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This study was design to evaluate the physiological properties of bacteriophage-insensitive Klebsiella pneumoniae (BIKP) mutants in association with the antibiotic cross-resistance, ß-lactamase activity, and gene expression. Klebsiella pneumoniae ATCC 23357(KPWT), ciprofloxacin-induced antibiotic-resistant K. pneumoniae ATCC 23357 (KPCIP), and clinically isolated antibiotic-resistant K. pneumoniae 10263 (KPCLI) were used to isolate BIKP mutants against KPB1, PBKP02, PBKP21, PBKP29, PBKP33, and PBKP35. PBKP35-induced mutants, including bacteriophage-insensitive K. pneumoniae ATCC 23357 (BIKPWT), ciprofloxacin-induced K. pneumoniae ATCC 23357 (BIKPCIP), and clinically isolated antibiotic-resistant K. pneumoniae CCARM 10263 (BIKPCLI). BIKPWT, BIKPCIP, and BIKPCLI were resistant to Klebsiella bacteriophages, KPB1, PBKP02, PBKP21, PBKP29, and PBKP33. The antibiotic cross-resistance to cefotaxime, cephalothin, chloramphenicol, ciprofloxacin, erythromycin, kanamycin, levofloxacin, and nalidixic acid was observed in BIKPWT. The relative expression levels of vagC was increased by more than 8-folds in BIKPWT, corresponding to the increased ß-lactamase activity. The aac(6')-Ib-cr was overexpressed in BIKP mutants, responsible for aminoglycoside and quinolone resistance. The phage-resistant mutants decreased the antibiotic susceptibilities in association with ß-lactamase activity and antibiotic resistance-related gene expression. The results pointed out the cross-resistance of BIKP mutants to antibiotics, which might be considered when applying for the therapeutic use of bacteriophage.
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Antibacterianos/farmacologia , Bacteriófagos/fisiologia , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Klebsiella pneumoniae/efeitos dos fármacos , Klebsiella pneumoniae/virologia , Aminoglicosídeos/genética , Cefotaxima/farmacologia , Cefalotina/farmacologia , Cloranfenicol/farmacologia , Ciprofloxacina/farmacologia , Farmacorresistência Bacteriana Múltipla/genética , Regulação Bacteriana da Expressão Gênica , Humanos , Levofloxacino/farmacologia , Terapia por Fagos , Quinolonas/farmacologia , beta-Lactamases/genéticaRESUMO
We study the dynamics of cyclic competing mobile five species on spatially extended systems originated from asymmetric initial populations and investigate the basins for the three possible asymptotic states, coexistence of all species, existences of only two independent species, and the extinction. Through extensive numerical simulations, we find a prosperous dependence on initial conditions for species biodiversity. In particular, for fixed given equal densities of two relevant species, we find that only five basins for the existence of two independent species exist and they are spirally entangled for high mobility. A basin of coexistence is outbreaking when the mobility parameter is decreased through a critical value and surrounded by the other five basins. For fixed given equal densities of two independent species, however, we find that basin structures are not spirally entangled. Further, final states of two independent species are totally different. For all possible considerations, the extinction state is not witnessed which is verified by the survival probability. To provide the validity of basin structures from lattice simulations, we analyze the system in mean-field manners. Consequently, results on macroscopic levels are matched to direct lattice simulations for high mobility regimes. These findings provide a good insight into the fundamental issue of the biodiversity among many species than previous cases.
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Comportamento Competitivo , Teoria dos Jogos , Simulação por Computador , Probabilidade , Especificidade da Espécie , Análise de SobrevidaRESUMO
Traumatic brain injuries(TBI) pose significant challenges to human health, specifically neurological disorders and related motor activities. After TBI, the injured neuronal tissue is known for hardly regenerated and recovered to their normal neuron physiology and tissue compositions. For this reason, tissue engineering strategies that promote neuronal regeneration have gained increasing attention. This study explored the development of a novel neural tissue regeneration cryogel by combining brain-derived decellularized extracellular matrix (ECM) with heparin sulfate crosslinking that can perform nerve growth factor (NGF) release ability. Morphological and mechanical characterizations of the cryogels were performed to assess their suitability as a neural regeneration platform. After that, the heparin concnentration dependent effects of varying NGF concentrations on cryogel were investigated for their controlled release and impact on neuronal cell differentiation. The results revealed a direct correlation between the concentration of released NGF and the heparin sulfate ratio in cryogel, indicating that the cryogel can be tailored to carry higher loads of NGF with heparin concentration in cryogel that induced higher neuronal cell differentiation ratio. Furthermore, the study evaluated the NGF loaded cryogels on neuronal cell proliferation and brain tissue regeneration in vivo. The in vivo results suggested that the NGF loaded brain ECM derived cryogel significantly affects the regeneration of brain tissue. Overall, this research contributes to the development of advanced neural tissue engineering strategies and provides valuable insights into the design of regenerative cryogels that can be customized for specific therapeutic applications.
