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1.
Ecotoxicol Environ Saf ; 181: 481-490, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31228824

RESUMO

Peroxidases and catalases are well-known antioxidant enzymes produced in almost all living organisms for the elimination of reactive oxygen species (ROS) and thus they prevent the occurrence of oxidative stress. In our study we focused on two soil fungi of the family Chaetomiaceae (mesophilic Chaetomium cochliodes and its thermophilic counterpart C. thermophilum var. dissitum) in order to explore the presence of peroxidase and catalase genes, formation of their native transcripts and protective effect of corresponding translation products in a case study. Predicted genes of our interest were confirmed by genomic PCR and their inducible transcripts by RT-PCR. We were able to quantify the expression levels of newly discovered fungal heme peroxidases and catalases with the reverse-transcription quantitative real-time PCR method. We compared obtained quantitative levels of mRNA production with the level of corresponding extracellular protein occurrence as detected with monitoring their specific peroxidase and catalase activities directly in the cultivation media at optimal growth temperatures. The presence of secretory Catalase 2 from C. thermophilum var. dissitum was detected and identified with mass spectrometry approach directly in the growth medium. This unique catalase is phylogenetically closely related with a previously described catalase-phenol oxidase thus representing an effective and versatile antioxidant in the environment of the fungal mycelia also involved in the catabolism of recalcitrant phenolic substances.


Assuntos
Ascomicetos/metabolismo , Catalase/metabolismo , Espaço Extracelular/enzimologia , Estresse Oxidativo , Peroxidases/metabolismo , Antioxidantes/metabolismo , Ascomicetos/classificação , Ascomicetos/enzimologia , Ascomicetos/genética , Catalase/genética , Meios de Cultura/metabolismo , Espaço Extracelular/metabolismo , Oxirredução , Peroxidases/genética , Filogenia , Temperatura Ambiente
2.
Inorg Chem ; 58(11): 7488-7498, 2019 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-31083932

RESUMO

It was shown that His3 of human copper transporter 1 (hCtr1) prompts the ATCUN-like Cu(II) coordination for model peptides of the hCtr1 N-terminus. Its high Cu(II) affinity is a potential driving force for the transfer of Cu(II) from extracellular Cu(II) carriers to hCtr1. Having a sequence similar to that of hCtr1, hCtr2 has been proposed as another human copper transporter. However, the N-terminal domain of hCtr2 is much shorter than that of hCtr1, with different copper binding motifs at its N-terminus. Employing a model peptide of the hCtr2 N-terminus, MAMHF-am, we demonstrated that His4 provides a unique pattern of Cu(II) complexes, involving Met sulfurs in their Cu(II) coordination sphere. The affinity of Cu(II) for MAMHF-am is a few orders of magnitude lower than that reported for the hCtr1 model peptides at the extracellular pH of 7.4, suggesting a maximal complementary role of Cu(II) binding to hCtr2 in the import of copper from the extracellular space to the cytoplasm. On the other hand, the ability of the hCtr2 model peptide to capture Cu(II) from amino acids and short peptides (potential degradation products of proteins) at pH 5.0 and the known predominant lysosomal localization of hCtr2 support an important potential role of the Cu(II)-hCtr2 interaction in the recovery of copper from lysosomes.


Assuntos
Proteínas de Transporte de Cátions/química , Cobre/metabolismo , Espaço Extracelular/química , Lisossomos/química , Fragmentos de Peptídeos/metabolismo , Sequência de Aminoácidos , Espaço Extracelular/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Lisossomos/metabolismo , Modelos Moleculares , Fragmentos de Peptídeos/química , Ligação Proteica , Conformação Proteica
3.
Adv Neurobiol ; 22: 233-250, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31073939

