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1.
Naturwissenschaften ; 106(9-10): 51, 2019 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-31455975

RESUMO

Endophytic actinomycetes, a prolific source of natural products, are well known for their diverse metabolic versatility, and their association with medicinal plants and antimicrobial potential are well worth exploring. We isolated and identified the Streptomyces cavourensis strain MH16 inhabiting the tree Millingtonia hortensis Linn. using phylogenetic analysis based on a 16S rRNA molecular approach. We used the disc diffusion method to evaluate the impact of differences in the compositions of the media on the production of secondary metabolites from strain MH16. The production of antimicrobial metabolites was determined by the observation of inhibition zones on intensive bands when using a TLC-bioautography assay. Biosynthesis of secondary metabolites was optimal when the strain MH16 was cultured in ISP-2 medium as depicted by a zone of inhibition. Strain MH16 effectively inhibited methicillin-resistant Staphylococcus aureus, Escherichia coli, Candida albicans, and other multi drug-resistant pathogens. The minimum inhibitory concentration of the antimicrobial metabolites was 25-100 µg mL-1. The study manifests the optimization and utilization of different fermentation media which best suits for increased production of the secondary metabolites from Streptomyces cavourensis. This research suggests that the antimicrobial metabolites of strain MH16 found in M. hortensis has great potential for the biodiscovery of new anti-infective drugs against a wide range of multidrug-resistant pathogens.


Assuntos
Peptídeos Catiônicos Antimicrobianos/biossíntese , Peptídeos Catiônicos Antimicrobianos/farmacologia , Resistência Microbiana a Medicamentos/efeitos dos fármacos , Streptomyces/genética , Streptomyces/metabolismo , Bactérias/efeitos dos fármacos , Meios de Cultura/farmacologia , Fungos/efeitos dos fármacos , Lamiales/microbiologia , Testes de Sensibilidade Microbiana , RNA Ribossômico 16S/genética , Streptomyces/efeitos dos fármacos , Streptomyces/crescimento & desenvolvimento
2.
Int J Food Microbiol ; 306: 108273, 2019 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-31382055

RESUMO

Ochratoxin A (OTA) is a nephrotoxic mycotoxin naturally found in a wide range of food commodities throughout the world. Aspergillus carbonarius is the most important source of OTA in food commodities such as wine, grapes and dried vine fruits and is also responsible for the formation of OTA in coffee. The aim of this study was to determine the simultaneous effect of three culture media (Czapek Yeast Extract Broth (CYB); Synthetic Grape Juice Medium (SGM) and White grape juice (WGJ)) at three water activity (aw) levels (0.90; 0.95 and 0.98-0.99), and three incubation temperatures (15 °C, 25 °C and 35 °C) on the growth and OTA production by 16 strains of A. carbonarius. The strains were mainly isolated from grapes from areas with a Mediterranean climate. All the strains were confirmed for identity by sequencing of the calmodulin gene. The assay was performed in microtiter plates, determining the absorbance at 530 nm and the concentration of OTA after 1, 2, 4 and 10 days of incubation. No significant differences were observed in absorbance values between the strains. The highest absorbance values were recorded in CYB at 0.99 aw and at 0.95 aw after 10 days of incubation at 25 °C and 35 °C. None of the strains were able to grow at 0.90 aw and 15 °C in any culture media after 10 days of incubation. OTA concentration was statistically higher at 15 °C than at 25 °C or 35 °C. The highest significant OTA values were obtained at 0.98-0.99 aw and the best culture media for OTA production was CYB, followed by WGJ and SGM. While strains isolated from Mediterranean climate foods had a similar behavior despite being isolated from different geographical areas, OTA concentration produced by one Robusta coffee strain from Thailand was statistically higher at 25 °C than at 15 °C. This would suggest that the type of food matrices and consequently the adaptation of A. carbonarius strains to different climatic conditions would have a greater influence on the ecophysiology of the strains than only their geographical origin.


Assuntos
Aspergillus/crescimento & desenvolvimento , Aspergillus/metabolismo , Meios de Cultura/farmacologia , Micotoxinas/biossíntese , Ocratoxinas/biossíntese , Aspergillus/patogenicidade , Clima , Meio Ambiente , Microbiologia de Alimentos , Temperatura Ambiente , Tailândia , Vitis/microbiologia , Água/análise , Vinho/microbiologia
3.
Bioengineered ; 10(1): 335-344, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31322471

RESUMO

Selenium-enriched yeast can transform toxic inorganic selenium into absorbable organic selenium, which is of great significance for human health and pharmaceutical industry. A yeast Rhodotorula glutinis X-20 we obtained before has good selenium-enriched ability, but its selenium content is still low for industrial application. In this study, strategies of process optimization and transport regulation of selenium were thus employed to further improve the cell growth and selenium enrichment. Through engineering phosphate transporters from Saccharomyces cerevisiae into R. glutinis X-20, the selenium content was increased by 21.1%. Through using mixed carbon culture (20 g L-1, glycerol: glucose 3:7), both biomass and selenium content were finally increased to 5.3 g L-1 and 5349.6 µg g-1 (cell dry weight, DWC), which were 1.14 folds and 6.77 folds compared to their original values, respectively. Our results indicate that high selenium-enrichment ability and biomass production can be achieved through combining process optimization and regulation of selenium transport.


