RESUMO
High-mobility group box 1 (HMGB1), a highly conserved chromosome protein, is considered as a potential therapeutic target and novel biomarker because of its regulation in the proliferation and metastasis of Hepatocellular carcinoma (HCC). Calenduloside E (CE), a natural active product, has been reported to anti-cancer effect. However, the role and underlying molecular mechanism of CE in HCC is still unclear. The purpose of this study is to investigate the effects of CE on the proliferation and migration of HCC, and then explore the possible underlying molecular mechanism. HepG2 cells were treated with CE or transfected with HMGB1 shRNA plasmids, EdU and colony formation assays were used to detect cell proliferation ability. Wound healing and transwell assays were used to determine the role of CE in cell migration. The expression of Cyclins, PCNA, MMPs, HMGB1, N-cadherin, E-cadherin and phosphorylation of p38, ERK and JNK were all detected using Western blotting. Our results showed that CE inhibited HepG2 cells proliferation and migration in a dose dependent manner; reduced the expression levels of Cycins, PCNA, HMGB1, MMPs and N-cadherin; up-regulated E-cadherin expression; enhanced the phosphorylation of p38 and JNK signalling pathways. Blocking the activation of p38 and JNK obviously reversed CE-mediated inhibitory effects on HepG2 cell proliferation and migration; reversed CE-induced down-regulation of Cyclins, PCNA, MMPs, N-cadherin and HMGB1, as well as E-cadherin up-regulation. In conclusion, our study suggested that CE reduces the expression levels of Cyclins, MMPs and epithelial-mesenchymal transformation (EMT) through p38/JNK-HMGB1 signaling axis and then inhibits HepG2 cells proliferation and migration in HepG2 cells. This study provides a new perspective for the anti-tumour molecular mechanism of CE in HCC.
Assuntos
Carcinoma Hepatocelular/patologia , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/genética , Proteína HMGB1/metabolismo , Neoplasias Hepáticas/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Ácido Oleanólico/análogos & derivados , Saponinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Carcinoma Hepatocelular/tratamento farmacológico , Células Hep G2 , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Ácido Oleanólico/farmacologia , FitoterapiaRESUMO
A potential method called microwave-assisted aqueous two-phase extraction (MA-ATPE) was developed for concurrent extraction and purification of gentiopicroside from Gentiana scabra Bunge. Formation characteristics of aqueous two-phase system (ATPS) composed of ethanol and 25 kinds of salts were investigated; K2HPO4 (w/w, 21.71%) and ethanol (w/w, 40.72%) were determined to be the optimal compositions of ATPS. Response surface methodology based on Box-Behnken design was used to investigate the extraction conditions, the optimal parameters were summarized as follows: 80°C of extraction temperature, 31 s of extraction time, 11:1 (mL/g) of liquid-to-solid ratio, 100 meshes of particle size and 806 W of microwave power. Under these conditions, the extraction yield of gentiopicroside was 65.32 ± 0.24 mg/g with a recovery of 96.51%. Compared with other four methods, the purity of gentiopicroside in the crude extracts reached 17.16 ± 0.25%, which was significantly higher than that of smashing tissue extraction, microwave assisted-extraction, ultrasonic-assisted extraction and heat reflux extraction, respectively. In addition, the phase-forming salt can be recyclable. Therefore, MA-ATPE was an excellent and alternative technique to the conventional extraction approaches of gentiopicroside.
Assuntos
Etanol/química , Gentiana/química , Glucosídeos Iridoides/química , Glucosídeos Iridoides/isolamento & purificação , Micro-Ondas , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Reprodutibilidade dos TestesRESUMO
The IL-17 receptor (IL-17R) has a perplexing role in cancer, which may be explained by its yin-yang signaling pathways. Recently, the critical role of IL-17R in maintaining basal levels of A20-a key negative regulator of NF-κB and JNK-c-Jun pathways has been demonstrated in cancer cell lines. Cross-cancer analyses of somatic copy number alterations in IL-17RA, IL-17RC and A20 genes reveal that IL-17RA-deletion is common in colorectal cancer (CRC) patients, representing 24, 26, 37 and 49% of stage I, II, III and IV of patients, respectively, and mutually exclusive with patients displaying microsatellite instability. Importantly, patients with IL-17R-deletion or concurrent deletions of A20 show significantly reduced overall survival. Analysis of multiple published microarray studies confirms that IL-17RA expression is significantly reduced in CRC samples compared to normal counterparts, and its level is closely associated with A20 expression. Analyses of RNAseq data indicate that tumors with IL-17R-deletion express strong molecular markers of tumor invasion, growth and metastasis. Notably, approximately 20 genes responsible for protein synthesis and mitochondrial metabolism are inversely correlated with both IL-17RA and A20. Immunohistochemistry staining in human colorectal tissue arrays further reveals that high-grade tumors have significantly reduced IL-17RA staining compared to low-grade tumors. Thus, collective evidence strongly supports a previously unrecognized CRC-promoting mechanism triggered by IL-17RA-deletion and highlights its utility as a prognostic marker in CRC.