RESUMO

The recent years have seen unprecedented growth in the manufacturing of neurotechnological tools. The latest technological advancements presented the neuroscientific community with neuronal probes containing thousands of recording sites. These next-generation probes are capable of simultaneously recording neuronal signals from a large number of channels. Numerically, a simple 128-channel neuronal data acquisition system equipped with a 16 bits A/D converter digitizing the acquired analog waveforms at a sampling frequency of 20 kHz will generate approximately 17 GB uncompressed data per hour. Today's biggest challenge is to mine this staggering amount of data and find useful information which can later be used in decoding brain functions, diagnosing diseases, and devising treatments. To this goal, many automated processing and analysis tools have been developed and reported in the literature. A good amount of them are also available as open source for others to adapt them to individual needs. Focusing on extracellularly recorded neuronal signals in vitro, this chapter provides an overview of the popular open-source tools applicable on these signals for spike trains and local field potentials analysis, and spike sorting. Towards the end, several future research directions have also been outlined.


Assuntos
Potenciais de Ação , Eletrofisiologia/métodos , Espaço Extracelular/metabolismo , Técnicas In Vitro , Neurônios/citologia , Neurônios/metabolismo , Processamento de Sinais Assistido por Computador , Humanos , Processamento de Sinais Assistido por Computador/instrumentação
4.
J Phys Chem A ; 123(17): 3928-3934, 2019 May 02.
Artigo em Inglês | MEDLINE | ID: mdl-30957999

RESUMO

Detailed knowledge of the water status in living organisms is crucial for understanding their physiology and pathophysiology. Here, we developed a technique to spectroscopically image water at high resolution using ultrabroadband multiplex coherent anti-Stokes Raman scattering (CARS) microscopy equipped with a supercontinuum light source. This system allows for the visualization of a wide spectrum of CARS signals from the fingerprint to the end of O-H stretching at a spectral resolution of ∼10 cm-1. Application of the system to living mammalian cells revealed a spectral red shift of the O-H stretching vibrational band inside compared to outside the cells, suggesting the existence of stronger hydrogen bonds inside the cells. Furthermore, potential changes in spectra were examined by adding mannitol to the extracellular solution, which increases the osmolality outside the cells and thereby induces dehydration of the cells. Under this treatment, the red shift of the O-H stretching band was further enhanced, revealing the effects of mannitol on water states inside the cells. The methodology developed here should serve as a powerful tool for the chemical imaging of water in living cells in various biological and medical contexts.


Assuntos
Espaço Extracelular/metabolismo , Espaço Intracelular/metabolismo , Análise Espectral Raman , Água/metabolismo , Animais , Células CHO , Cricetulus , Osmose
5.
Biomed Pharmacother ; 112: 108714, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30970518

RESUMO

Changes in the expression and subcellular localization of high mobility group box 1 (HMGB1), a damage-associated molecular pattern (DAMP) molecule, have been implicated in tumorigenesis and tumor cell death in response to cancer therapy. Specifically, HMGB1 release has been shown to occur with a specific form of induced cell death known as necroptosis. In the present study, we examined the role of HMGB1 in the necroptosis of acute myeloid leukemia (AML) cells. In two AML cell lines and primary AML cells from two patients, etoposide induced necroptosis via cIAP1/2 degradation when caspase activity was inhibited by Z-VAD-fmk, but treatment with extracellular HMGB1 prevented this necroptosis. Interestingly, HMGB1 did not prevent the degradation of cIAP1/2, but rather activated the nuclear factor kappa B pathway. The results of the present study provide evidence that extracellular HMGB1 is not only an important DAMP molecule released by cells upon necrosis, but also a regulatory factor that prevents necroptosis in AML cells.


