RESUMO
Codonopsis pilosula polysaccharides (CPP), the main active ingredient of Codonopsis pilosula, has gained significant attention as a liver-protective agent. Previous studies have demonstrated that CPP could alleviate gut microbiota dysbiosis in colitis or obese mice. However, the effects of CPP on mycotoxin-induced liver injury and gut microbiota dysbiosis are still poorly understood. In this study, we aimed to investigate the protective effects of CPP on sterigmatocystin (STC)-induced liver injury, as well as its regulatory effects on gut microbiota. Our results revealed that CPP intervention significantly alleviated STC-induced liver injury, as evidenced by decreased liver index, reduced liver histopathological changes, and modulation of related molecular markers. Additionally, we found that CPP could alleviate liver injury by reducing liver inflammation and oxidative stress, inhibiting hepatocyte apoptosis, and regulating lipid metabolism. Notably, we also observed that CPP could alleviate STC-induced gut microbiota dysbiosis by modulating the diversity and richness of gut microbiota, suggesting that gut microbiota modulation may also serve as a mechanism for CPP-mediated remission of liver injury. In summary, our study not only provided a new theoretical basis for understanding the hepatotoxicity of STC and the protective effects of CPP against STC-induced liver injury, but also provided new perspectives for the application of CPP in the fields of food, healthcare products, and medicine.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas , Codonopsis , Disbiose , Microbioma Gastrointestinal , Polissacarídeos , Esterigmatocistina , Microbioma Gastrointestinal/efeitos dos fármacos , Animais , Disbiose/tratamento farmacológico , Codonopsis/química , Camundongos , Polissacarídeos/farmacologia , Polissacarídeos/química , Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Masculino , Esterigmatocistina/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologiaRESUMO
Microglia play a significant role in immune defense and tissue repair in the central nervous system (CNS). Microglial activation and the resulting neuroinflammation play a key role in the pathogenesis of neurodegenerative disorders. Recently, inflammation reduction strategies in neurodegenerative diseases have attracted increasing attention. Herein, we discovered and evaluated the anti-neuroinflammatory potential of compounds from the Antarctic fungi strain Aspergillus sp. SF-7402 in lipopolysaccharide (LPS)-stimulated BV2 cells. Four metabolites were isolated from the fungi through chemical investigations, namely, 5-methoxysterigmatocystin (1), sterigmatocystin (2), aversin (3), and 6,8-O-dimethylversicolorin A (4). Their chemical structures were elucidated by extensive spectroscopic analysis and HR-ESI-MS, as well as by comparison with those reported in literature. Anti-neuroinflammatory effects of the isolated metabolites were evaluated by measuring the production of nitric oxide (NO), tumor necrosis factor (TNF)-α, and interleukin (IL)-6 in LPS-activated microglia at non-cytotoxic concentrations. Sterigmatocystins (1 and 2) displayed significant effects on NO production and mild effects on TNF-α and IL-6 expression inhibition. The molecular mechanisms underlying this activity were investigated using Western blot analysis. Sterigmatocystin treatment inhibited NO production via downregulation of inducible nitric oxide synthase (iNOS) expression in LPS-stimulated BV2 cells. Additionally, sterigmatocystins reduced nuclear translocation of NF-κB. These results suggest that sterigmatocystins present in the fungal strain Aspergillus sp. are promising candidates for the treatment of neuroinflammatory diseases.
