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1.
Environ Res ; 216(Pt 4): 114837, 2023 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-36400223

RESUMO

Nitrate pollution of groundwater has become an increasingly serious environmental problem that poses a great threat to aquatic ecosystems and to human health. Previous studies have shown that solid-phase humin (HM) can act as an additional electron donor to support microbial denitrification in the bioremediation of nitrate-contaminated groundwater where electron donor is deficient. However, the electron-donating capacities of HMs vary widely. In this study, we introduced ferrihydrite and prepared ferrihydrite-humin (Fh-HM) coprecipitates via biotic means to strengthen their electron-donating capacities. The spectroscopic results showed that the crystal phase of Fh did not change after coprecipitation with HM in the presence of Shewanella oneidensis MR-1, and iron may have complexed with the organic groups of HM. The Fh-HM coprecipitate prepared with an optimal initial Fh-HM mass ratio of 14:1 enhanced the microbial denitrification of Pseudomonas stutzeri with an electron-donating capacity 2.4-fold higher than that of HM alone, and the enhancement was not caused by greater bacterial growth. The alginate bead embedding assay indicated that the oxidation pathway of Fh-HM coprecipitate was mainly through direct contact between P. stutzeri and the coprecipitate. Further analyses suggested that quinone and organic-complexed Fe were the main electron-donating fractions of the coprecipitate. The results of the column experiments demonstrated that the column filled with Fh-HM-coated quartz sand exhibited a higher denitrification rate than the one filled with quartz sand, indicating its potential for practical applications.


Assuntos
Pseudomonas stutzeri , Humanos , Pseudomonas stutzeri/metabolismo , Nitratos/química , Desnitrificação , Elétrons , Areia , Quartzo/metabolismo , Ecossistema , Compostos Férricos/química , Oxirredução , Compostos Orgânicos
2.
BMC Microbiol ; 20(1): 355, 2020 11 19.
Artigo em Inglês | MEDLINE | ID: mdl-33213384

RESUMO

BACKGROUND: Manure application and sewage irrigation release many intestinal pathogens into the soil. After being introduced into the soil matrix, pathogens are commonly found to attach to soil minerals. Although the survival of mineral-associated Escherichia coli O157:H7 has been studied, a comprehensive understanding of the attachment process and physiological properties after attachment is still lacking. RESULTS: In this study, planktonic and attached Escherichia coli O157:H7 cells on quartz were investigated using RNA sequencing (RNA-seq) and the isobaric tagging for relative and absolute quantitation (iTRAQ) proteomic method. Based on the transcriptomic and proteomic analyses and gene knockouts, functional two-component system pathways were required for efficient attachment; chemotaxis and the Rcs system were identified to play determinant roles in E. coli O157:H7 attachment on quartz. After attachment, the pyruvate catabolic pathway shifted from the tricarboxylic acid (TCA) cycle toward the fermentative route. The survival rate of attached E. coli O157:H7 increased more than 10-fold under penicillin and vancomycin stress and doubled under alkaline pH and ferric iron stress. CONCLUSIONS: These results contribute to the understanding of the roles of chemotaxis and the Rcs system in the attachment process of pathogens and indicate that the attachment of pathogens to minerals significantly elevates their resistance to antibiotics and environmental stress, which may pose a potential threat to public health.


Assuntos
Aderência Bacteriana , Escherichia coli O157/fisiologia , Quartzo/metabolismo , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Quimiotaxia , Farmacorresistência Bacteriana , Escherichia coli O157/efeitos dos fármacos , Escherichia coli O157/genética , Escherichia coli O157/metabolismo , Perfilação da Expressão Gênica , Proteômica , Transdução de Sinais , Estresse Fisiológico
3.
Langmuir ; 35(17): 5911-5920, 2019 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-30955325

RESUMO

Molecular recognition between peptides and metal oxide surfaces is a fundamental process in biomineralization, self-assembly, and biocompatibility. Yet, the underlying driving forces and dominant mechanisms remain unclear, bringing obstacles to understand and control this process. To elucidate the mechanism of peptide/surface recognition, specifically the role of serine phosphorylation, we employed molecular dynamics simulation and metadynamics-enhanced sampling to study five artificial peptides, DDD, DSS, DpSpS, DpSpSGKK, and DpSKGpSK, interacting with two surfaces: rutile TiO2 and quartz SiO2. On both surfaces, we observe that phosphorylation increases the binding energy. However, the interfacial peptide conformation reveals a distinct binding mechanism on each surface. We also study the impact of peptide sequence to binding free energy and interfacial conformation on both surfaces, specifically the impact on the behavior of phosphorylated serine. Finally, the results are discussed in context of prior studies investigating the role of serine phosphorylation in peptide binding to silica.