Assuntos
Alarminas/metabolismo , Apoptose , Espaço Extracelular/metabolismo , Proteína HMGB1/metabolismo , Leucemia Mieloide Aguda/patologia , Necrose , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Etoposídeo/farmacologia , Células HL-60 , Humanos , Leucemia Mieloide Aguda/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais
6.
Eur Phys J E Soft Matter ; 42(4): 47, 2019 Apr 18.
Artigo em Inglês | MEDLINE | ID: mdl-31011840

RESUMO

Drosophila wing discs show a remarkable variability when subject to mechanical perturbation. We developed a stretching bench that allows accurate measurements of instantaneous and time-dependent material behaviour of the disc as a whole, while determining the exact three-dimensional structure of the disc during stretching. Our experiments reveal force relaxation dynamics on timescales that are significant for development, along with a surprisingly nonlinear force-displacement relationship. Concurrently our imaging indicates that the disc is a highly heterogeneous tissue with a complex geometry. Using image-based 3D finite element modelling we are able to identify the contributions of size, shape and materials parameters to the measured force-displacement relations. In particular, we find that simulating the stretching of a disc with stiffness patterns in the extra-cellular matrix (ECM) recapitulates the experimentally found stretched geometries. In our simulations, linear hyperelasticity explains the measured nonlinearity to a surprising extent. To fully match the experimental force-displacement curves, we use an exponentially elastic material, which, when coupled to material relaxation also explains time-dependent experiments. Our simulations predict that as the disc develops, two counteracting effects, namely the discs foldedness and the hardening of the ECM lead to force-relative displacement curves that are nearly conserved during development.


Assuntos
Drosophila/anatomia & histologia , Elasticidade , Asas de Animais , Animais , Fenômenos Biomecânicos , Espaço Extracelular/metabolismo , Modelos Lineares , Dinâmica não Linear , Viscosidade , Asas de Animais/citologia
7.
Lab Chip ; 19(8): 1448-1457, 2019 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-30887972

RESUMO

Developing tools to enable non-invasive, high-throughput electrophysiology measurements of large functional-networks of electrogenic cells used as in vitro disease models for the heart and brain remains an outstanding challenge for preclinical drug discovery, where failures are costly and can prove to be fatal during clinical trials. Here we demonstrate, for the first time, that it is possible to perform non-contact monitoring of extra-cellular field potentials with a multi-electrode array (MEA). To do this preliminary demonstration we built a prototype with a custom mechanical stage to micro-position cells grown on conventional glass coverslips over the recording surface of a MEA sensor. The prototype can monitor extra-cellular fields generated by multi-cellular networks in a non-contact configuration, enabling a single MEA sensor to probe different cultures in succession, without fouling or degrading its sensitive electronic surface. This first demonstration with easy to culture cardiomyocyte cells and a prototype device points to the exciting possibility for instrument development leading to more efficient and cost-effective drug screening paradigms for cardiovascular and neurological diseases.


Assuntos
Espaço Extracelular/metabolismo , Microeletrodos , Avaliação Pré-Clínica de Medicamentos , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Desenho de Equipamento , Espaço Extracelular/efeitos dos fármacos , Humanos , Análise Espaço-Temporal
8.
Mol Cancer ; 18(1): 50, 2019 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-30925930

RESUMO

Increasing evidence indicates that the ability of cancer cells to convey biological information to recipient cells within the tumor microenvironment (TME) is crucial for tumor progression. Microvesicles (MVs) are heterogenous vesicles formed by budding of the cellular membrane, which are secreted in larger amounts by cancer cells than normal cells. Recently, several reports have also disclosed that MVs function as important mediators of intercellular communication between cancerous and stromal cells within the TME, orchestrating complex pathophysiological processes. Chemokines are a family of small inflammatory cytokines that are able to induce chemotaxis in responsive cells. MVs which selective incorporate chemokines as their molecular cargos may play important regulatory roles in oncogenic processes including tumor proliferation, apoptosis, angiogenesis, metastasis, chemoresistance and immunomodulation, et al. Therefore, it is important to explore the association of MVs and chemokines in TME, identify the potential prognostic marker of tumor, and develop more effective treatment strategies. Here we review the relevant literature regarding the role of MVs and chemokines in TME.