Assuntos
Microglia , NF-kappa B , Regiões Antárticas , Anti-Inflamatórios/química , Aspergillus/metabolismo , Interleucina-6/metabolismo , Lipopolissacarídeos/metabolismo , Lipopolissacarídeos/farmacologia , NF-kappa B/metabolismo , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Transdução de Sinais , Esterigmatocistina/metabolismo , Esterigmatocistina/farmacologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
The chemical investigation on endophytic fungus Annulohypoxylon cf. stygium in leaves of Anoectochilus roxburghii (Wall.) Lindl. has been performed. Sixteen compounds were isolated and their structures were identified as (-)-notoamide A, (-)-notoamide B, (+)-versicolamide B, notoamide C, notoamide D, stephacidin A, sterigmatocystin, dihydrosterigmatocystin, secosterigmatocystin, versiconol, averufanin, kipukasin D, kipukasin E, diorcinal, palmarumycin CP2 and (-)-(3R)-mellein methyl ether, respectively, by spectroscopic analysis and comparison with literature data. All the compounds were isolated from Annulohypoxylon genus for the first time. Sterigmatocystin and palmarumycin CP2 showed selective cytotoxic activities against HepG2, HeLa, MCF-7 and HT-29.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Ascomicetos/química , Naftalenos/farmacologia , Orchidaceae/microbiologia , Folhas de Planta/microbiologia , Compostos de Espiro/farmacologia , Esterigmatocistina/farmacologia , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/isolamento & purificação , Ascomicetos/metabolismo , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Estrutura Molecular , Naftalenos/química , Naftalenos/isolamento & purificação , Compostos de Espiro/química , Compostos de Espiro/isolamento & purificação , Esterigmatocistina/química , Esterigmatocistina/isolamento & purificaçãoRESUMO
Since humans are exposed to different mycotoxins through daily intake, there is increasing concern about the adverse effects of the interactions between them. Cytotoxicity of sterigmatocystin (STE) and nivalenol (NIV) alone and in combination in human hepatocarcinoma (HepG2) cells was evaluated by MTT assay. Furthermore, ROS production and alteration of ΔΨm as mechanisms of action were assessed. Cells were treated with concentrations ranging from 0.15 to 5 µM for NIV and from 0.78 to 50 µM for STE individually and in binary combinations. The combination ratio between the mixture STE + NIV was 10:1. The IC50 values of NIV ranged from 0.96 to 0.66 µM, whereas no IC50 values were obtained for STE at any time tested. For the combinations studied, synergistic, antagonistic and addictive effects were obtained with the two type of analyses performed, the isobologram analysis and the Combenefit method. No relevant effects on ROS and ΔΨm were observed. In conclusion, predictive models based on combination data could help to better understand the interactions between mycotoxins and their implications in food safety assessment. However, a further analysis of the molecular mechanism underlying these interactive effects is required.
Assuntos
Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Esterigmatocistina/farmacologia , Tricotecenos/farmacologia , Antineoplásicos/farmacologia , Sinergismo Farmacológico , Células Hep G2 , Humanos , Estrutura Molecular , Esterigmatocistina/química , Tricotecenos/químicaRESUMO
Molecular networking approach was applied for the targeted isolation of new sterigmatocystin derivatives, sterigmatocystins A-C, from the marine sponge-derived fungus Aspergillus versicolor. Sterigmatocystin A features a rare 6/6/6/6/5 polycyclic system. The structures of sterigmatocystins A-C, including absolute configurations, were determined on the basis of spectroscopic data and ECD calculations. Sterigmatocystin A showed more stronger promoting angiogenesis activity than the positive control at 1.25â µM level in transgenic fluorescent zebrafish. Sterigmatocystins A-C also exhibited moderate antiviral activity by the inhibition of HSV-2.
Assuntos
Antivirais/química , Aspergillus/química , Esterigmatocistina/análogos & derivados , Inibidores da Angiogênese/química , Inibidores da Angiogênese/isolamento & purificação , Inibidores da Angiogênese/farmacologia , Animais , Animais Geneticamente Modificados/metabolismo , Antivirais/isolamento & purificação , Antivirais/farmacologia , Aspergillus/metabolismo , Dicroísmo Circular , Herpesvirus Humano 2/efeitos dos fármacos , Humanos , Espectroscopia de Ressonância Magnética , Conformação Molecular , Neovascularização Fisiológica/efeitos dos fármacos , Poríferos/microbiologia , Esterigmatocistina/isolamento & purificação , Esterigmatocistina/farmacologia , Peixe-Zebra/metabolismoRESUMO
Sterigmatocystin (STC) is structurally close to the mycotoxin aflatoxin B1 as it shares its biosynthetic pathway with aflatoxins. The purpose of the present study was to investigate the short-term (24â¯h) effects of STC contaminated diet at different doses (1â¯mg, 2â¯mg and 4â¯mg STC kg-1 feed) in one year old common carp juveniles. Liver samples were taken in 8-h intervals. The markers of the lipid peroxidation showed moderate changes after the application of sterigmatocystin-contaminated diet, significant elevations were only observed in the lowest applied dose group of sterigmatocystin after 16â¯h of exposure. Reduced glutathione content showed higher levels than control group after 16â¯h of exposure as effect of low dose of sterigmatocystin. Glutathione peroxidase (GPX4) activity was lower than control in the group treated with 2â¯mg STC kg-1 feed after 24â¯h of exposure. Gene expression measurements of keap1, nrf2, gpx4a, gpx4b and gss genes revealed a dual response. Down-regulation or near control values were observed 8â¯h after exposure which was followed by an induction 16 and 24â¯h after exposure. In case of gsr, gene expression values returned to control levels by the 24th hour. In summary, these results suggest that lower doses of STC caused oxidative stress earlier than higher doses, which efficiently activated the Keap1-Nrf2 pathway, while higher doses revealed long-drawn activation of this pathway.