Assuntos
Neoplasias Colorretais/patologia , Regulação para Baixo , Deleção de Genes , Receptores de Interleucina-17/genética , Receptores de Interleucina-17/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Instabilidade de Microssatélites , Gradação de Tumores , Invasividade Neoplásica , Metástase Neoplásica , Estadiamento de Neoplasias , Análise de Sobrevida , Proteína 3 Induzida por Fator de Necrose Tumoral alfa/genéticaRESUMO
Insects and pathogenic infections (bacteria, viruses and fungi) cause huge losses in agriculturally important crops yearly. Due to the rise in pesticide and antibiotic resistance, our crops and livestock are increasingly at risk. There is a rising demand for environmentally friendly solutions to prevent crop decreases. Components of Ascophyllum nodosum seaweed extracts were recently found to boost plant immunity. The stimulatory activities of the A.nodosum marine alga-derived extract (Stella Maris®) were investigated in a broad range of immune assays. Elevated hydrogen peroxide production measured in a chemiluminescence assay suggested that the extract elicited a strong burst of reactive oxygen species. Arabidopsis seedlings treated with Stella Maris® activated the expression of WRKY30, CYP71A12 and PR-1 genes, the induction of which represent early, mid and late plant immune response, respectively. Finally, this study found that Stella Maris® inhibited the growth of multiple bacterial pathogens, including an opportunistic human pathogen that has demonstrated pathogenicity in plants. In summary, the pre-treatment with the seaweed extract protected Arabidopsis against subsequent infection by these pathogens.
Assuntos
Arabidopsis/efeitos dos fármacos , Imunidade Inata/efeitos dos fármacos , Doenças das Plantas/imunologia , Extratos Vegetais/farmacologia , Alga Marinha/química , Arabidopsis/imunologia , Ascophyllum/química , Substâncias Protetoras/farmacologia , Espécies Reativas de Oxigênio/metabolismoRESUMO
Toll-like receptors (TLRs) recognize the conserved molecular patterns in microorganisms and trigger myeloid differentiation primary response 88 (MyD88) and/or TIR-domain-containing adapter-inducing interferon-ß (TRIF) pathways that are critical for host defense against microbial infection. However, the molecular mechanisms that govern TLR signaling remain incompletely understood. Regulator of calcineurin-1 (RCAN1), a small evolutionarily conserved protein that inhibits calcineurin phosphatase activity, suppresses inflammation during Pseudomonas aeruginosa infection. Here, we define the roles for RCAN1 in P. aeruginosa lipopolysaccharide (LPS)-activated TLR4 signaling. We compared the effects of P. aeruginosa LPS challenge on bone marrow-derived macrophages from both wild-type and RCAN1-deficient mice and found that RCAN1 deficiency increased the MyD88-NF-κB-mediated cytokine production (IL-6, TNF and MIP-2), whereas TRIF-interferon-stimulated response elements (ISRE)-mediated cytokine production (IFNß, RANTES and IP-10) was suppressed. RCAN1 deficiency caused increased IκBα phosphorylation and NF-κB activity in the MyD88-dependent pathway, but impaired ISRE activation and reduced IRF7 expression in the TRIF-dependent pathway. Complementary studies of a mouse model of P. aeruginosa LPS-induced acute pneumonia confirmed that RCAN1-deficient mice displayed greatly enhanced NF-κB activity and MyD88-NF-κB-mediated cytokine production, which correlated with enhanced pulmonary infiltration of neutrophils. By contrast, RCAN1 deficiency had little effect on the TRIF pathway in vivo. These findings demonstrate a novel regulatory role of RCAN1 in TLR signaling, which differentially regulates MyD88 and TRIF pathways.