Assuntos
Comunicação Celular , Micropartículas Derivadas de Células/metabolismo , Quimiocinas/metabolismo , Neoplasias/patologia , Microambiente Tumoral , Animais , Progressão da Doença , Espaço Extracelular/metabolismo , Humanos , Neoplasias/etiologia , Neoplasias/metabolismo
9.
Biomater Sci ; 7(5): 1852-1862, 2019 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-30899922

RESUMO

Galectins are attractive therapeutic candidates to control aberrant immune system activation because they can alter the phenotype and function of various innate and adaptive immune cells. However, use of exogenous galectin-1 ("G1") and galectin-3 ("G3") as immunomodulators is challenged by their high dosing requirements and dynamic quaternary structures. Here we report a chimeric assembly of G1 and G3 with enhanced extracellular activity ("G1/G3 Zipper"), which was created by recombinant fusion of G1 and G3 via a peptide linker that forms a two-stranded α-helical coiled-coil. G1/G3 Zipper had higher apparent binding affinity for immobilized lactose and a lower concentration threshold for inducing soluble glycoprotein crosslinking than G1, a recombinant fusion of G1 and G3 with a flexible peptide linker ("G1/G3"), or a recently reported stable G1 dimer crosslinked by poly(ethylene glycol) diacrylate ("G1-PEG-G1"). As a result, G1/G3 Zipper was more effective at inducing Jurkat T cell apoptosis in media containing serum, and was the only variant that could induce apoptosis at low concentrations under serum-free conditions. The monomeric G1/G3 fusion protein lacked extracellular activity under all conditions tested, suggesting that the enhanced activity of G1/G3 Zipper was due to its quaternary structure and increased carbohydrate-recognition domain valency. Thus, combining G1 and G3 into a non-native chimeric assembly provides a new candidate therapeutic with greater immunomodulatory potency than the wild-type proteins and previously reported engineered variants.


Assuntos
Espaço Extracelular/efeitos dos fármacos , Galectina 1/farmacologia , Galectina 3/farmacologia , Proteínas Recombinantes de Fusão/farmacologia , Metabolismo dos Carboidratos , Espaço Extracelular/metabolismo , Galectina 1/química , Galectina 1/metabolismo , Galectina 3/química , Galectina 3/metabolismo , Humanos , Células Jurkat , Modelos Moleculares , Conformação Proteica , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/metabolismo
10.
Nat Commun ; 10(1): 1355, 2019 03 22.
Artigo em Inglês | MEDLINE | ID: mdl-30902976

RESUMO

Extracellular electron uptake (EEU) is the ability of microbes to take up electrons from solid-phase conductive substances such as metal oxides. EEU is performed by prevalent phototrophic bacterial genera, but the electron transfer pathways and the physiological electron sinks are poorly understood. Here we show that electrons enter the photosynthetic electron transport chain during EEU in the phototrophic bacterium Rhodopseudomonas palustris TIE-1. Cathodic electron flow is also correlated with a highly reducing intracellular redox environment. We show that reducing equivalents are used for carbon dioxide (CO2) fixation, which is the primary electron sink. Deletion of the genes encoding ruBisCO (the CO2-fixing enzyme of the Calvin-Benson-Bassham cycle) leads to a 90% reduction in EEU. This work shows that phototrophs can directly use solid-phase conductive substances for electron transfer, energy transduction, and CO2 fixation.