Assuntos
Carpas/genética , Carpas/metabolismo , Glutationa Peroxidase/metabolismo , Glutationa/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Fígado/efeitos dos fármacos , Esterigmatocistina/farmacologia , Animais , Regulação da Expressão Gênica/efeitos dos fármacos , Genes Reguladores/efeitos dos fármacos , Glutationa Peroxidase/genética , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Fígado/enzimologia , Fígado/metabolismo , Oxirredução/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Fosfolipídeo Hidroperóxido Glutationa Peroxidase , RNA/genética , RNA/metabolismoRESUMO
Sterigmatocystin (ST), being a precursor of aflatoxin, is categorized as Group 2B carcinogen. Our previous studies found that both mismatch repair (MMR) pathways and p53 signaling pathway were involved in ST-induced G2 cell cycle arrest in human esophageal squamous epithelial cell line, HET-1A, in vitro. Studies showed that ERK, JNK and p38 signaling pathways played important roles in cell cycle arrest induced by several other carcinogens. However, the role of MAPK pathway and the links between the MMR and p53 signaling pathways in ST induced G2 phase arrest is still not clarified. In the present study, we first explored the role of MAPK pathway upon ST induced G2 arrest, and found that ST up-regulated the expression of G2/M regulatory factors through MAPK signaling pathway (both ERK and p38, but not JNK pathway). The inhibition of ERK and p38 significantly inhibited p53 activation by ST. Blockage of MMR pathway by silencing hMLH1 expression inhibited ERK, p38 and p53 activation and then attenuated G2 arrest by ST. Thus, in conclusion, the current study demonstrated that in response to ST induced DNA damage, hMLH1 was first activated, then triggered ERK, p38 and p53 activation and finally resulted in G2 arrest in HET-1A cells.
Assuntos
Células Epiteliais/efeitos dos fármacos , Esôfago/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Proteína 1 Homóloga a MutL/metabolismo , Esterigmatocistina/farmacologia , Proteína Supressora de Tumor p53/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Linhagem Celular Tumoral , Dano ao DNA/efeitos dos fármacos , Células Epiteliais/metabolismo , Esôfago/citologia , Esôfago/metabolismo , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteína 1 Homóloga a MutL/genética , Proteína Supressora de Tumor p53/genética , Proteínas Quinases p38 Ativadas por Mitógeno/genéticaRESUMO
BACKGROUND/AIMS: Adult cardiomyocytes can re-enter cell cycle as stimulated by prohypertrophic factors although they withdraw from cell cycle soon after birth. p21WAF1/CIP1, a cyclin-dependent kinase inhibitor, has been implicated in cardiac hypertrophy, however, its precise contribution to this process remains largely unclear. METHODS: The gene expression profile in left ventricle (LV) of spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats was determined using quantitative PCR array and verified by real-time PCR and Western blotting. Hypertrophic response of H9c2 cells and neonatal rat ventricular myocytes (NRVM) were induced by angiotensin II (1 µmol/L). Cardiac hypertrophy of mice was elicited by isoproterenol (ISO) infusion (40 mg/kg per day for 14 days). p21-adenovirus and p21-siRNA were employed to transfect NRVM, and sterigmatocystin (STE, 3 mg/kg, ip, qd) was used to inhibit p21 activity. mRNA and protein expression levels of α- and ß-myosin heavy chain (MHC), p21WAF1/CIP1, calcineurin (CaN) and atrial natriuretic peptide (ANP) were assayed by realtime PCR and WB, respectively. RESULTS: Sixteen genes showed two-fold or greater changes between SHR and WKY rats, in which the expression of p21WAF1/CIP1 was upregulated by 4.15-fold (P=0.002) and reversed by losartan. Surface area, protein content, mRNA and protein expressions of ß-MHC, ANP and p21WAF1/CIP1 in H9c2 cells treated with AngII elevated significantly compared with control group. p21-Ad transfection markedly increased the surface area and ß-MHC mRNA expression of normal NRVMs, and p21-siRNA transfection decreased them in AngII-treated NRVMs. STE treatment decreased HW/BW and cross-sectional area, expression levels of ß-MHC, ANP and p21 significantly in ISO-treated mice. CONCLUSION: Our findings suggest that p21 facilitates the development of cardiac hypertrophy, and regulating the expression of p21 may be an approach to attenuate hypertrophic growth of cardiomyocytes.