Assuntos
Engenharia Metabólica/métodos , Fosfatos/metabolismo , Rhodotorula/genética , Saccharomyces cerevisiae/genética , Selênio/metabolismo , Transgenes , Transporte Biológico , Biomassa , Meios de Cultura/química , Meios de Cultura/farmacologia , Fermentação , Expressão Gênica , Glucose/química , Glucose/metabolismo , Glicerol/química , Glicerol/metabolismo , Proteínas de Transporte de Fosfato/genética , Proteínas de Transporte de Fosfato/metabolismo , Plasmídeos/química , Plasmídeos/metabolismo , Simportadores de Próton-Fosfato/genética , Simportadores de Próton-Fosfato/metabolismo , Rhodotorula/metabolismo , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas Cotransportadoras de Sódio-Fosfato Tipo III/genética , Proteínas Cotransportadoras de Sódio-Fosfato Tipo III/metabolismo
4.
Int J Food Microbiol ; 306: 108258, 2019 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-31362161

RESUMO

Propionic acid is widely used as a preservative in (poultry) feed. In this study we have isolated and identified fungal strains from nine samples poultry feed originating from different countries. The majority of the strains were Aspergilli with a eurotium-morph, such as Aspergillus proliferans and A. chevalieri. These and three other species were selected and tested for their sensitivity towards the feed preservative propionic acid, among them Penicillium lanosocoeruleum. The determined MIC values of 6.1-31 mM of these poultry feed specific fungi were well in the range as described in literature. Propionic acid (at 31 mM) damages conidia (spores) in a species dependent fashion after a 24-hour-treatment. The majority of the conidia (over 70%) of P. lanosocoeruleum germinated within 60 h on agar medium, while 50 and 80% of the A. chevalieri and A. proliferans conidia did not, respectively. Dependent on the species, cell damage was visible after incubation with propionic acid. Germ tubes of P. lanosocoeruleum in a biofilm showed extensive (85%) cell death after a 30 min treatment with propionic acid and slightly lower sensitivity was observed with A. proliferans (62% cell death). Microscopic analysis of these fungal biofilms revealed extensive damage to the cell membrane and showed distorted intracellular structures. Fluorescent life-dead staining of the germ tubes showed a clear dose response of propionic acid indicating a fungicidal effect on these growing cells. These results show that conidia can be inactivated by propionic acid, but that germ tubes show a much higher sensitivity. These observations shed new light on the mode of action of this important preservative to prevent fungal contamination of feed.


Assuntos
Ração Animal/microbiologia , Aspergillus/efeitos dos fármacos , Fungicidas Industriais/farmacologia , Penicillium/efeitos dos fármacos , Propionatos/farmacologia , Esporos Fúngicos/efeitos dos fármacos , Animais , Aspergillus/classificação , Aspergillus/isolamento & purificação , Biofilmes/efeitos dos fármacos , Meios de Cultura/farmacologia , Eurotium , Microbiologia de Alimentos/métodos , Testes de Sensibilidade Microbiana , Penicillium/classificação , Penicillium/isolamento & purificação , Aves Domésticas
5.
Nature ; 571(7763): 117-121, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31142833

RESUMO

Multipotent self-renewing haematopoietic stem cells (HSCs) regenerate the adult blood system after transplantation1, which is a curative therapy for numerous diseases including immunodeficiencies and leukaemias2. Although substantial effort has been applied to identifying HSC maintenance factors through the characterization of the in vivo bone-marrow HSC microenvironment or niche3-5, stable ex vivo HSC expansion has previously been unattainable6,7. Here we describe the development of a defined, albumin-free culture system that supports the long-term ex vivo expansion of functional mouse HSCs. We used a systematic optimization approach, and found that high levels of thrombopoietin synergize with low levels of stem-cell factor and fibronectin to sustain HSC self-renewal. Serum albumin has long been recognized as a major source of biological contaminants in HSC cultures8; we identify polyvinyl alcohol as a functionally superior replacement for serum albumin that is compatible with good manufacturing practice. These conditions afford between 236- and 899-fold expansions of functional HSCs over 1 month, although analysis of clonally derived cultures suggests that there is considerable heterogeneity in the self-renewal capacity of HSCs ex vivo. Using this system, HSC cultures that are derived from only 50 cells robustly engraft in recipient mice without the normal requirement for toxic pre-conditioning (for example, radiation), which may be relevant for HSC transplantation in humans. These findings therefore have important implications for both basic HSC research and clinical haematology.