Assuntos
Fosfopeptídeos/metabolismo , Quartzo/metabolismo , Titânio/metabolismo , Adsorção , Simulação de Dinâmica Molecular , Fosfopeptídeos/química , Ligação Proteica , Quartzo/química , Eletricidade Estática , Termodinâmica , Titânio/química
4.
Appl Microbiol Biotechnol ; 97(3): 1317-24, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22410741

RESUMO

The attachment of Acidithiobacillus ferrooxidans and Leptospirillum ferriphilum spp. grown on ferrous medium or adapted to a pyrite mineral concentrate to four mineral substrata, namely, chalcopyrite and pyrite concentrates, a low-grade chalcopyrite ore (0.5 wt%) and quartzite, was investigated. The quartzite represented a typical gangue mineral and served as a control. The attachment studies were carried out in a novel particle-coated column reactor. The saturated reactor containing glass beads, which were coated with fine mineral concentrates, provided a quantifiable surface area of mineral concentrate and maintained good fluid flow. A. ferrooxidans and Leptospirillum spp. had similar attachment characteristics. Enhanced attachment efficiency occurred with bacteria grown on sulphide minerals relative to those grown on ferrous sulphate in an ore-free environment. Selective attachment to sulphide minerals relative to gangue materials occurred, with mineral adapted cultures attaching to the minerals more efficiently than ferrous grown cultures. Mineral-adapted cultures showed highest levels of attachment to pyrite (74% and 79% attachment for A. ferrooxidans and L. ferriphilum, respectively). This was followed by attachment of mineral-adapted cultures to chalcopyrite (63% and 58% for A. ferrooxidans and L. ferriphilum, respectively). A. ferrooxidans and L. ferriphilum exhibited lower levels of attachment to low-grade ore and quartz relative to the sulphide minerals.


Assuntos
Bactérias/crescimento & desenvolvimento , Bactérias/metabolismo , Aderência Bacteriana , Fenômenos Fisiológicos Bacterianos , Cobre/metabolismo , Ferro/metabolismo , Quartzo/metabolismo , Sulfetos/metabolismo , Reatores Biológicos , Minerais/metabolismo
5.
Biotechnol Lett ; 35(12): 2129-36, 2013 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23974497

RESUMO

Attachments of Acidithiobacillus ferrooxidans ATCC 23270 onto elemental sulfur, quartz and complex chalcopyrite were investigated by analysis of its extracellular polymeric substances as well as applying Langmuir and Freundlich equations. The two equations fitted the adsorption equilibrium data with significant correlation coefficient over 0.9. This indicated that bacterial attachment is complicated and involves Langmuir and Freundlich characterizations. Sulfur-grown cells showed the highest affinity for the three solid substrates. The investigated complex chalcopyrite possessed a higher maximum adsorption capacity for A. ferrooxidans than elemental sulfur or quartz. The Freundlich fitting parameters suggested that quartz had a weaker adsorption capacity and smaller adsorption areas than elemental sulfur or the complex chalcopyrite. It is not the content of total carbohydrates or proteins in EPS but their ratios that determine the affinity differences between cells and substrates.


Assuntos
Acidithiobacillus/metabolismo , Aderência Bacteriana/fisiologia , Cobre/metabolismo , Modelos Teóricos , Quartzo/metabolismo , Enxofre/metabolismo , Acidithiobacillus/química , Acidithiobacillus/citologia , Adsorção , Espaço Extracelular/química , Espaço Extracelular/metabolismo , Cinética , Modelos Lineares , Polissacarídeos Bacterianos/química , Polissacarídeos Bacterianos/metabolismo
6.
Chemistry ; 18(24): 7388-93, 2012 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-22615170

RESUMO

Using a highly sensitive flow-type 27 MHz quartz crystal microbalance, we could detect a small mass change during stepwise and alternating one-sugar transfer of glucuronic acid (GlcA) and N-acetylgalactosamine (GalNAc) to an acceptor, catalyzed by chondroitin polymerase from Escherichia coli strain K4 (K4CP), and analyze the elongation mechanism of K4CP. K4CP was found to bind strongly to a chondroitin acceptor (K(d)=0.97 µM). Although the binding affinity and the catalytic rate constant for each monomer were considerably different, the apparent catalytic efficiency (k(cat)/K(m)) was similar (6.3×10(4) M(-1) s(-1) for GlcA transfer and 3.4×10(4) M(-1) s(-1) for the GalNAc transfer). This is reasonable for the smooth alternating elongation of GlcA and GalNAc on the acceptor. This is the first study to report the determination of kinetic parameters for enzymatic, alternated, sugar elongation.