Assuntos
Cardiomegalia/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Ventrículos do Coração/metabolismo , Miócitos Cardíacos/metabolismo , Animais , Fator Natriurético Atrial/genética , Fator Natriurético Atrial/metabolismo , Calcineurina/genética , Calcineurina/metabolismo , Cardiomegalia/induzido quimicamente , Cardiomegalia/metabolismo , Cardiomegalia/patologia , Linhagem Celular , Inibidor de Quinase Dependente de Ciclina p21/antagonistas & inibidores , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Ventrículos do Coração/efeitos dos fármacos , Ventrículos do Coração/patologia , Isoproterenol , Losartan/farmacologia , Masculino , Camundongos Endogâmicos C57BL , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Esterigmatocistina/farmacologia , Miosinas Ventriculares/genética , Miosinas Ventriculares/metabolismoRESUMO
A method is reported for the analysis of sterigmatocystin in various food and feed matrices using a commercial sterigmatocystin immunoaffinity column (IAC) for sample clean-up prior to HPLC analysis by UV with mass spectrometric detection (LC-MS/MS). Cereals (wheat, oats, rye, maize and rice), sunflower seeds and animal feed were spiked with sterigmatocystin at levels from 0.75 to 50 µg kg(-1) to establish method performance. Using acetonitrile/water extraction followed by IAC clean-up, and analysis by HPLC with detection at 325 nm, recoveries ranged from 68% to 106%, with repeatability from 4.2% to 17.5%. The limit of quantification with UV detection in these matrices was 1.5 µg kg(-1). For the analysis of beer and cheese the sample preparation prior to IAC clean-up was changed to accommodate the different properties of the matrix, prior to analysis by LC-MS/MS. For beer and cheese spiked at 5.0 µg kg(-1) the recoveries were 94% and 104%, and precision (RSDs) were 1.9% and 2.9% respectively. The limits of quantification by LC-MS/MS in beer and cheese were 0.02 and 0.6 µg kg(-1) respectively. The sterigmatocystin IAC was demonstrated to provide an efficient clean-up of various matrices to enable this mycotoxin to be determined by either HPLC with UV detection or LC-MS/MS.
Assuntos
Ração Animal/análise , Cerveja/análise , Queijo/análise , Grão Comestível/química , Micotoxinas/isolamento & purificação , Esterigmatocistina/isolamento & purificação , Animais , Cromatografia de Afinidade , Cromatografia Líquida de Alta Pressão , Grão Comestível/efeitos dos fármacos , Contaminação de Alimentos/análise , Humanos , Limite de Detecção , Micotoxinas/farmacologia , Reprodutibilidade dos Testes , Esterigmatocistina/farmacologia , Espectrometria de Massas em TandemRESUMO
UNLABELLED: In screening for natural products with antiparasitic activity, an endophytic fungus, strain F2611, isolated from above-ground tissue of the tropical grass Paspalum conjugatum (Poaceae) in Panama, was chosen for bioactive principle elucidation. Cultivation on malt extract agar (MEA) followed by bioassay-guided chromatographic fractionation of the extract led to the isolation of the new polyketide integrasone B (1) and two known mycotoxins, sterigmatocystin (2) and secosterigmatocystin (3). Sterigmatocystin (2) was found to be the main antiparasitic compound in the fermentation extract of this fungus, possessing potent and selective antiparasitic activity against Trypanosoma cruzi, the cause of Chagas disease, with an IC50 value of 0.13 µmol l(-1) . Compounds 2 and 3 showed high cytotoxicity against Vero cells (IC50 of 0.06 and 0.97 µmol l(-1) , respectively). The new natural product integrasone B (1), which was co-purified from the active fractions, constitutes the second report of a natural product possessing an epoxyquinone with a lactone ring and exhibited no significant biological activity. Strain F2611 represents a previously undescribed taxon within the Microthyriaceae (Dothideomycetes, Ascomycota). SIGNIFICANCE AND IMPACT OF THE STUDY: The present study attributes new antiparasitic and psychoactive biological activities to sterigmatocystin (2), and describes the structure elucidation of the new natural product integrasone B (1), which possesses a rare epoxyquinone with a lactone ring moiety. This is also the first report of sterigmatocystin (2) isolation in a fungal strain from this family, broadening the taxonomic range of sterigmatocystin-producing fungi. The study also presents taxonomic analyses indicating that strain F2611 is strongly supported as a member of the Microthyriaceae (Ascomycota), but is not a member of any previously known or sequenced genus.