Assuntos
Técnicas de Cultura de Células/métodos , Autorrenovação Celular/efeitos dos fármacos , Transplante de Células-Tronco Hematopoéticas/métodos , Células-Tronco Hematopoéticas/citologia , Animais , Proliferação de Células/efeitos dos fármacos , Células Clonais/citologia , Células Clonais/efeitos dos fármacos , Meios de Cultura/química , Meios de Cultura/farmacologia , Feminino , Fibronectinas/farmacologia , Células-Tronco Hematopoéticas/efeitos dos fármacos , Masculino , Camundongos , Álcool de Polivinil/farmacologia , Albumina Sérica , Fator de Células-Tronco/farmacologia , Trombopoetina/farmacologia , Fatores de Tempo , Condicionamento Pré-Transplante
6.
BMC Res Notes ; 12(1): 250, 2019 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-31053168

RESUMO

OBJECTIVE: The objective of this experiment was to identify transcripts in baker's yeast (Saccharomyces cerevisiae) that could have originated from previously non-coding genomic regions, or de novo. We generated this data to be able to compare the transcriptomes of different species of Ascomycota. DATA DESCRIPTION: We generated high-depth RNA sequencing data for 11 species of yeast: Saccharomyces cerevisiae, Saccharomyces paradoxus, Saccharomyces mikatae, Saccharomyces kudriavzevii, Saccharomyces bayanus, Naumovia castelii, Kluyveromyces lactis, Lachancea waltii, Lachancea thermotolerans, Lachancea kluyveri, and Schizosaccharomyces pombe. Using RNA-Seq from yeast grown in rich and oxidative conditions we created genome-guided de novo assemblies of the transcriptomes for each species. We included synthetic spike-in transcripts in each sample to determine the lower limit of detection of the sequencing platform as well as the reliability of our de novo transcriptome assembly pipeline. We subsequently compared the de novo transcripts assemblies to the reference gene annotations and generated assemblies that comprised both annotated and novel transcripts.


Assuntos
Meios de Cultura/farmacologia , Estresse Oxidativo/genética , Transcriptoma/genética , Leveduras/crescimento & desenvolvimento , Leveduras/genética , Estresse Oxidativo/efeitos dos fármacos , Transcriptoma/efeitos dos fármacos , Leveduras/efeitos dos fármacos
7.
J Appl Microbiol ; 127(1): 109-120, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31067345

RESUMO

AIMS: To determine how the microbicide ceragenin-13 (CSA-13) kills Bacillus subtilis spores prepared on growth or sporulation media, and these spores' properties. METHODS AND RESULTS: Spores made on Luria broth (LB) growth or double-strength Schaeffer's-glucose (2xSG) sporulation plates found that spores made on LB plates have coat defects as evidenced by their lower hypochlorite resistance, faster germination with dodecylamine and slower germination with Ca2+ -dipicolinic acid (CaDPA) than 2xSG plate spores. CSA-13 triggered CaDPA release from spores, an early step in germination, but only well at 70°C and better with spores made on LB than on 2xSG plates. Approximately 90% of spores with elevated levels of SpoVA proteins that form a CaDPA release channel, released CaDPA with CSA-13 at 70°C, and faster with spores made on LB than 2xSG plates. Levels of CSA-13 killing of spores made on LB and 2xSG plates were similar to levels of CaDPA release triggered by this agent. CONCLUSIONS: CSA-13 kills bacterial spores, but only at high concentrations and temperatures, and is preceded by CaDPA release. SIGNIFICANCE AND IMPACT OF THE STUDY: CSA-13 is not a direct sporicide as reported previously, but most likely germinates spores via activation of spores' CaDPA channel, albeit inefficiently, and then killing the germinated spores.


Assuntos
Anti-Infecciosos/farmacologia , Bacillus subtilis/efeitos dos fármacos , Meios de Cultura/química , Meios de Cultura/farmacologia , Esporos Bacterianos/efeitos dos fármacos , Esporos Bacterianos/crescimento & desenvolvimento , Esteroides/farmacologia , Aminas , Ácidos Picolínicos/metabolismo , Esporos Bacterianos/metabolismo
8.
Environ Sci Pollut Res Int ; 26(20): 20815-20828, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31111387