Assuntos
Hexosiltransferases/metabolismo , Polissacarídeos/metabolismo , Catálise , Ácido Glucurônico/química , Ácido Glucurônico/metabolismo , Glicosilação , Cinética , Modelos Químicos , Polissacarídeos/química , Quartzo/metabolismo
7.
Environ Sci Technol ; 46(3): 1511-8, 2012 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-22243624

RESUMO

Enterococcus was selected by US EPA as a Gram-positive indicator microorganism for groundwater fecal contamination. It was recently reported that enterococcal surface protein (esp) was more prevalent in Enterococcus from human sources than in Enterococcus from nonhuman sources and esp could potentially be used as a source tracking tool for fecal contamination (Scott et al., 2005). In this research, we performed laboratory column transport experiments to investigate the transport of Enterococcus faecium within saturated quartz sands. Particularly, we used a wild type strain (E1162) and a mutant (E1162Δesp) to examine the influence of esp on the transport behavior of E. faecium. Our results showed that esp could significantly enhance the attachment of E. faecium cells onto the surface of silica sands and thus lower the mobility of E. faecium within sand packs. Cell surface properties (e.g., zeta potential) were determined and the extended Derjaguin-Landau-Verwey-Overbeek (XDLVO) theory was applied to explain the effects of esp on the retention of E. faecium. Overall, our results suggested that E. faecium strains with esp could display lower mobility within saturated sand packs than E. faecium strains without esp. The disparity in the transport behavior of E. faecium with and without esp could limit the effectiveness of esp as a source tracking tool within the groundwater system.


Assuntos
Enterococcus faecium/metabolismo , Água Subterrânea/microbiologia , Proteínas de Membrana/metabolismo , Quartzo/metabolismo , Adesão Celular/fisiologia , Monitoramento Ambiental/métodos , Fezes/microbiologia , Humanos , Proteínas de Membrana/fisiologia , Modelos Químicos , Movimento
8.
J Contam Hydrol ; 248: 104016, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35512510

RESUMO

Pathogenic bacteria can be discharged in the environment through natural as well as anthropogenic activities. Once in the environment, they may contaminate soil and sediments and migrate towards water bodies. Transient chemical conditions may occur in soil/sediments and favor mobilization of bacteria, e.g., upon the reduction of salinity (or ionic strength). However, the magnitude of this phenomenon and its relationship with particle size is not well understood, yet. In this work, we investigated the transport of Escherichia coli under variable salinity conditions (between 1 and 20 part per thousand, ppt) and for different soil grain sizes (between 150 and 710 µm). A model developed in our group was applied in this work. It couples bacteria and salinity transport equations in order to account for transient water composition in the description of bacteria migration. The model was calibrated and validated with laboratory experiments. The tests were monitored continuously with UV-Vis spectroscopy, which allowed to record highly resolved concentration fronts. The results show that salinity increases the retardation of the bacteria. Upon salinity drop, a release of bacteria occurs forming a peak whose magnitude increases with salinity change. This effect becomes more important as the grain size decreases. Simulations suggest that the dominant retention mechanism is attachment for coarse sand and straining for fine sand. The retention can be reversed as the salinity is reduced causing a sudden bacteria mobilization. Such a behaviour may have important implications on microbial contamination of water bodies when soil/sediments undergo transient chemical conditions.


Assuntos
Quartzo , Areia , Escherichia coli/metabolismo , Porosidade , Quartzo/metabolismo , Salinidade , Solo , Água/química
9.
Nature ; 433(7024): 399-402, 2005 Jan 27.
Artigo em Inglês | MEDLINE | ID: mdl-15674287

RESUMO

Silicon is the second most abundant element on Earth. It is an important nutrient for phytoplankton and is readily absorbed by terrestrial vegetation; it also assists the removal of carbon dioxide from the atmosphere through the weathering of silicates. But the continental cycle of silicon is not well known, and only a few studies have attempted to use silicon stable isotopes (28Si, 29Si and 30Si) to quantify the continental silicon reservoirs. Dissolved silicon in sea and river waters forms a reservoir of mean isotopic value +1.1 per thousand (refs 7, 10). It is enriched in 30Si with respect to the igneous rocks reservoir, which has a mean isotopic value of -0.3 per thousand (refs 4, 9). This enrichment can only be produced by a major fractionation during weathering, and should result in the formation of a continental 30Si-depleted reservoir. Such a reservoir, however, has not been identified to date. Here we analyse silicon isotopes of in situ quartz from a sandstone series in France, using a new-generation secondary ion mass spectrometry apparatus. We show that quartz that precipitates as siliceous cements forms a strongly 30Si-depleted reservoir with isotopic values down to -5.7 per thousand, a more negative value than any previously published for terrestrial samples. Our findings suggest that quartz re-precipitation plays an important role in the biogeochemical cycle of silicon.