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Lesões Encefálicas Traumáticas , Engenharia Tecidual , Humanos , Encéfalo , Lesões Encefálicas Traumáticas/terapia , Criogéis , Matriz Extracelular , Heparina , Fator de Crescimento Neural/farmacologia , Regeneração Nervosa , Sulfatos , Engenharia Tecidual/métodosRESUMO
The tissue-specific heart decellularized extracellular matrix (hdECM) demonstrates a variety of therapeutic advantages, including fibrosis reduction and angiogenesis. Consequently, recent research for myocardial infarction (MI) therapy has utilized hdECM with various delivery techniques, such as injection or patch implantation. In this study, a novel approach for hdECM delivery using a wet adhesive paintable hydrogel is proposed. The hdECM-containing paintable hydrogel (pdHA_t) is simply applied, with no theoretical limit to the size or shape, making it highly beneficial for scale-up. Additionally, pdHA_t exhibits robust adhesion to the epicardium, with a minimal swelling ratio and sufficient adhesion strength for MI treatment when applied to the rat MI model. Moreover, the adhesiveness of pdHA_t can be easily washed off to prevent undesired adhesion with nearby organs, such as the rib cages and lungs, which can result in stenosis. During the 28 days of in vivo analysis, the pdHA_t not only facilitates functional regeneration by reducing ventricular wall thinning but also promotes neo-vascularization in the MI region. In conclusion, the pdHA_t presents a promising strategy for MI treatment and cardiac tissue regeneration, offering the potential for improved patient outcomes and enhanced cardiac function post-MI.
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Matriz Extracelular Descelularizada , Modelos Animais de Doenças , Hidrogéis , Infarto do Miocárdio , Ratos Sprague-Dawley , Animais , Ratos , Hidrogéis/química , Matriz Extracelular Descelularizada/química , Masculino , Matriz Extracelular/química , MiocárdioRESUMO
Axon regeneration and Schwann cell proliferation are critical processes in the repair and functional recovery of damaged neural tissues. Biomaterials can play a crucial role in facilitating cell proliferative processes that can significantly impact the target tissue repair. Chemical decellularization and supercritical fluid-based decellularization methods are similar approaches that eliminate DNA from native tissues for tissue-mimetic biomaterial production by using different solvents and procedures to achieve the final products. In this study, we conducted a comparative analysis of these two methods in the context of nerve regeneration and neuron cell differentiation efficiency. We evaluated the efficacy of each method in terms of biomaterial quality, preservation of extracellular matrix components, promotion of neuronal cell differentiation and nerve tissue repair ability in vivo. Our results indicate that while both methods produce high-quality biomaterials, supercritical fluid-based methods have several advantages over conventional chemical decellularization, including better preservation of extracellular matrix components and mechanical properties and superior promotion of cellular responses. We conclude that supercritical fluid-based methods show great promise for biomaterial production for nerve regeneration and neuron cell differentiation applications.
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Regeneração Nervosa , Tecido Nervoso , Matriz Extracelular/química , Axônios , Materiais Biocompatíveis/química , Engenharia Tecidual/métodos , Alicerces Teciduais/químicaRESUMO
In this study, blended perfluorinated sulfonic acid (PFSA) ionomers with equivalent weights (EWs, g/mol) of ~1000, 980, and 830 are prepared. Catalyst layers (CLs), using blended PFSA ionomers, with different side chain lengths and EWs are investigated and compared to CLs using single ionomers. The ion exchange capacity results confirm that blended ionomers have the target EWs. As a result, blended ionomers exhibit higher ion conductivity than single ionomers at all temperatures due to the higher water uptake of the blended ionomers. This implies that blended ionomers have a bulk structure to form a competent free volume compared to single ionomers. Blended ionomers with short side chains and low EWs can help reduce the activation energy in proton conduction due to enhanced hydrophobic and hydrophilic segregation. In addition, when using the blended ionomer, the CLs form a more porous microstructure to help reduce the resistance of oxygen transport and contributes to lower mass transfer loss. This effect is proven in fuel cell operations at not a lower temperature (70 °C) and full humidification (100%) but at an elevated temperature (80 °C) and lower relative humidity (50 and 75%). Blended ionomer-based CLs with a higher water uptake and porous CL structure result in improved fuel cell performance with better mass transport than single ionomer-based CLs.