RESUMO

The effects of iron (Fe), zinc (Zn), and molybdenum (Mo) on the biomass yield, lipid content, lipid yield, and fatty acid composition of Chlorella sp. NC-MKM, Graesiella emersonii NC-M1, Scenedesmus acutus NC-M2, and Chlorophyta sp. NC-M5 were studied. Among them, G. emersonii NC-M1 recorded the highest percentage increase in lipid content (140.3%) and neutral lipid (50.9%) under Zn-supplemented condition compared to the control. Also, it showed a 105% and 41.88% increase in lipid yield and neutral lipid under Fe-supplemented condition compared to the control. However, Chlorella sp. NC-MKM recorded an elevation in lipid yield (70.3% rise) and neutral lipid (24.32% rise) compared to the control in Mo-supplemented condition. The enhanced production of reactive oxygen species (ROS) and antioxidant enzyme (SOD and POD) under Fe-, Zn-, and Mo-supplemented condition supports the lipid accumulation. FAME analysis showed that the overall percentage of SFA and MUFA increased after the addition of Fe, Zn, and Mo in a culture medium compared to the control which is vital for a good-quality biodiesel. Further, biodiesel properties derived from FAMEs such as CN, SV, IV, CFPP, OS, υ, ρ, and HHV were found in accordance with biodiesel standard.


Assuntos
Ferro/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Microalgas/efeitos dos fármacos , Molibdênio/farmacologia , Zinco/farmacologia , Biocombustíveis , Chlorella/efeitos dos fármacos , Chlorella/metabolismo , Clorófitas/efeitos dos fármacos , Clorófitas/metabolismo , Meios de Cultura/química , Meios de Cultura/farmacologia , Ácidos Graxos/análise , Lipídeos/química , Microalgas/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Scenedesmus/efeitos dos fármacos , Scenedesmus/metabolismo
9.
Malar J ; 18(1): 155, 2019 May 02.
Artigo em Inglês | MEDLINE | ID: mdl-31046772

RESUMO

BACKGROUND: The protective efficacy of the most promising malaria whole-parasite based vaccine candidates critically depends on the parasite's potential to migrate in the human host. Key components of the parasite motility machinery (e.g. adhesive proteins, actin/myosin-based motor, geometrical properties) have been identified, however the regulation of this machinery is an unknown process. METHODS: In vitro microscopic live imaging of parasites in different formulations was performed and analysed, with the quantitative analysis software SMOOTIn vitro, their motility; their adherence capacity, movement pattern and velocity during forward locomotion. RESULTS: SMOOTIn vitro enabled the detailed analysis of the regulation of the motility machinery of Plasmodium berghei in response to specific (macro)molecules in the formulation. Albumin acted as an essential supplement to induce parasite attachment and movement. Glucose, salts and other whole serum components further increased the attachment rate and regulated the velocity of the movement. CONCLUSIONS: Based on the findings can be concluded that a complex interplay of albumin, glucose and certain salts and amino acids regulates parasite motility. Insights in parasite motility regulation by supplements in solution potentially provide a way to optimize the whole-parasite malaria vaccine formulation.


Assuntos
Meios de Cultura/química , Locomoção/efeitos dos fármacos , Plasmodium berghei/efeitos dos fármacos , Esporozoítos/fisiologia , Albuminas/farmacologia , Animais , Culicidae/parasitologia , Meios de Cultura/farmacologia , Feminino , Glucose/farmacologia , Microscopia Intravital , Malária/parasitologia , Camundongos , Plasmodium berghei/fisiologia , Software
10.
Comp Immunol Microbiol Infect Dis ; 63: 131-135, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30961808

RESUMO

The aim of the study was to determine whether the presence of the Yersinia virulence plasmid could affect the production of enterotoxin YstA by Y. enterocolitica strains isolated from pigs which are the main source of infection for humans. The phenotypic features characteristic for the Yersinia virulence plasmid were detected on CRMOX agar in 8 out of 12 strains producing enterotoxin YstA, in 5 out of 12 doubtful strains, and in 11 out of 12 strains not producing YstA. Autoagglutination ability was detected in all 12 Y. enterocolitica strains that were positive in the suckling mice bioassay, in 11 doubtful strains and 10 negative strains. CRMOX+ colonies were generally ystA, myfA, virF and yadA positive, while CRMOX- colonies were only ystA and myfA positive. The amplicons of yadA were not detected in 2 (8.3%) out of 24 CRMOX+ and virF positive strains. The results of this study indicate that the presence of pYV does not affect the enterotoxin-producing ability of Y. enterocolitica strains.