Assuntos
Sedimentos Geológicos/química , Silício/química , Silício/metabolismo , Precipitação Química , Clima , Cristalização , Diatomáceas/metabolismo , França , Isótopos , Fitoplâncton/metabolismo , Quartzo/química , Quartzo/metabolismo , Água/química , Água/metabolismo
10.
J Exp Med ; 189(9): 1497-506, 1999 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-10224290

RESUMO

Alveolar macrophages (AMs) avidly bind and ingest unopsonized environmental particles and bacteria through scavenger-type receptors (SRs). AMs from mice with a genetic deletion of the major macrophage SR (types AI and AII; SR-/-) showed no decrease in particle binding compared with SR+/+ mice, suggesting that other SRs are involved. To identify these receptors, we generated a monoclonal antibody (mAb), PAL-1, that inhibits hamster AM binding of unopsonized particles (TiO2, Fe2O3, and latex beads; 66 +/- 5, 77 +/- 2, and 85 +/- 2% inhibition, respectively, measured by flow cytometry). This antibody identifies a protein of approximately 70 kD on the AM surface (immunoprecipitation) that is expressed by AMs and other macrophages in situ. A cDNA clone encoding the mAb PAL-1-reactive protein isolated by means of COS cell expression was found to be 84 and 77% homologous to mouse and human scavenger receptor MARCO mRNA, respectively. Transfection of COS cells with MARCO cDNA conferred mAb-inhibitable TiO2 binding. Hamster MARCO also mediates AM binding of unopsonized bacteria (67 +/- 5 and 47 +/- 4% inhibition of Escherichia coli and Staphylococcus aureus binding by mAb PAL-1). A polyclonal antibody to human MARCO identified the expected approximately 70-kD band on Western blots of lysates of normal bronchoalveolar lavage (BAL) cells (>90% AMs) and showed strong immunolabeling of human AMs in BAL cytocentrifuge preparations and within lung tissue specimens. In normal mouse AMs, the anti-MARCO mAb ED31 also showed immunoreactivity and inhibited binding of unopsonized particles (e.g., TiO2 approximately 40%) and bacteria. The novel function of binding unopsonized environmental dusts and pathogens suggests an important role for MARCO in the lungs' response to inhaled particles.


Assuntos
Macrófagos Alveolares/metabolismo , Proteínas de Membrana , Receptores Imunológicos/fisiologia , Receptores de Lipoproteínas , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/imunologia , Sequência de Bases , Células COS , Clonagem Molecular , Cricetinae , DNA Complementar , Escherichia coli/metabolismo , Humanos , Camundongos , Dados de Sequência Molecular , Testes de Precipitina , Quartzo/metabolismo , Receptores Imunológicos/genética , Receptores Imunológicos/metabolismo , Receptores Depuradores , Receptores Depuradores Classe B , Staphylococcus aureus/metabolismo , Titânio/metabolismo
11.
Microb Ecol ; 60(4): 730-9, 2010 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-20336290

RESUMO

The hypolithic microbial community associated with quartz pavement at a high-altitude tundra location in central Tibet is described. A small-scale ecological survey indicated that 36% of quartz rocks were colonized. Community profiling using terminal restriction fragment length polymorphism revealed no significant difference in community structure among a number of colonized rocks. Real-time quantitative PCR and phylogenetic analysis of environmental phylotypes obtained from clone libraries were used to elucidate community structure across all domains. The hypolithon was dominated by cyanobacterial phylotypes (73%) with relatively low frequencies of other bacterial phylotypes, largely represented by the chloroflexi, actinobacteria, and bacteriodetes. Unidentified crenarchaeal phylotypes accounted for 4% of recoverable phylotypes, while algae, fungi, and mosses were indicated by a small fraction of recoverable phylotypes.