Assuntos
Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Musculares/metabolismo , Fator 88 de Diferenciação Mieloide/metabolismo , Transdução de Sinais , Receptores Toll-Like/metabolismo , Animais , Proteínas de Ligação ao Cálcio , Quimiocinas/metabolismo , Regulação para Baixo/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Fator Regulador 7 de Interferon/genética , Fator Regulador 7 de Interferon/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/deficiência , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Proteínas Musculares/deficiência , Inibidor de NF-kappaB alfa/metabolismo , NF-kappa B/metabolismo , Infiltração de Neutrófilos/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Pseudomonas aeruginosa/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Elementos de Resposta/genética , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacosRESUMO
Schisandra chinensis is one of the most famous herbal medicines in China, Korea and Japan. It has been widely used as a tonic, sedative, anti-aging and astringent agent. Lignans are one of its main bioactive components. The classical methods for extracting lignans, however, were tedious and energy-consuming. With the aim to develop an effective extraction method of lignans, the smashing tissue extraction (STE) technique was adopted and optimized in this study. Extraction conditions of STE have been optimized by the response surface methodology based on the Box-Behnken design. Results showed that 75% aqueous ethanol was the optimal extraction solvent, and the other optimal conditions were as follows: extraction voltage of 180 V, extraction time of 1 min, solid-liquid ratio of 1 : 19 and sample particle size of 120 mesh. Under these optimized conditions, the total content of the five lignans (Schisandrol A, Schisantherin A, Deoxyschisandrin, Schisandrin B and Schisandrin C) in S. chinensis collected from Baishan City located in the northeast of China was 13.89 ± 0.014 mg/g, which was well matched with the value predicted by the model. Other techniques, including heat reflux, Soxhlet, ultrasonic-assisted and microwave-assisted extraction, were further compared. Results suggested that STE had the highest extraction efficiency of lignans with the shortest time. It indicates that the approach proposed in this study is a simple and efficient technique for the extraction of lignans in S. chinensis.
Assuntos
Métodos Analíticos de Preparação de Amostras/métodos , Frutas/química , Lignanas/isolamento & purificação , Extratos Vegetais/isolamento & purificação , Schisandra/química , Métodos Analíticos de Preparação de Amostras/instrumentação , Lignanas/análise , Micro-Ondas , Extratos Vegetais/análiseRESUMO
A microwave-assisted enzymatic extraction (MAEE) method had been developed, which was optimized by response surface methodology (RSM) and orthogonal test design, to enhance the extraction of crude polysaccharides (CPS) from the fruit of Schisandra chinensis Baill. The optimum conditions were as follows: microwave irradiation time of 10 min, extraction pH of 4.21, extraction temperature of 47.58°C, extraction time of 3h and enzyme concentration of 1.5% (wt% of S. chinensis powder) for cellulase, papain and pectinase, respectively. Under these conditions, the extraction yield of CPS was 7.38 ± 0.21%, which was well in close agreement with the value predicted by the model. The three methods including heat-refluxing extraction (HRE), ultrasonic-assisted extraction (UAE) and enzyme-assisted extraction (EAE) for extracting CPS by RSM were further compared. Results indicated MAEE method had the highest extraction yields of CPS at lower temperature. It was indicated that the proposed approach in this study was a simple and efficient technique for extraction of CPS in S. chinensis Baill.
Assuntos
Frutas/química , Micro-Ondas , Extratos Vegetais/química , Polissacarídeos/química , Schisandra/química , Fracionamento Químico/métodos , Polissacarídeos/ultraestruturaRESUMO
A method for two-steps extraction of essential oil, polysaccharides and lignans from Schisandra chinensis Baill had been established. Firstly, S. chinensis was extracted by hydro-distillation, the extracted solution was separated from the water-insoluble residue and precipitated by adding dehydrated alcohol after the essential oil was collected, and then the precipitate as polysaccharide was collected. Finally, second extraction was performed to obtained lignans from the water-insoluble residue with ultrasonic-microwave assisted extraction (UMAE) method. Response surface methodology was employed to optimize the UMAE parameters, the optimal conditions were as follows: microwave power 430W, ethanol concentration 84%, particle size of sample 120-mesh sieves, ratio of water to raw material 15 and extraction time 2.1min. Under these optimized conditions, the total extraction yields of five lignans (Schisandrol A, Schisantherin A, Deoxyschisandrin, Schisandrin B and Schisandrin C) had reached 14.22±0.135mg/g. Compared with the traditional method of direct extraction of different bioactive components in respective procedure, the extraction yields of polysaccharides and the five lignans had reached 99% and 95%, respectively. The mean recoveries of the 5 lignan compounds and polysaccharides were 97.75-101.08% and their RSD value was less than 3.88%.The approach proposed in this study not only improved the extraction yield of lignans, but also elevated the utilization of Schisandra resources.