Assuntos
Toxinas Bacterianas/biossíntese , Enterotoxinas/biossíntese , Plasmídeos/genética , Yersinia enterocolitica/genética , Yersinia enterocolitica/metabolismo , Adesinas Bacterianas/metabolismo , Animais , Antígenos de Bactérias/metabolismo , Proteínas da Membrana Bacteriana Externa/metabolismo , Meios de Cultura/farmacologia , DNA Bacteriano/genética , Humanos , Camundongos , Suínos , Doenças dos Suínos/microbiologia , Yersinia enterocolitica/patogenicidade
11.
Mater Sci Eng C Mater Biol Appl ; 100: 862-873, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30948124

RESUMO

Mineralization capability is an important issue in developing bone repairing biomaterials, while it is not quite clear how this feature would act in the presence of cells and influence cell osteogenic differentiation without adding extra osteoinductive factors such as ß­sodium glycerophosphate and dexamethasone. Poly(l­lactide) (PLLA) and gelatin composite fibers (PG, 1:1 in weight) were electrospun, treated with CaCl2 solution (PG-Ca), and used for mineralization studies by using cell culture media (αMEM, and αMEM + serum). Bone mesenchymal stromal cells (BMSCs) were then seeded and cultured on both PG and PG-Ca fibrous mats for 28 days by only using αMEM + serum. Interestingly, mineral depositions on both PG and PG-Ca fibers were detected in the environment of αMEM or αMEM + serum, in which, PG-Ca fibers demonstrated stronger ability in inducing hydroxyapatite formation than PG fibers, especially in the presence of fetal bovine serum. When BMSCs were cultured on the two kinds of fibrous mats, apatite depositions were still clearly detected, while the depositing amounts decreased in comparison with corresponding cell-free cases. It was ascribed to the consumption of ions by the continuously proliferating BMSCs, whose osteogenic differentiation was significantly promoted even without extra osteoinductive factors, especially on PG-Ca fibrous mats, in comparison with the control group. Therefore, it was confirmed the capability of scaffolding materials in enriching ions like calcium and phosphate around cells was an efficient way to promote bone regeneration.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Meios de Cultura/farmacologia , Gelatina/farmacologia , Células-Tronco Mesenquimais/citologia , Minerais/química , Osteogênese/efeitos dos fármacos , Poliésteres/farmacologia , Fosfatase Alcalina/metabolismo , Animais , Cloreto de Cálcio/farmacologia , Bovinos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Células-Tronco Mesenquimais/efeitos dos fármacos , Ratos Sprague-Dawley , Espectroscopia de Infravermelho com Transformada de Fourier , Termogravimetria , Difração de Raios X
12.
Methods Mol Biol ; 1968: 3-10, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30929201

RESUMO

Control of Streptococcus pneumoniae is mainly achieved by the use of existing vaccines. Capsular polysaccharides are the major antigenic component and are also the main virulence factor.Capsular polysaccharides must fulfill requirements of purity, uniformity, and an accurate molecular weight to be used as vaccine antigens. Vaccine production largely relies on cultivation of the pathogen in appropriate conditions.Here we describe widely used techniques to culture S. pneumoniae based on solid or complex liquid media, which are successfully applied in the diagnosis of the pathogen and in development and production of S. pneumoniae vaccines. Furthermore, we present a new chemically defined medium that can be used at lab scale.


Assuntos
Meios de Cultura/farmacologia , Streptococcus pneumoniae/metabolismo , Antígenos de Bactérias/metabolismo , Vacinas Bacterianas/metabolismo , Polissacarídeos Bacterianos , Streptococcus pneumoniae/efeitos dos fármacos
13.
Lipids Health Dis ; 18(1): 89, 2019 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-30954075

RESUMO

BACKGROUND: Elevation of exogenous free fatty acid (FFA) level leads to insulin resistance (IR) in liver, IR is manifested by elevated hepatic glucose production. We aim to study whether inhibition of endogenous fatty acid synthesis could decrease hepatic glucose production. METHODS: Low-passage HepG2 cells derived from human liver tissue were cultured in medium supplemented with FFA to induce IR, the influences of sterol regulatory element binding protein-1c (SREBP-1c) silencing on glucose production of HepG2 cells were investigated, and genes responsible for fatty acid and glucose metabolism were detected by real-time PCR. RESULTS: Compared with HepG2 cells cultured in normal growth medium, glucose production of HepG2 cells treated by FFA was significantly increased {[(0.28 ± 0.01) vs (0.83 ± 0.02)] umol.ug- 1 protein, n = 6 wells, P < 0.01}; the mRNA expression of phosphoenolpyruvate carboxylase kinase (PEPCK) and glucose-6-phosphatase (G6PC) in HepG2 cells increased by more than 5-fold and 3-fold, respectively; the mRNA expression of fatty acid synthase (FAS) and stearoyl-CoA desaturase-1 (SCD1) increased by approximately 4-fold and 1.1-fold, respectively; the mRNA expression of carnitine palmitoyltransferase-1 (CPT-1) changed slightly. Compared with the scrambled siRNA control, glucose production of HepG2 cells treated by FFA significantly increased after SREBP-1c silencing {[(0.018 ± 0.001) vs (0.028 ± 0.002)] umol.ug- 1 protein, n = 6 wells, P < 0.01}; the mRNA expression of PEPCK and G6PC increased by approximately 1.5-fold and 5-fold, respectively, but the mRNA expression of FAS, SCD1 and CPT-1 changed slightly. CONCLUSIONS: SREBP-1c silencing further augmented glucose production of HepG2 cells treated by FFA significantly, genes responsible for fatty acid synthesis and gluconeogenesis played an important role in this process. SREBP-1c functions not only as a lipid regulator but also plays an important role in regulation of glucose metabolism.