Assuntos
Ciclo do Carbono , Dióxido de Carbono/metabolismo , Elétrons , Espaço Extracelular/metabolismo , Processos Fototróficos , Rodopseudomonas/metabolismo , Hidrogênio/metabolismo , Espaço Intracelular/metabolismo , Modelos Biológicos , Oxirredução , Fotossíntese , Rodopseudomonas/crescimento & desenvolvimento , Ribulose-Bifosfato Carboxilase/metabolismo
11.
Int J Mol Sci ; 20(6)2019 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-30909368

RESUMO

BACKGROUND: Vascular endothelial injury during ischemia generates apoptotic cell death and precedes apoptosis of underlying tissues. We aimed at studying the role of extracellular adenosine triphosphate (ATP) on endothelial cells protection against hypoxia injury. METHODS: In a hypoxic model on endothelial cells, we quantified the extracellular concentration of ATP and adenosine. The expression of mRNA (ectonucleotidases, adenosine, and P2 receptors) was measured. Apoptosis was evaluated by the expression of cleaved caspase 3. The involvement of P2 and adenosine receptors and signaling pathways was investigated using selective inhibitors. RESULTS: Hypoxic stress induced a significant increase in extracellular ATP and adenosine. After a 2-h hypoxic injury, an increase of cleaved caspase 3 was observed. ATP anti-apoptotic effect was prevented by suramin, pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS), and CGS15943, as well as by selective A2A, A2B, and A3 receptor antagonists. P2 receptor-mediated anti-apoptotic effect of ATP involved phosphoinositide 3-kinase (PI3K), extracellular signal-regulated kinases (ERK1/2), mitoKATP, and nitric oxide synthase (NOS) pathways whereas adenosine receptor-mediated anti-apoptotic effect involved ERK1/2, protein kinase A (PKA), and NOS. CONCLUSIONS: These results suggest a complementary role of P2 and adenosine receptors in ATP-induced protective effects against hypoxia injury of endothelial. This could be considered therapeutic targets to limit the development of ischemic injury of organs such as heart, brain, and kidney.


Assuntos
Trifosfato de Adenosina/metabolismo , Apoptose , Células Endoteliais da Veia Umbilical Humana/metabolismo , Hipóxia/metabolismo , Receptores Purinérgicos P1/metabolismo , Receptores Purinérgicos P2/metabolismo , Adenosina/metabolismo , Apoptose/genética , Biomarcadores , Espaço Extracelular/metabolismo , Expressão Gênica , Humanos , Hipóxia/genética , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Óxido Nítrico Sintase/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , RNA Mensageiro/genética , Receptores Purinérgicos P1/genética , Receptores Purinérgicos P2/genética , Transdução de Sinais , Estresse Fisiológico/genética
12.
Appl Microbiol Biotechnol ; 103(8): 3521-3535, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30852658

RESUMO

At present, anti-virulence drugs are being considered as potential therapeutic alternatives and/or adjuvants to currently failing antibiotics. These drugs do not kill bacteria but inhibit virulence factors essential for establishing infection and pathogenesis through targeting non-essential metabolic pathways reducing the selective pressure to develop resistance. We investigated the effect of naturally isolated plant compounds on the repression of the quorum sensing (QS) system which is linked to virulence/pathogenicity in Pseudomonas aeruginosa. Our results show that trans-cinnamaldehyde (CA) and salicylic acid (SA) significantly inhibit expression of QS regulatory and virulence genes in P. aeruginosa PAO1 at sub-inhibitory levels without any bactericidal effect. CA effectively downregulated both the las and rhl QS systems with lasI and lasR levels inhibited by 13- and 7-fold respectively compared to 3- and 2-fold reductions with SA treatment, during the stationary growth phase. The QS inhibitors (QSI) also reduced the production of extracellular virulence factors with CA reducing protease, elastase and pyocyanin by 65%, 22% and 32%, respectively. The QSIs significantly reduced biofilm formation and concomitantly with repressed rhamnolipid gene expression, only trace amount of extracellular rhamnolipids were detected. The QSIs did not completely inhibit virulence factor expression and production but their administration significantly lowered the virulence phenotypes at both the transcriptional and extracellular levels. This study shows the significant inhibitory effect of natural plant-derived compounds on the repression of QS systems in P. aeruginosa.