Assuntos
Ascomicetos/química , Policetídeos/farmacologia , Esterigmatocistina/análogos & derivados , Esterigmatocistina/farmacologia , Tripanossomicidas/farmacologia , Animais , Ascomicetos/classificação , Ascomicetos/genética , Sobrevivência Celular/efeitos dos fármacos , Chlorocebus aethiops , DNA Fúngico/genética , DNA Espaçador Ribossômico/genética , Endófitos/química , Concentração Inibidora 50 , Dados de Sequência Molecular , Tipagem Molecular , Técnicas de Tipagem Micológica , Poaceae/microbiologia , Policetídeos/isolamento & purificação , Esterigmatocistina/isolamento & purificação , Tripanossomicidas/isolamento & purificação , Trypanosoma cruzi/efeitos dos fármacos , Células VeroRESUMO
Sterigmatocystin (ST) is a carcinogenic mycotoxin that is commonly found in human food, animal feed and in the indoor environment. Although the correlation between ST exposure and lung cancer has been widely reported in many studies, the cytotoxicity of ST on human pulmonary cells is not yet fully understood. In the current study, we found that ST could induce DNA double-strand breaks in a human immortalized bronchial epithelial cell line (BEAS-2B cells) and a human lung cancer cell line (A549 cells). In addition, the effects of ST on cell cycle arrest were complex and dependent on the tested ST concentration and cell type. Low concentrations of ST arrested cells in the G2/M phase in BEAS-2B cells and in the S phase in A549 cells, while at high concentration both cells lines were arrested in S and G2/M phases. Furthermore, we observed that the modulation of cyclins and CDK expression showed concomitant changes with cell cycle arrest upon ST exposure in BEAS-2B and A549 cells. In conclusion, ST induced DNA damage and affected key proteins involved in cell cycle regulation to trigger genomic instability, which may be a potential mechanism underlying the developmental basis of lung carcinogenesis.
Assuntos
Ciclo Celular/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Esterigmatocistina/farmacologia , Western Blotting , Linhagem Celular Tumoral , Ensaio Cometa , Dano ao DNA , Citometria de Fluxo , Humanos , Técnicas In Vitro , Pulmão/citologiaRESUMO
During the systematic screening of active compounds from marine-derived fungi, the extract of a strain of Aspergillus versicolor MF359 isolated from a marine sponge of Hymeniacidon perleve was identified for detailed chemical investigation. Three new secondary metabolites, named hemiacetal sterigmatocystin (1), acyl-hemiacetal sterigmatocystin (2), and 5-methoxydihydrosterigmatocystin (3), together with a known compound, aversin (4), were characterized. 1 represents a first structure of sterigmatocystin hemiacetal from nature. The antibacterial activities of these identified compounds were evaluated against Staphylococcus aureus, methicillin-resistant Staphylococcus aureus, Bacillus subtilis, and Pseudomonas aeruginosa. Compound 3 showed activity against S. aureus and B. subtilis with MIC values of 12.5 and 3.125 µg/mL, respectively.