Assuntos
Bactérias/isolamento & purificação , Ecossistema , Sedimentos Geológicos/microbiologia , Altitude , Bactérias/classificação , Bactérias/genética , Bactérias/metabolismo , DNA Bacteriano/genética , Dados de Sequência Molecular , Filogenia , Quartzo/metabolismo , RNA Ribossômico 16S/genética , Tibet
12.
J Basic Microbiol ; 50(2): 160-70, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-20082378

RESUMO

An aqueous mixture of goethite, quartz, and lead chloride (PbCl(2)) was treated with the sulfate-reducing bacterium, Desulfovibrio desulfuricans G20 (D. desulfuricans G20), in a medium specifically designed to assess metal toxicity. In the presence of 26 muM of soluble Pb, together with the goethite and quartz, D. desulfuricans G20 grew after a lag time of 5 days compared to 2 days in Pb-, goethite-, and quartz-free treatments. In the absence of goethite and quartz, however, with 26 microM soluble Pb, no measurable growth was observed. Results showed that D. desulfuricans G20 first removed Pb from solutions then growth began resulting in black precipitates of Pb and iron sulfides. Transmission electron microscopic analyses of thin sections of D. desulfuricans G20 treated with 10 microM PbCl(2) in goethite- and quartz-free treatment showed the presence of a dense deposit of lead sulfide precipitates both in the periplasm and cytoplasm. However, thin sections of D. desulfuricans G20 treated with goethite, quartz, and PbCl(2) (26 microM soluble Pb) showed the presence of a dense deposit of iron sulfide precipitates both in the periplasm and cytoplasm. Energy-dispersive X-ray spectroscopy, selected area electron diffraction patterns, or X-ray diffraction analyses confirmed the structure of precipitated Pb inside the cell as galena (PbS) in goethite- and quartz-free treatments, and iron sulfides in treatments with goethite, quartz, and PbCl(2). Overall results suggest that even at the same soluble Pb concentration (26 microM), in the presence of goethite and quartz, apparent Pb toxicity to D. desulfuricans G20 decreased significantly. Further, accumulation of lead/iron sulfides inside D. desulfuricans G20 cells depended on the presence of goethite and quartz.


Assuntos
Desulfovibrio desulfuricans/efeitos dos fármacos , Compostos de Ferro/metabolismo , Chumbo/metabolismo , Chumbo/toxicidade , Quartzo/metabolismo , Citoplasma/química , Citoplasma/ultraestrutura , Desulfovibrio desulfuricans/crescimento & desenvolvimento , Desulfovibrio desulfuricans/metabolismo , Desulfovibrio desulfuricans/ultraestrutura , Ferro/análise , Chumbo/análise , Microscopia Eletrônica de Transmissão , Minerais , Periplasma/química , Periplasma/ultraestrutura , Espectrometria por Raios X , Sulfetos/análise
13.
Proteomics ; 9(8): 2052-63, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19337994

RESUMO

This paper describes a novel strategy to create a microarray of G-protein coupled receptors (GPCRs), an important group of membrane proteins both physiologically and pharmacologically. The H(1)-histamine receptor and the M(2)-muscarinic receptor were both used as model GPCRs in this study. The receptor proteins were embedded in liposomes created from the cellular membrane extracts of Spodoptera frugiperda (Sf9) insect cell culture line with its accompanying baculovirus protein insert used for overexpression of the receptors. Once captured onto a surface these liposomes provide a favourable lipidic environment for the integral membrane proteins. Site directed immobilisation of these liposomes was achieved by introduction of cholesterol-modified oligonucleotides (oligos). These oligo/cholesterol conjugates incorporate within the lipid bilayer and were captured by the complementary oligo strand exposed on the surface. Sequence specific immobilisation was demonstrated using a quartz crystal microbalance with dissipation (QCM-D). Confirmatory results were also obtained by monitoring fluorescent ligand binding to GPCRs captured on a spotted oligo microarray using Confocal Laser Scanning Microscopy and the Zepto-READER microarray imaging system. Sequence specific immobilisation of such biologically important membrane proteins could lead to the development of a heterogeneous self-sorting liposome array of GPCRs which would underpin a variety of future novel applications.