Assuntos
Acroleína/análogos & derivados , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/genética , Percepção de Quorum/efeitos dos fármacos , Ácido Salicílico/farmacologia , Fatores de Virulência/genética , Acroleína/farmacologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biofilmes/efeitos dos fármacos , Biofilmes/crescimento & desenvolvimento , Espaço Extracelular/metabolismo , Pseudomonas aeruginosa/crescimento & desenvolvimento , Pseudomonas aeruginosa/patogenicidade , Percepção de Quorum/genética , Virulência/efeitos dos fármacos , Fatores de Virulência/metabolismo
13.
Hum Cell ; 32(2): 88-94, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30730038

RESUMO

New neurons are continuously generated in the adult brain. This generation primarily occurs in the subventricular zone (SVZ) of the lateral ventricles and the subgranular zone (SGZ) of the hippocampal dentate gyrus. In the SGZ, neural stem cells (NSCs) give rise to glutamatergic granule cells that integrate into the hippocampal circuitry. Reduction of neurogenesis in the hippocampus impairs learning and memory, which suggests that this process is important for adult hippocampal function. Indeed, the neurogenesis is reduced in the progression of aging, which is thought to contribute to age-related cognitive impairment. Although the mechanism of age-dependent decline in neurogenesis remains largely obscure, astrocytes are thought to play a vital role in regulating NSC proliferation and differentiation. Both astrocytes and NSCs secrete nucleotides to the extracellular space and extracellular nucleotides bind to their receptors on the surface of target cells. In this review, the recent knowledge on adult neurogenesis in the hippocampus is summarized briefly, and possible role of extracellular nucleotides in the age-dependent changes of the adult neurogenesis is discussed.


Assuntos
Envelhecimento/metabolismo , Envelhecimento/patologia , Espaço Extracelular/metabolismo , Hipocampo/patologia , Hipocampo/fisiologia , Células-Tronco Neurais/metabolismo , Células-Tronco Neurais/fisiologia , Neurogênese/fisiologia , Nucleotídeos/metabolismo , Envelhecimento/fisiologia , Animais , Astrócitos/fisiologia , Diferenciação Celular , Proliferação de Células , Disfunção Cognitiva/etiologia , Hipocampo/citologia , Humanos , Aprendizagem , Memória , Células-Tronco Neurais/citologia , Via de Sinalização Wnt/fisiologia
14.
Nat Mater ; 18(4): 397-405, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30778227

RESUMO

The generation of organoids is one of the biggest scientific advances in regenerative medicine. Here, by lengthening the time that human pluripotent stem cells (hPSCs) were exposed to a three-dimensional microenvironment, and by applying defined renal inductive signals, we generated kidney organoids that transcriptomically matched second-trimester human fetal kidneys. We validated these results using ex vivo and in vitro assays that model renal development. Furthermore, we developed a transplantation method that utilizes the chick chorioallantoic membrane. This approach created a soft in vivo microenvironment that promoted the growth and differentiation of implanted kidney organoids, as well as providing a vascular component. The stiffness of the in ovo chorioallantoic membrane microenvironment was recapitulated in vitro by fabricating compliant hydrogels. These biomaterials promoted the efficient generation of renal vesicles and nephron structures, demonstrating that a soft environment accelerates the differentiation of hPSC-derived kidney organoids.


Assuntos
Espaço Extracelular/metabolismo , Rim/citologia , Organoides/citologia , Células-Tronco Pluripotentes/citologia , Técnicas de Cultura de Tecidos/métodos , Diferenciação Celular , Microambiente Celular , Feminino , Humanos , Cinética , Células-Tronco Pluripotentes/metabolismo , Gravidez , Terceiro Trimestre da Gravidez , Transcriptoma
16.
Int J Mol Sci ; 20(3)2019 Feb 11.
Artigo em Inglês | MEDLINE | ID: mdl-30754672

