-7 cells. These results indicate that HM inhibits PMA-induced MMP-9 expression by blocking the activation of activator protein-1 (AP-1) via extracellular signal regulated kinase 1/2 (ERK 1/2) signaling pathway.
Assuntos
Medicamentos de Ervas Chinesas/farmacologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Expressão Gênica/efeitos dos fármacos , Metaloproteinase 9 da Matriz/metabolismo , Transdução de Sinais/fisiologia , Acetato de Tetradecanoilforbol/farmacologia , Fator de Transcrição AP-1/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , DNA/genética , DNA/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/antagonistas & inibidores , Feminino , Expressão Gênica/genética , Genes Reporter/genética , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Metaloproteinase 9 da Matriz/genética , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Mutação/genética , NF-kappa B/metabolismo , Fosforilação/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Ligação Proteica/genética , Inibidores de Proteínas Quinases/farmacologia , Elementos de Resposta/genética , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição RelA/metabolismo , TransfecçãoRESUMO
Geiji-Bokryung-Hwan (GBH), consisting of herbes of Cinnamomi ramulus (Geiji), Poria cocos (Bokryun), Mountan cortex radicis (Mokdanpi), Paeoniae radix (Jakyak), and Persicae semen (Doin), on antiplatelet activity in human platelet suspensions was studied. The mechanism involved in the antiplatelet activity of GBH in human platelet suspensions was investigated. GBH did not significantly affect the thromboxane synthetase activity of aspirin-treated platelet microsomes and GBH (15 and 30 microg/ml) significantly inhibited [3H]arachidonic acid released in collagen-activated platelets but not in unactivated-platelets. Nitric oxide (NO) production in human platelets was measured by a chemiluminesence detection method in this study. GBH did not significantly affect nitrate production in collagen (10 microg/ml)-induced human platelet aggregation. Various concentrations of GBH (0, 5, 10, 15, and 30 microg/ml) dose-dependently inhibited [3H]inositol monophosphate formation stimulated by collagen (10 microg/ml) in [3H]myoinositol-loaded platelets at different incubation times (1, 2, 3, and 5 min). These results indicated that the antiplatelet activity of GBH may possibly be due to the inhibition of phospholipase C (PLC) activity, leading to reduce phosphoinositide breakdown, followed by the inhibition of thromboxane A(2) formation, and then inhibition of [Ca(2+)](i) mobilization of platelet aggregation stimulated by agonists. In conclusion, GBH suppressed PLC in a dose-dependent manner, and may have pharmaceutical applications. These data suggest that GBH extracts merit investigation as a potential anti-atherosclerogenic agent in humans.