Assuntos
Lignanas/isolamento & purificação , Óleos Voláteis/isolamento & purificação , Polissacarídeos/isolamento & purificação , Schisandra/química , Compostos de Bifenilo/análise , Compostos de Bifenilo/isolamento & purificação , Ciclo-Octanos/análise , Ciclo-Octanos/isolamento & purificação , Frutas , Lignanas/análise , Micro-Ondas , Óleos Voláteis/análise , Extratos Vegetais/química , Polissacarídeos/análise , Ultrassom/métodosRESUMO
Baicalin purified from the root of Radix scutellariae is widely used in clinical practices. This study aimed to evaluate the effect of baicalin on the pharmacokinetics of nifedipine, a CYP3A probe substrate, in rats in vivo and in vitro. In a randomised, three-period crossover study, significant changes in the pharmacokinetics of nifedipine (2 mg/kg) were observed after treatment with a low (0.225 g/kg) or high (0.45 g/kg) dose of baicalin in rats. In the low- and high-dose groups of baicalin-treated rats, C max of total nifedipine decreased by 40%±14% (P<0.01) and 65%±14% (P<0.01), AUC0-∞ decreased by 41%±8% (P<0.01) and 63%±7% (P<0.01), Vd increased by 85%±43% (P<0.01) and 224%±231% (P<0.01), and CL increased by 97%±78% (P<0.01) and 242%±135% (P<0.01), respectively. Plasma protein binding experiments in vivo showed that C max of unbound nifedipine significantly increased by 25%±19% (P<0.01) and 44%±29% (P<0.01), respectively, and there was a good correlation between the unbound nifedipine (%) and baicalin concentrations (P<0.01). Furthermore, in vitro results revealed that baicalin was a competitive displacer of nifedipine from plasma proteins. In vitro incubation experiments demonstrated that baicalin could also competitively inhibit CYP3A activity in rat liver microsomes in a concentration-dependent manner. In conclusion, the pharmacokinetic changes of nifedipine may be modulated by the inhibitory effects of baicalin on plasma protein binding and CYP3A-mediated metabolism.
Assuntos
Proteínas Sanguíneas/metabolismo , Inibidores do Citocromo P-450 CYP3A , Flavonoides/farmacologia , Nifedipino/farmacocinética , Ligação Proteica/efeitos dos fármacos , Animais , Estudos Cross-Over , Interações Medicamentosas , Masculino , Microssomos Hepáticos/metabolismo , Extratos Vegetais/farmacologia , Ratos , Ratos Sprague-DawleyRESUMO
Baicalin, a flavonoid compound isolated from Scutellaria baicalensis, has been shown to possess antiinflammatory, antiviral, antitumour, and immune regulatory properties. The present study evaluated the potential herb-drug interaction between baicalin and midazolam in rats. Coadministration of a single dose of baicalin (0.225, 0.45, and 0.90 g/kg, i.v.) with midazolam (10 mg/kg, i.v.) in rats resulted in a dose-dependent decrease in clearance (CL) from 25% (P < 0.05) to 34% (P < 0.001) with an increase in AUC0-∞ from 47% (P < 0.05) to 53% (P < 0.01). Pretreatment of baicalin (0.90 g/kg, i.v., once daily for 7 days) also reduced midazolam CL by 43% (P < 0.001), with an increase in AUC0-∞ by 87% (P < 0.01). Multiple doses of baicalin decreased the expression of hepatic CYP3A2 by approximately 58% (P < 0.01) and reduced midazolam 1'-hydroxylation by 23% (P < 0.001) and 4'-hydroxylation by 21% (P < 0.01) in the liver. In addition, baicalin competitively inhibited midazolam metabolism in rat liver microsomes in a concentration-dependent manner. Our data demonstrated that baicalin induced changes in the pharmacokinetics of midazolam in rats, which might be due to its inhibition of the hydroxylation activity and expression of CYP3A in the liver.
RESUMO
Baicalin has been shown to possess many pharmacological effects, including antiviral, antioxidant, anti-cancer and anti-inflammatory properties. In the current study, we reveal the inhibitory effects of baicalin on the metabolism of dextromethorphan (DXM), a dual probe substrate of CYP2D and CYP3A, in rats. Lineweaver-Burk plots demonstrated that baicalin inhibited the activities of CYP2D and CYP3A in a non-competitive manner in rat liver microsomes (RLMs). Concomitant administration of baicalin (0.90 g/kg, i.v.) and DXM (10 mg/kg, i.v.) increased the maximum drug concentration (C(max)) (37%) and the area under concentration-time curve (AUC) (42%) and decreased the clearance (CL) (27%) of DXM in a randomised, crossover study in rats (P < 0.01). The change in the AUC of DXM was significantly correlated with the C(max) and AUC of baicalin (P < 0.05). The inhibitory effects of multiple doses of baicalin (0.90 g/kg, i.v., 12 days) on the metabolism of DXM were similar to those observed following a single dose in rats. The activity of CYP3A in excised liver samples from rats following multiple baicalin treatment was significantly decreased compared to that of the control group (P < 0.05), whereas multiple doses of baicalin had no obvious effect on the activity of CYP2D. Taken together, these data demonstrate that baicalin inhibits the metabolism of DXM in a concentration-dependent manner in rats, possibly through inhibiting hepatic CYP2D and CYP3A activities.