Assuntos
Lipossomos/metabolismo , Análise Serial de Proteínas/métodos , Receptor Muscarínico M2/metabolismo , Receptores Histamínicos H1/metabolismo , Animais , Linhagem Celular , Ligantes , Microscopia Confocal , Oligodesoxirribonucleotídeos/metabolismo , Quartzo/metabolismo , Receptor Muscarínico M2/química , Receptores Histamínicos H1/química
14.
Toxicol Lett ; 176(2): 131-7, 2008 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-18079072

RESUMO

Inflammation is considered as a key event in adverse health effects associated with exposure to ambient particulate matter. The inflammatory potential of particles is often compared using in vitro cell systems, where the particle-induced release of pro-inflammatory cytokines is measured. A major concern in these assays is the potential of particles to bind cytokines, which may lead to an underestimation of the inflammatory potential. We therefore investigated the cytokine binding to a selection of particle samples, including particles collected from outdoor sources (wood combustion, traffic) and particles commonly used to model environmental sources (ultrafine carbon black, diesel, quartz), for a range of pro-inflammatory cytokines (TNF-alpha, IL-1beta, IL-6, IL-8). Furthermore, the influence of serum proteins and particle- and cytokine concentrations on the cytokine binding was studied. Cytokines primarily bound to carbonaceous particles (up to 85%), not to mineral particles. Furthermore, depending on the type of cytokine, the cytokine binding could be reduced partly or completely by adding serum proteins to the cell growth medium or particle suspensions. Based on these observations we recommend either to adjust culturing and exposure conditions to prevent cytokine binding, or to adjust the measured cytokine release by application of correction factors obtained from cytokine binding experiments.


Assuntos
Interleucinas/metabolismo , Material Particulado/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Adsorção , Análise de Variância , Animais , Ligação Competitiva , Proteínas Sanguíneas/análise , Proteínas Sanguíneas/metabolismo , Bovinos , Meios de Cultura/química , Monitoramento Ambiental/métodos , Interleucinas/genética , Minerais/análise , Minerais/química , Minerais/metabolismo , Tamanho da Partícula , Material Particulado/análise , Material Particulado/química , Ligação Proteica , Quartzo/análise , Quartzo/química , Quartzo/metabolismo , Proteínas Recombinantes/metabolismo , Soro/química , Soro/efeitos dos fármacos , Soro/metabolismo , Soroalbumina Bovina/análise , Soroalbumina Bovina/metabolismo , Fumaça/análise , Fuligem/química , Fuligem/metabolismo , Fator de Necrose Tumoral alfa/genética , Emissões de Veículos/análise , Madeira
15.
Toxicol In Vitro ; 21(3): 438-48, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17125965

RESUMO

This paper describes the in vitro cytotoxicity assessment of single walled carbon nanotubes (SWCNT) on A549 cells, a human lung cell line. Cellular viability was determined using the alamar blue (AB), neutral red (NR) and MTT assays, which evaluated metabolic, lysosomal and mitochondrial activity respectively. In addition, the total protein content of the cells was measured using the coomassie brilliant (CB) blue assay. Supernatants were also assayed for Adenylate Kinase (AK) release and Interleukin 8 (IL-8) which indicated a loss of cell membrane integrity and an inflammation response respectively. To investigate the interactions between serum components in the test medium and the test materials, exposures were conducted both in serum containing (5%) and serum-free medium. Results from the cytotoxicity tests (AB, CB, MTT) revealed the SWCNT to have very low acute toxicity to the A549 cells as all but one of the reported 24h EC(50) values exceeded the top concentration tested (800 microg/ml). The SWCNT were found to interfere with a number of the dyes used in the cytotoxicity assessment and we are currently conducting a comprehensive spectroscopic study to further investigate these interactions. Of the multiple cytotoxicity assays used, the AB assay was found to be the most sensitive and reproducible. Transmission electron microscopy (TEM) studies confirmed that there was no intracellular localization of SWCNT in A549 cells following 24h exposure; however, increased numbers of surfactant storing lamellar bodies were observed in exposed cells.


Assuntos
Células Epiteliais/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Nanotecnologia , Nanotubos de Carbono/toxicidade , Adenilato Quinase/metabolismo , Linhagem Celular , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/ultraestrutura , Sobrevivência Celular/efeitos dos fármacos , Meios de Cultivo Condicionados/química , Citoplasma/efeitos dos fármacos , Citoplasma/ultraestrutura , Relação Dose-Resposta a Droga , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Formazans/metabolismo , Humanos , Indicadores e Reagentes/metabolismo , Interleucina-8/metabolismo , Pulmão/metabolismo , Pulmão/patologia , Microscopia Eletrônica de Transmissão , Nanotubos de Carbono/química , Nanotubos de Carbono/ultraestrutura , Necrose/induzido quimicamente , Vermelho Neutro/metabolismo , Oxazinas/metabolismo , Quartzo/metabolismo , Quartzo/toxicidade , Corantes de Rosanilina/metabolismo , Sais de Tetrazólio/metabolismo , Xantenos/metabolismo
16.
Asian Pac J Cancer Prev ; 16(12): 5115-8, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26163652