Assuntos
Meios de Cultura/farmacologia , Ácidos Graxos não Esterificados/farmacologia , Glucose/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Carnitina O-Palmitoiltransferase/genética , Ácido Graxo Sintases/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Inativação Gênica/efeitos dos fármacos , Células Hep G2 , Humanos , Resistência à Insulina/genética , Metabolismo dos Lipídeos/efeitos dos fármacos , Lipídeos/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/antagonistas & inibidores , Triglicerídeos/metabolismo
14.
Braz J Microbiol ; 50(2): 527-532, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30850978

RESUMO

This study aimed to evaluate the effects of the solid and semisolid culture medium on the mycelial viability of A. subrufescens after 5-year cryopreservation at - 70 °C. Mycelia were grown in three types of whole or ground grains, with or without 5% glycerol addition in the substrate and/or in a cryotube. After 5 years of cryopreservation at - 70 °C, every treatment was thawed and recovered in malt extract culture medium with 15 (solid culture medium) or 5 g L-1 (semisolid culture medium) of agar. The semisolid recovery culture medium increased the mycelial viability recovery capacity of A. subrufescens cryopreserved for 5 years in grains with glycerol only in the cryotube, and specifically with medium-hard wheat grain without glycerol addition at all. Agar-based substrates such as malt extract agar, agar-ground grain, or the one with glycerol addition to the substrate were not effective to keep the mycelial viability, regardless of the recovery culture medium consistency. Hard and medium-hard endosperm wheat grains or hard endosperm rye grains with addition of glycerol as cryoprotectant only to the cryotube were effective to cryopreserve the fungus for 5 years without cryoprotectant addition in the substrate.


Assuntos
Agaricus/crescimento & desenvolvimento , Criopreservação/métodos , Crioprotetores/farmacologia , Meios de Cultura/farmacologia , Grão Comestível/microbiologia , Glicerol/farmacologia , Micélio/crescimento & desenvolvimento , Ágar/farmacologia , Sobrevivência Celular
15.
Biomed Res Int ; 2019: 8146948, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30915361

RESUMO

It was found that Bacillus sp. Y1 could secrete alkaline pectinase with suitable enzyme system for powerful and fast degumming of ramie fiber. In this study, the medium components and fermentation conditions were optimized by some statistical methods including mixture design, fractional factorial design, central composite design and response surface methodology, and single factor method for enhancing the alkaline pectinase production. The optimized conditions for pectinase production were that the culture was shaken at 34°C for 60 h in 50 mL of medium containing 10.5% (w/v) carbon source (consisting of 3.8% starch, 4.2% wheat bran, and 2.5% sucrose), 0.37% (NH4)2SO4, 0.3% MgSO4, and 0.1% Tween-80, with initial pH 8.2 and inoculation amount of 1.3 mL (with the OD600 of the seed medium about 5.77). Using the optimizing conditions, the activities of polygalacturonate lyase (PGL) and polygalacturonase (PG) in fermentation liquor were increased to 2.00-fold and 3.44-fold, respectively, and the fermentation time shortened 12 hours (from 72 h to 60 h), which showed good application potential in degumming of ramie.


Assuntos
Bacillus/enzimologia , Meios de Cultura/química , Poligalacturonase/biossíntese , Meios de Cultura/farmacologia , Poligalacturonase/química , Poligalacturonase/isolamento & purificação
16.
Zygote ; 27(2): 69-77, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30834849

RESUMO

SummaryDirect swim-up procedure is widely used to separate the motile competent spermatozoa from the antioxidant-rich semen. Subsequently, spermatozoa become more vulnerable to reactive oxygen species (ROS) due to their cytological characteristics. The effect of vitamin C, a highly concentrated antioxidant in the semen, on direct swim-up-enriched sperm population is not fully investigated. Therefore, the aim of the present study was to assess the effect of vitamin C on sperm functional properties during direct swim-up procedure. Semen samples were collected from 22 participants. Each semen sample was divided into several aliquots. The first portion was overlaid with sperm medium without ascorbic acid (0 µM AA). The second and third fractions were overlaid with sperm medium supplemented with 300 µM and 600 µM AA; respectively. After 1 h of incubation, basic sperm parameters, intracellular ROS levels, acrosome reaction, chromatin integrity, and glucose uptake were assessed. Swim-up without AA significantly increased the percentage of ROS(+) spermatozoa compared with the raw semen (P<0.01). Interestingly, swim-up with 300 µM AA did not increase the percentage of ROS(+) sperm compared with the raw semen. In parallel, the percentage of sperm with altered chromatin integrity was significantly lower in the 300 µM AA group compared with that in the raw semen (P<0.05). These findings suggest that supplementation of vitamin C to sperm medium could be beneficial for direct swim-up-derived spermatozoa.