RESUMO

Since Biblical times, honey has been utilized in "folk medicine", and in recent decades the positive qualities of honey have been re-discovered and are gaining acceptance. Scientific literature states that honey has been successfully utilized on infections not responding to classic antiseptic and antibiotic therapy, because of its intrinsic H2O2 production. In our study, we demonstrated the involvement of H2O2 as a main mediator of honey regenerative effects on an immortalized human keratinocyte cell line. We observed that this extracellularly released H2O2 could pass across the plasma membrane through a specific aquaporin (i.e., AQP3). Once in the cytoplasm H2O2, in turn, induces the entry of extracellular Ca2+ through Melastatin Transient Receptor Potential 2 (TRPM2) and Orai1 channels. Honey-induced extracellular Ca2+ entry results in wound healing, which is consistent with the role played by Ca2+ signaling in tissue regeneration. This is the first report showing that honey exposure increases intracellular Ca2+ concentration ([Ca2+]i), due to H2O2 production and redox regulation of Ca2+-permeable ion channels, opening up a new horizon for the utilization of the honey as a beneficial tool.


Assuntos
Aquaporina 3/genética , Mel , Cicatrização , Aquaporina 3/metabolismo , Cálcio/metabolismo , Bloqueadores dos Canais de Cálcio/farmacologia , Canais de Cálcio/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Linhagem Celular , Relação Dose-Resposta a Droga , Espaço Extracelular/metabolismo , Humanos , Peróxido de Hidrogênio/metabolismo
17.
Bioresour Technol ; 279: 189-194, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30735927

RESUMO

This study aimed to investigate the effect of food to microorganisms rate (F/M) on organic removal, extracellular polymeric substances (EPS) and soluble microbial products (SMP) of the pure oxygen aerated activated sludge running in batch mode. The F/M rates were controlled by adjusting the MLSS concentrations (2000, 5000, 8000 mg/L) and/or the initial TOC concentrations (100, 500 mg/L). Results showed that at high F/M rate (0.25 kg TOC/kg MLSS), the substrate degradation rate in the oxygen aerated reactor could reach 1.347 mg TOC/(L·min)), much higher than that in the air aerated reactor (0.640 mg TOC/(L·min)). The SMP concentrations with oxygen aeration were also higher than those with air aeration under high F/M conditions. The total EPS contents in the pure oxygen aerated sludge were significantly lower regardless of the different F/M rates. High F/M condition would lead to more amount of polysaccharides synthesis rather than proteins synthesis in EPS.


Assuntos
Alimentos , Oxigênio/metabolismo , Esgotos , Reatores Biológicos , Espaço Extracelular/metabolismo , Microbiologia de Alimentos , Esgotos/química
18.
Eur J Pharmacol ; 852: 125-133, 2019 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-30797785

RESUMO

Stearoyl lysophosphatidylcholine (sLPC) has protective effects against several lethal sepsis models, even after induction of sepsis, which is associated with sLPC-mediated inhibition of high mobility group box 1 (HMGB1) release. This study investigated the mechanism by which sLPC inhibits lipopolysaccharide (LPS)-induced extracellular secretion of HMGB1 after the onset of sepsis. sLPC increased AMPK phosphorylation and the binding of AMPK to calcium/calmodulin-dependent protein kinase kinase ß (CaMKKß), one of the upstream signals of AMPK. Inhibition of CaMKKß activity decreased sLPC-mediated inhibition of HMGB1 release, and sLPC increased the concentration of intracellular calcium. Blocking of the macrophage G protein-coupled receptor G2A (G2A) suppressed AMPK phosphorylation, suppressed increases in the intracellular levels of calcium, and prevented the inhibition of HMGB1 release by sLPC. In particular, when macrophages were incubated with sLPC even after LPS treatment, sLPC increased the phosphorylation of AMPK and the binding of CaMKKß and AMPK, and suppressed the secretion of HMGB1. In addition, sLPC administered 1 h before or 4 h after establishment of sepsis significantly diminished circulating HMGB1 levels in mice. sLPC inhibited LPS-induced extracellular release of HMGB1 through the activation of the G2A/calcium/CaMKKß/AMPK pathway. These findings suggest that sLPC may be a potential anti-inflammatory agent for acute inflammatory conditions such as sepsis.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Quinase da Proteína Quinase Dependente de Cálcio-Calmodulina/metabolismo , Cálcio/metabolismo , Espaço Extracelular/efeitos dos fármacos , Proteína HMGB1/metabolismo , Lisofosfatidilcolinas/farmacologia , Receptores CCR10/metabolismo , Animais , Espaço Extracelular/metabolismo , Lipopolissacarídeos/farmacologia , Lisofosfatidilcolinas/uso terapêutico , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Células RAW 264.7 , Sepse/tratamento farmacológico , Sepse/imunologia , Sepse/metabolismo , Transdução de Sinais/efeitos dos fármacos
19.
Methods Mol Biol ; 1928: 353-363, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30725464