RESUMO

BACKGROUND: The p16INK4a is a protein that expressed in Liquid-based cervical cytology specimens and has been proved link to cervical cancer. The p16INK4a could be detection by piezoelectric immunosensor and the immobilization of the p16INK4a antibody influence the sensitivity of the piezoelectric immunosensor. MATERIALS AND METHODS: 5µL mouse polyclonal antibody against p16INK4a was bound onto the surface of immonosensor through two methods. (directly immobilized method; protein A method). Absorb of the p16INK4a antibody on the surface of immonosensor caused a shift in the resonant frequency of the immunosensor and The frequency changes recorded showed a better reproducibility. The activity of the immobilization antibody with the directly method and protein A method was tested with p16INK4a antigen. RESULTS: The resonant frequency for different antibody immobilization methods were different, and the sensitivity for p16INK4a detection also different. CONCLUSIONS: The protein A method was found to be much more better than the directly method for the immobilization of the p16INK4A antibody on the gold electrode of the quartz crystal for cervical lesion detection. The Protein A method created more reproducible and stable immobilization antibody layers with p16INK4A antigen.


Assuntos
Anticorpos Imobilizados/imunologia , Técnicas Biossensoriais/instrumentação , Inibidor p16 de Quinase Dependente de Ciclina/imunologia , Imunoensaio/instrumentação , Quartzo/química , Neoplasias do Colo do Útero/diagnóstico , Animais , Técnicas Biossensoriais/métodos , Inibidor p16 de Quinase Dependente de Ciclina/sangue , Detecção Precoce de Câncer , Eletrodos , Feminino , Ouro/química , Humanos , Imunoensaio/métodos , Camundongos , Quartzo/metabolismo , Proteína Estafilocócica A/imunologia , Proteína Estafilocócica A/metabolismo , Neoplasias do Colo do Útero/sangue , Neoplasias do Colo do Útero/imunologia
17.
Biosens Bioelectron ; 19(9): 987-97, 2004 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-15018953

RESUMO

A method for eliminating the mass transport limitation on biosensor surfaces is introduced. The measurement of macromolecular binding kinetics on plane surfaces is the key objective of many evanescent wave (e.g. total internal reflection fluorescence (TIRF)), and surface plasmon resonance (SPR) based biosensor systems, allowing the determination of binding constants within minutes or hours. However, these methods are limited in not being rigorously applicable to large macromolecules like proteins or DNA, since the on-rates are transport limited due to a Nernst diffusion layer of 5-10 microm thickness. Thus, for the binding of fibrinogen (340 kDa) to a surface current SPR biosensors will show a mass transport coefficient of ca. 2 x 10(-6) m/s. In a novel approach with an immiscible fluid vesicle (e.g. air bubble), it has been possible to generate nanoscopic fluid films of ca. 200 nm thickness on the sensor surface of an interfacial TIRF rheometer system. The thickness of the liquid film can be can be easily probed and measured by evanescent wave technology. This nanofilm technique increases the mass transport coefficient for fibrinogen to ca. 1 x 10(-4) m/s eliminating the mass transport limitation, making the binding rates reaction-rate limited. From the resulting exponential kinetic functions, lasting only 20-30s, the kinetic constants for the binding reaction can easily be extracted and the binding constants calculated. As a possible mechanism for the air bubble effect it is suggested that the aqueous fluid flow in the rheometer cell is separated by the air bubble below the level of the Nernst boundary layer into two independent laminar fluid flows of differing velocity: (i) a slow to stationary nanostream ca. 200 nm thick strongly adhering to the surface; and (ii) the bulk fluid streaming over it at a much higher rate in the wake of the air bubble. Surprising properties of the nanofluidic film are: (i) its long persistence for at least 30-60s after the air bubble has passed (2.5s); and (ii) the absence of solute depletion. It is suggested that a new liquid-liquid interface (i.e. a "vortex sheet") between the two fluid flows plays a decisive role, lending metastability to the nanofluidic film and replenishing its protein concentration via the vortices-thus upholding exponential binding kinetics. Finally, the system relaxes via turbulent reattachment of the two fluid flows to the original velocity profile. It is concluded that this technique opens a fundamentally novel approach to the construction of macromolecular biosensors.