Assuntos
Ácido Ascórbico/farmacologia , Separação Celular/métodos , Espermatozoides/fisiologia , Reação Acrossômica , Adulto , Ácido Ascórbico/administração & dosagem , Cromatina/patologia , Meios de Cultura/química , Meios de Cultura/farmacologia , Relação Dose-Resposta a Droga , Glucose/farmacocinética , Humanos , Masculino , Espécies Reativas de Oxigênio/metabolismo , Sêmen/fisiologia , Motilidade Espermática/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos
17.
Ann Clin Lab Sci ; 49(1): 63-71, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30814079

RESUMO

This study aimed to evaluate the use of Insulin-Transferrin-Selenium (ITS) medium in place of fetal bovine serum (FBS) to culture human amnion mesenchymal stem cells (hAMSCs). Cell morphology, ultrastructure, proliferation, migration and MSC related markers were assessed accordingly. The hAMSCs were induced to osteocyte, chondrocyte, adipocyte and keratinocyte by culturing in appropriate induction medium. hAMSCs mRNA expression was detected for the matrix metalloproteinases 2 (MMP2), keratinocyte growth factor (KGF), vascular endothelial growth factor (VEGF), insulin-like growth factor-I (IGF-I), Platelet-derived Growth Factor (PDGF), and transforming growth factor beta 1 (TGF-ß) by real-time quantitative RT-PCR. Our results showed that hAMSCs cultured in ITS medium exhibited similar proliferation rates, demonstrated a statistically significant increased migration and expressed similar levels of MSC markers(CD73+, CD90+, CD105+, CD45-, CD34-) compared with those cultured in FBS. Osteoblasts, chondrocytes, adipocytes and keratinocytes were differentiated. Results of transmission electron microscope (TEM) revealed that hAMSCs cultured in ITS medium underwent active metabolism. The mRNA expression of MMP2, VEGF, KGF, TGF-ß, IGF-I and PDGF upregulated in ITS medium. In conclusion, ITS medium has the potential to be used for the expansion of hAMSCs before clinical application.


Assuntos
Adipócitos/citologia , Âmnio/citologia , Condrócitos/citologia , Meios de Cultura/farmacologia , Queratinócitos/citologia , Células-Tronco Mesenquimais/citologia , Osteócitos/citologia , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Âmnio/efeitos dos fármacos , Âmnio/metabolismo , Antioxidantes/farmacologia , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Condrócitos/efeitos dos fármacos , Condrócitos/metabolismo , Humanos , Hipoglicemiantes/farmacologia , Insulina/farmacologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Osteócitos/efeitos dos fármacos , Osteócitos/metabolismo , Selênio/farmacologia , Transdução de Sinais , Transferrina/farmacologia
18.
Int J Oncol ; 54(5): 1843-1852, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30864702

RESUMO

Mesenchymal stem cells (MSCs) have been demonstrated to be involved in tumor progression and the modulation of the tumor microenvironment, partly through their secretome. Extracellular vesicles (EVs) are membranous nanovesicles secreted by multiple types of cells and have been demonstrated to mediate intercellular communication in both physiological and pathological conditions. However, numerous questions still remain regarding the underlying mechanisms and functional consequences of these interactions. The purpose of this study was to investigate the effects of human umbilical cord mesenchymal stem cell­derived EVs (hUC­MSC­EVs) on the proliferation, migration and invasion of human breast cancer cells. We successfully generated and identified hUC­MSCs and hUC­MSC­EVs which were used in this study. The results revealed that treatment of the MDA­MB­231 and MCF­7 human breast cancer cells with medium containing hUC­MSC­EVs significantly enhanced the proliferation, migration and invasion of the cells in vitro. Treatment of the cells with medium containing hUC­MSC­EVs also reduced E­cadherin expression and increased N­cadherin expression, thus promoting the epithelial­mesenchymal transition (EMT) of the breast cancer cells. Treatment of the breast cancer cells with extracellular signal­regulated kinase (ERK) inhibitor prior to the interaction with hUC­MSC­EVs significantly reversed the enhanced proliferation, migration and invasion, as well as the EMT of the breast cancer cells induced by the hUC­MSC­EVs. On the whole, these data indicate that hUC­MSC­EVs promote the invasive and migratory potential of breast cancer cells through the induction of EMT via the ERK pathway, leading to malignant tumor progression and metastasis. Taken together, the findings of this study suggest that targeting pathways to reverse EMT may lead to the development of novel therapeutic approaches with which to combat breast cancer.