Assuntos
Antibacterianos/isolamento & purificação , Aspergillus/química , Aspergillus/classificação , Esterigmatocistina/isolamento & purificação , Animais , Antibacterianos/química , Antibacterianos/farmacologia , Organismos Aquáticos/química , Organismos Aquáticos/classificação , Organismos Aquáticos/genética , Aspergillus/genética , Aspergillus/isolamento & purificação , Bacillus subtilis/efeitos dos fármacos , DNA Fúngico/química , DNA Fúngico/genética , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Poríferos/microbiologia , Pseudomonas aeruginosa/efeitos dos fármacos , Análise de Sequência de DNA , Staphylococcus aureus/efeitos dos fármacos , Esterigmatocistina/química , Esterigmatocistina/farmacologiaRESUMO
Cell migration is fundamental to the inflammatory response, but uncontrolled cell migration and excess recruitment of neutrophils and other leukocytes can cause damage to the tissue. Here we describe the use of an in vivo model - the Tg(mpx:GFP)(i114) zebrafish line, in which neutrophils are labelled by green fluorescent protein (GFP) - to screen a natural product library for compounds that can affect neutrophil migratory behaviour. Among 1040 fungal extracts screened, two were found to inhibit neutrophil migration completely. Subfractionation of these extracts identified sterigmatocystin and antibiotic PF1052 as the active components. Using the EZ-TAXIScan chemotaxis assay, both compounds were also found to have a dosage-dependent inhibitory effect on murine neutrophil migration. Furthermore, neutrophils treated with PF1052 failed to form pseudopods and appeared round in shape, suggesting a defect in PI3-kinase (PI3K) signalling. We generated a transgenic neutrophil-specific PtdIns(3,4,5)P3 (PIP3) reporter zebrafish line, which revealed that PF1052 does not affect the activation of PI3K at the plasma membrane. In human neutrophils, PF1052 neither induced apoptosis nor blocked AKT phosphorylation. In conclusion, we have identified an antibiotic from a natural product library with potent anti-inflammatory properties, and have established the utility of the mpx:GFP transgenic zebrafish for high-throughput in vivo screens for novel inhibitors of neutrophil migration.
Assuntos
Antibacterianos/farmacologia , Quimiotaxia/efeitos dos fármacos , Naftalenos/farmacologia , Neutrófilos/citologia , Pirrolidinonas/farmacologia , Esterigmatocistina/farmacologia , Animais , Animais Geneticamente Modificados , Apoptose , Membrana Celular/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Humanos , Inflamação , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Infiltração de Neutrófilos , Neutrófilos/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Frações Subcelulares , Transgenes , Peixe-ZebraRESUMO
Sterigmatocystin (ST), which is commonly detected in food and feed commodities, is a mutagenic and carcinogenic mycotoxin that has been recognized as a possible human carcinogen. Our previous study showed that ST can induce G2 phase arrest in GES-1 cells in vitro and that the MAPK and PI3K signaling pathways are involved in the ST-induced G2 arrest. It is now widely accepted that DNA damage plays a critical role in the regulation of cell cycle arrest and apoptosis. In response to DNA damage, a complex signaling network is activated in eukaryotic cells to trigger cell cycle arrest and facilitate DNA repair. To further explore the molecular mechanism through which ST induces G2 arrest, the current study was designed to precisely dissect the role of DNA damage and the DNA damage sensor ataxia telangiectasia-mutated (ATM)/p53-dependent pathway in the ST-induced G2 arrest in GES-1 cells. Using the comet assay, we determined that ST induces DNA damage, as evidenced by the formation of DNA comet tails, in GES-1 cells. We also found that ST induces the activation of ATM and its downstream molecules, Chk2 and p53, in GES-1 cells. The ATM pharmacological inhibitor caffeine was found to effectively inhibit the activation of the ATM-dependent pathways and to rescue the ST-induced G2 arrest in GES-1 cells, which indicating its ATM-dependent characteristic. Moreover, the silencing of the p53 expression with siRNA effectively attenuated the ST-induced G2 arrest in GES-1 cells. We also found that ST induces apoptosis in GES-1 cells. Thus, our results show that the ST-induced DNA damage activates the ATM/53-dependent signaling pathway, which contributes to the induction of G2 arrest in GES-1 cells.