RESUMO

A large amount of energy used for nutrient processing and cellular functions is essential for tumorigenesis. Total intracellular adenosine triphosphate (ATP) is mainly generated by glycolysis and mitochondrial oxidative phosphorylation. Here, we provide a protocol for measurements of energy metabolism in cancer cells by using Seahorse XF24 Extracellular Flux analyzer. Specifically, this machine measures glycolysis by analyzing the extracellular acidification rate (ECAR) and measures mitochondrial oxidative phosphorylation on the basis of the oxygen consumption rate (OCR), through real-time and live cell analysis. This protocol is provided for researchers who are unfamiliar with the method and to aid them in carrying out the technique successfully.


Assuntos
Bioensaio/instrumentação , Bioensaio/métodos , Espaço Extracelular/metabolismo , Concentração de Íons de Hidrogênio , Neoplasias/metabolismo , Consumo de Oxigênio , Animais , Linhagem Celular , Análise de Dados , Metabolismo Energético , Glicólise , Humanos , Mitocôndrias/metabolismo , Fosforilação Oxidativa , Software
20.
Int J Biol Macromol ; 129: 571-578, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30753879

RESUMO

A novel extracellular xylanase was purified and characterized from Pediococcus acidilactici GC25 (GenBank number: MF289522). The purification was 4.6-fold with a yield of 43.61% through acetone precipitation, Q-Sepharose, and CM-Sepharose ion change chromatography. The molecular weight of the enzyme was 48.15 kDa, and the optimum pH and temperature were 7.0 and 40 °C, respectively. The maximum activity was observed between 20 and 50 °C. Although it was active within a wide pH range (pH 2.0-9.0), it retained over 85% of its activity after 24 h incubation; and over 70% of its activity after 168 h incubation in neutral and alkaline pH. It was observed that the enzyme showed high stability with K+, Ba2+, Cd2+, Co2+, Sr2+, Mg2+, Ca2+, Al3+, Zn2+, and Ni2+ ions. The Km and Vmax for the xylanase were 3.10 mg mL-1 and 4.66 U/mg protein, respectively. It was determined that treatment of different fruit juices with P. acidilactici GC25 xylanase improved the clarification. The highest increase in the reducing sugar amount and decrease in the turbidity was 24.47 ±â€¯1.08 and 21.22 ±â€¯0.58 for peach juice at 0.15 U/mL enzyme concentration. These results showed that the xylanase purified from P. acidilactici GC25 may have a wide potential in biotechnological processes of the food and baking industry.


Assuntos
Endo-1,4-beta-Xilanases/isolamento & purificação , Endo-1,4-beta-Xilanases/metabolismo , Manipulação de Alimentos , Sucos de Frutas e Vegetais , Pediococcus acidilactici/enzimologia , Estabilidade Enzimática , Espaço Extracelular/metabolismo , Concentração de Íons de Hidrogênio , Hidrólise , Cinética , Metais/farmacologia , Pediococcus acidilactici/citologia , Temperatura Ambiente
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