Assuntos
Neoplasias da Mama/metabolismo , Meios de Cultura/farmacologia , Vesículas Extracelulares/patologia , Sistema de Sinalização das MAP Quinases , Células-Tronco Mesenquimais/citologia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Técnicas de Cocultura , Meios de Cultura/química , Transição Epitelial-Mesenquimal , Vesículas Extracelulares/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Células MCF-7 , Células-Tronco Mesenquimais/metabolismo , Microambiente Tumoral , Cordão Umbilical/citologia , Cordão Umbilical/metabolismo
19.
J Cancer Res Ther ; 15(1): 176-184, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30880776

RESUMO

Context: Tilorone dihydrochloride is a therapeutic agent with a different mechanism in cancer. The species of Lactobacillus have an important role in cytotoxic effect. Aims: Because of unknown effects of tilorone and culture supernatants from Lactobacillus reuteri on hepatoma, the aim of this study is to evaluate apoptotic, cytotoxic, and therapeutic effects of tilorone on mouse hepatoma cell line with and without culture supernatants from L. reuteri. Materials and Methods: To do so, after cell line culture, cells were divided into different groups such as negative control, treatment with four doses of tilorone, positive control of supernatant (single dose), and combination therapy groups of different doses of tilorone with supernatant (constant doses), for 48 h. All groups were studied with pathologic tests, biochemical study, tetrazolium dye (3-(4, 5- dimethylthiazol -2-yl)-2, 5-diphenyltetrazolium bromide [MTT]) assay, and absolute real-time-polymerase chain reaction (RT-PCR) were done to assess Bax and Bcl-2 genes expression, as molecular studies. Results: MTT assay results revealed that the tilorone tissue culture IC50 (TCIC50) on the Hepa1-6 cell line was 50 µg/ml. RT-PCR analysis showed that tilorone dihydrochloride induced upregulation and downregulation in expression of Bax and Bcl-2, respectively. Simultaneous, antioxidant effect has also seen in a way that prevented necrosis, in biochemical analysis. These results were dose dependent and statistically significant compared to the control group. Conclusions: Based on these results, it appeared that this agent could be a good candidate for further evaluation as effective chemotherapy acting through the induction of apoptosis in hepatoma. The cell death caused through bacterial supernatant was rather necrosis than apoptosis.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Lactobacillus reuteri/metabolismo , Neoplasias Hepáticas/tratamento farmacológico , Tilorona/farmacologia , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Apoptose/efeitos dos fármacos , Fatores Biológicos/farmacologia , Fatores Biológicos/uso terapêutico , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Meios de Cultura/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Neoplasias Hepáticas/patologia , Camundongos , Tilorona/uso terapêutico
20.
Int J Mol Med ; 43(5): 2230-2240, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30864673

RESUMO

Hair follicles (HFs) are a well­characterized niche for adult stem cells (SCs), and include epithelial and melanocytic SCs. HF cells are an accessible source of multipotent adult SCs for the generation of the interfollicular epidermis, HF structures and sebaceous glands in addition to the reconstitution of novel HFs in vivo. In the present study, it was demonstrated that HF cells are able to be induced to differentiate into cardiomyocyte­like cells in vitro under specific conditions. It was determined that HF cells cultured on OP9 feeder cells in KnockOut­Dulbecco's modified Eagle's medium/B27 in the presence of vascular endothelial growth factors differentiated into cardiomyocyte­like cells that express markers specific to cardiac lineage, but do not express non­cardiac lineage markers including neural stem/progenitor cell, HF bulge cells or undifferentiated spermatogonia markers. These cardiomyocyte­like cells exhibited a spindle­ and filament­shaped morphology similar to that presented by cardiac muscles and exhibited spontaneous beating that persisted for over 3 months. These results demonstrate that SC reprogramming and differentiation may be induced without resulting in any genetic modification, which is important for the clinical applications of SCs including tissue and organ regeneration.


Assuntos
Folículo Piloso/citologia , Miócitos Cardíacos/citologia , Animais , Biomarcadores/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem da Célula/efeitos dos fármacos , Células Cultivadas , Meios de Cultura/farmacologia , Células Alimentadoras/citologia , Células Alimentadoras/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Camundongos , Miócitos Cardíacos/efeitos dos fármacos , Fenótipo , Fator A de Crescimento do Endotélio Vascular/farmacologia
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