Assuntos
Dano ao DNA , Células Epiteliais/citologia , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Esterigmatocistina/farmacologia , Estômago/citologia , Proteína Supressora de Tumor p53/metabolismo , Apoptose/efeitos dos fármacos , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Cafeína/farmacologia , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Quinase do Ponto de Checagem 2/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/enzimologia , Inativação Gênica/efeitos dos fármacos , Humanos , Modelos Biológicos , RNA Interferente Pequeno/metabolismo , Fatores de TempoRESUMO
Sterigmatocystin (ST), a mycotoxin with mutagenic, cytotoxic and carcinogenic properties, is commonly found as the contaminant in grains and animal feeds. Of particular interest is the capacity of ST to alter normal immune function when presented in foods. As part of an on-going investigation of ST toxicological effects, we attempt to explore the short-term immunotoxic effects of ST, specifically on FoxP3(+) regulatory T cells (FoxP3(+) Tregs) and plasmacytoid dendritic cells (pDCs), by observing changes in number/expression of FoxP3(+) Tregs, pDCs and CD4(+), CD8(+) T cells in BALB/c mice 24h after a single intraperitoneal administration of ST at different dosages (3, 30, 300 and 3000 µg/kg body weight). The present study showed that 24h after ST treatment, the proportion of CD8(+) T cells was decreased in the thymus in ST 3 µg/kg group, while that of CD4(+) and CD8(+) T cells was increased in the spleen in two treatment groups (3 and 30 µg/kg). The proportion of FoxP3(+) Tregs and FoxP3 expressions were all significantly increased in mPBMCs, the thymus and the spleen. It is noteworthy that the population of pDCs significantly decreased in the thymus as we expected but increased in the spleen as compared with control, which we suspect is resulted from a temporary immune response triggered by the ST inhibition. We believe that ST may exert its immunotoxic effects by stimulating Treg, but inhibiting pDCs in the long-term to contribute its carcinogenic effects.
Assuntos
Carcinógenos/farmacologia , Células Dendríticas/efeitos dos fármacos , Fatores de Transcrição Forkhead/fisiologia , Esterigmatocistina/farmacologia , Linfócitos T Reguladores/efeitos dos fármacos , Animais , Western Blotting , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/efeitos dos fármacos , Contagem de Células , DNA Complementar/genética , DNA Complementar/isolamento & purificação , Citometria de Fluxo , Fatores de Transcrição Forkhead/genética , Imuno-Histoquímica , Subunidade alfa de Receptor de Interleucina-3/metabolismo , Contagem de Linfócitos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Reação em Cadeia da Polimerase , RNA/genética , RNA/isolamento & purificação , Baço/citologia , Timo/citologiaRESUMO
Sterigmatocystin (ST) is a toxic metabolite mainly produced by the fungi Aspergillus nidulans and Aspergillus versicolor. ST is considered a potent carcinogen, mutagen and teratogen. However, over the past few years, it has been demonstrated that it is less acutely toxic to rodents in vivo. In this study, we evaluated the putative effects of ST on the expression of tumor necrosis factor-α (TNF-α), interleukin (IL)-6 and IL-12 at mRNA levels in murine peripheral blood mononuclear cells (mPBMCs) and peritoneal macrophage cells and on the serum TNF-α and IL-6 levels in BALB/c mice. Our results show the downregulation of TNF-α, IL-6 and IL-12 mRNA expression in mPBMCs and peritoneal macrophage cells using semi-quantitative reverse transcription-polymerase chain reaction following ST treatment by intraperitoneal injection. Additionally, serum TNF-α and IL-6 levels were also decreased as shown by enzyme-linked immunosorbent assay (ELISA). These results suggest that ST contamination has negative immunomodulatory effects through the downregulation of cytokine expression and secretion.
Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Fatores Imunológicos/farmacologia , Interleucina-12/biossíntese , Interleucina-6/biossíntese , Leucócitos Mononucleares/metabolismo , Macrófagos Peritoneais/metabolismo , Esterigmatocistina/farmacologia , Fator de Necrose Tumoral alfa/biossíntese , Animais , Masculino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
In a screening for natural products with mosquito larvicidal activities, the endophytic fungus Podospora sp. isolated from the plant Laggera alata (Asteraceae) was conspicuous. Two xanthones, sterigmatocystin (1) and secosterigmatocystin (2), and an anthraquinone derivative (3) 13-hydroxyversicolorin B were isolated after fermentation on M(2) medium. These compounds were characterised using spectroscopic and X-ray analysis and examined against third instar larvae of Anopheles gambiae. The results demonstrated that compound 1 was the most potent one with LC(50) and LC(90) values of 13.3 and 73.5 ppm, respectively. Over 95% mortality was observed at a concentration 100 ppm after 24 h. These results compared farvorably with the commercial larvicide pylarvex® that showed 100% mortality at the same concentration. Compound 3 was less potent and had an LC(50) of 294.5 ppm and over 95% mortality was achieved at a concentration of 1,000 ppm. Secosterigmatocystin (2) revealed relatively weak activity and therefore LC values were not determined.