Assuntos
Técnicas Biossensoriais , Interpretação Estatística de Dados , Fibrinogênio/metabolismo , Quartzo/metabolismo , Ressonância de Plasmônio de Superfície , Fatores de Tempo
18.
Anal Sci ; 18(4): 403-7, 2002 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-11999512

RESUMO

A novel piezoelectric immunosensor has been developed for the determination of beta-indole acetic acid (IAA) in dilute solutions. The detection is based on competitive immunoreaction between a hapten (IAA) and an antigen (IAA-BSA, hapten-protein conjugation) bound to an anti-IAA antibody, immobilized on a quartz crystal microbalance (QCM). The frequency change (y) of the sensor caused by antigen is linearly related to the logarithm of the concentration of IAA (x) in the range of 0.5 ng/ml - 5 microg/ml with a regression equation of the form y = -23x + 151 (r = 0.9937).


Assuntos
Técnicas Biossensoriais/instrumentação , Técnicas Biossensoriais/métodos , Ácidos Indolacéticos/análise , Animais , Calibragem , Imunoensaio/métodos , Imunoconjugados/imunologia , Imunoglobulina G/isolamento & purificação , Imunoglobulina G/metabolismo , Ácidos Indolacéticos/imunologia , Ácidos Indolacéticos/metabolismo , Reguladores de Crescimento de Plantas/análise , Reguladores de Crescimento de Plantas/imunologia , Reguladores de Crescimento de Plantas/metabolismo , Quartzo/metabolismo , Coelhos , Soroalbumina Bovina/imunologia , Soroalbumina Bovina/metabolismo , Fatores de Tempo
19.
Hum Exp Toxicol ; 14(3): 266-72, 1995 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-7779457

RESUMO

1. A new specific protein kinase C (PKC) inhibitor, Ro 31-7549, was used to explore the mechanisms by which particulate stimuli, quartz and chrysotile, stimulate human polymorphonuclear leukocytes (PMNL) to produce reactive oxygen metabolites (ROM). Also soluble stimuli, formyl-Methionyl-Leucyl-Phenylalanine (fMLP) and phorbol myristate acetate (PMA) were used. 2. Ro 31-7549 inhibited chrysotile-induced free intracellular calcium ([Ca2+]i) elevations but did not have an effect on quartz-induced elevations of [Ca2+]i. Both quartz and chrysotile induced production of ROM were partially inhibited by Ro 31-7549. fMLP-induced elevation of [Ca2+]i was inhibited by Ro 31-7549 whereas PMA did not affect [Ca2+]i. Ro 31-7549 strongly inhibited fMLP-induced ROM production, and completely abolished that induced by PMA. 3. These result suggest that PKC may have an important role in the activation of PMNL to produce ROM by particulate and soluble stimuli. However, the inhibition of chrysotile-, but not of quartz-induced [Ca2+]i elevations by Ro 31-7549 provides evidence that both PKC-dependent and -independent mechanisms may play a role in the activation of human leukocytes to produce ROM.


Assuntos
Cálcio/análise , Indóis/farmacologia , Maleimidas/farmacologia , Neutrófilos/efeitos dos fármacos , Proteína Quinase C/antagonistas & inibidores , Espécies Reativas de Oxigênio/análise , Adulto , Asbestos Serpentinas/metabolismo , Cálcio/metabolismo , Humanos , N-Formilmetionina Leucil-Fenilalanina/metabolismo , Fagocitose/fisiologia , Quartzo/metabolismo , Acetato de Tetradecanoilforbol/metabolismo
20.
Ind Health ; 34(4): 379-88, 1996.
Artigo em Inglês | MEDLINE | ID: mdl-8908848

RESUMO

Bronchoalveolar lavage (BAL) is a useful and safe method for sampling cellular and biochemical components from the lung. Analysis of bronchoaveolar lavage fluid (BALF) constituents is useful for defining the stage of disease, and for assessing disease progression and the response to therapy in lung disorders. We studied the dynamic changes in various indices for BALF and the accompanying silicotic changes in the lungs of rats at different times after quartz instillation. Total cell counts, LDH activity, protein concentration, and lipoperoxide (LPO) in the BALF of experimental silicotic rats were significantly higher than those of control rats (P < 0.05 or 0.01). After instillation, quartz content, total cell counts, LDH activity and protein concentration in BALF tended to decrease over time. These findings suggested that in acute silicosis, quartz can induce serious inflammation and damage the lung, with acute lung proteinosis seen as the main change in this stage.


Assuntos
Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/citologia , Quartzo/metabolismo , Silicose/patologia , Animais , Modelos Animais de Doenças , Masculino , Distribuição Aleatória , Ratos , Ratos Wistar , Estatísticas não Paramétricas
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