Assuntos
Anopheles/efeitos dos fármacos , Insetos Vetores/efeitos dos fármacos , Inseticidas , Podospora/metabolismo , Animais , Antraquinonas/análise , Antraquinonas/química , Antraquinonas/isolamento & purificação , Antraquinonas/farmacologia , Descoberta de Drogas , Inseticidas/química , Inseticidas/isolamento & purificação , Larva/efeitos dos fármacos , Dose Letal Mediana , Espectroscopia de Ressonância Magnética , Esterigmatocistina/química , Esterigmatocistina/isolamento & purificação , Esterigmatocistina/farmacologia , Xantonas/química , Xantonas/farmacologiaRESUMO
Five known fungal metabolites, aurasperone A, aurasperone D, averufanin, flavasperone and sterigmatocystin, were isolated from the culture broths of Aspergillus species as inhibitors of acyl-CoA:cholesterol acyltransferase (ACAT) in the cell-based assay using ACAT1- and ACAT2-expressing CHO cells. These compounds share a similar polycyclic skeleton. Among them, flavasperone and sterigmatocystin, having an angular skeleton, showed selective inhibition toward ACAT2 isozyme, while the others having a linear one had no selectivity in inhibition.
Assuntos
Aspergillus/metabolismo , Cromonas/farmacologia , Inibidores Enzimáticos/farmacologia , Esterigmatocistina/farmacologia , Esterol O-Aciltransferase/antagonistas & inibidores , Animais , Células CHO , Cromonas/química , Cricetinae , Cricetulus , Esterigmatocistina/química , Esterol O-Aciltransferase 2RESUMO
Seven new aroyl uridine derivatives (kipukasins A-G; 1-7) were isolated from solid-substrate fermentation cultures of two different Hawaiian isolates of Aspergillus versicolor. The structures of compounds 1-7 were determined by analysis of NMR and MS data. The nucleoside portion of lead compound 1 was assigned as uracil-1-beta-D-ribofuranoside by spectral comparison with an authentic standard. The bioactivity of the original A. versicolor extracts was accounted for mainly by the presence of the known metabolite sterigmatocystin, but kipukasins A and B showed modest activity in assays against Gram-positive bacteria.
Assuntos
Antibacterianos/isolamento & purificação , Aspergillus/química , Micotoxinas/isolamento & purificação , Nucleosídeos/isolamento & purificação , Uridina/análogos & derivados , Antibacterianos/química , Antibacterianos/farmacologia , Bactérias Gram-Positivas/efeitos dos fármacos , Havaí , Testes de Sensibilidade Microbiana , Micotoxinas/química , Micotoxinas/farmacologia , Nucleosídeos/química , Nucleosídeos/farmacologia , Esterigmatocistina/isolamento & purificação , Esterigmatocistina/farmacologia , Uridina/química , Uridina/isolamento & purificação , Uridina/farmacologiaRESUMO
OBJECTIVE: To explore the effects of Sterigmatocystin (ST), Deoxynivalenol (DON) and Aflatoxin G1 (AFG1) on apoptosis of human peripheral blood lymphocytes (HPBLs) in vitro and thus to further elucidate the putative roles of these three mycotoxins on human immunosystem. METHODS: The effects of ST, DON and AFG1 on apoptosis of HPBLs were studied with cell culture, flow cytometric (FCM) DNA analysis and DNA agarose gel electrophoresis. RESULTS: DNA agarose gel electrophoresis results showed the characteristic "ladder" pattern of apoptosis in HPBLs treated with ST, DON and AFG1. Flow cytometric DNA analysis revealed that typical subdiploid peaks of apoptosis in DNA histogram could be seen in all groups treated with the three mycotoxins. Significant time-effect and dose-effect relationships were found between the apoptosis rates and treatment time as well as concentrations of the three mycotoxins. CONCLUSION: ST, DON and AFG1 can induce apoptosis of HPBLs in vitro and may have some negative effects on human immunosystem.