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BACKGROUND: Patients with kidney failure must make complicated decisions about the dialysis modalities used either at home or in-hospital. Different options have varying levels of impact on patients' physical and psychological conditions and their social life. The purpose of this study was to evaluate the implementation of an intervention designed to achieve shared decision making (SDM) in patients' options for dialysis. METHODS: SDM was performed after consent was written for stage 5 chronic kidney disease patients before dialysis, and 435 cases were performed in 408 patients from December 16, 2019 to June 30, 2021. Among these, 101 patients were compared by SDM measurement scale, patient satisfaction, disease recognition scale survey, and dialysis method. RESULTS: The average age of participants was 56â years, with a gender composition of 55 males (54.5%) and 46 females (45.5%). Following SDM, the final dialysis methods decided upon by patients and clinicians were peritoneal dialysis (67 patients, 66.3%), hemodialysis (22 patients, 21.8%), and kidney transplantation (1 patient, 1.0%). CONCLUSIONS: Among participating patients, SDM was effective when used to decide on dialysis treatment, and patients were satisfied with the dialysis method decision process. On the disease awareness scale, those who participated in this project had relatively high positive and low negative perceptions, so it can be concluded that SDM was relatively effective. The implementation of SDM was helpful in selecting patients' best dialysis methods, and SDM scale results were higher in the peritoneal dialysis group than in the hemodialysis group.
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Falência Renal Crônica , Diálise Peritoneal , Masculino , Feminino , Humanos , Pessoa de Meia-Idade , Diálise Renal/métodos , Tomada de Decisão Compartilhada , Falência Renal Crônica/terapia , Falência Renal Crônica/psicologia , Inquéritos e Questionários , Tomada de Decisões , Participação do Paciente/métodosRESUMO
Melanin nanoparticles (MNPs) used for biomedical applications are often synthesized via the chemical auto-oxidation of catecholic monomers such as dopamine and 3,4-dihydroxyphenylalanine (DOPA) under alkaline conditions. However, the synthetic method for the chemical synthesis of MNP (cMNP) is relatively straightforward and more robust to control their homogenous particle size and morphology than the corresponding enzymatic synthetic methods. In this study, we demonstrated that the simple enzymatic synthesis of MNPs (eMNPs) with homogenous and soluble (<20 nm diameter) properties is possible using dopamine and Burkholderia cepacia tyrosinase (BcTy) under acidic conditions (i.e., pH 3.0). BcTy was highly reactive under pH 5.0, where the natural and chemical oxidation of catechol is complex, and thus melanin was synthesized via the hydroxylation of phenolic substrates. The detailed chemical analysis and characterization of the physical properties of the eMNPs confirmed the higher preservation of the catechol and primary amine moieties in the monomer substrate such as dopamine under acidic conditions. The eMNPs showed enhanced antioxidant activity and conferred stickiness to the formed hydrogel compared to the chemical auto-oxidation method owing to the large number of hydroxyl groups remaining such as catechol and quinone moieties. Because of these advantages and characteristics, the synthesis of MNPs using BcTy under acidic conditions can open a new path for their biomedical applications.
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Supercritical fluid-based extraction technologies are currently being increasingly utilized in high purity extract products for food industries. In recent years, supercritical fluid-based extraction technology is transformed in biomaterials process fields to be further utilized for tissue engineering and other biomedical applications. In particular, supercritical fluid-based decellularization protocols have great advantage over the conventional decellularization as it may allow preservation of extracellular matrix components and structures. In this review, the latest technological development utilizing the supercritical fluid-based decellularization for regenerative medicine is introduced.
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Matriz Extracelular , Medicina Regenerativa , Materiais Biocompatíveis , Matriz Extracelular/química , Medicina Regenerativa/métodos , Tecnologia , Engenharia Tecidual/métodos , Alicerces Teciduais/químicaRESUMO
Thermotherapy is an effective method for pain relief, recovery from injury, and general healthcare. The ordinary heat pad used for thermotherapy at home is not usually tailored to the individual but supplied in a few different pre-fixed sizes and shapes for mass marketing. A customized wearable heat pad often requires expert support. Herein, an instant, custom-fit, and on-demand heat pad for thermotherapy is demonstrated. The heater is directly printed using silver nanoparticle ink on an off-the-shelf medical grade tape by inkjet technology. By coating the tape with silica nanoparticles as ink-absorbing layer and chloride ions as chemical sintering agent, stable heater patterns are printed without the need for subsequent high temperature sintering process. A 3D scanner is used to acquire body information, and a customized heater is produced using the information. The printed heat pad is attached to the shoulder and the effect of thermotherapy is verified objectively through electroencephalography and subjectively through survey. This printed heat pad produced by simple and low-cost fabrication provides wearable medical devices for personal thermotherapy.
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Hipertermia Induzida , Nanopartículas Metálicas , Dispositivos Eletrônicos Vestíveis , Impressão Tridimensional , PrataRESUMO
An all three-dimensional (3D)-printed flexible ZnO ultraviolet (UV) photodetector is demonstrated, where the 3D-printing method is used not only for the electrode and photosensitive material but also for creating a substrate. An ultraflat and flexible substrate capable of serving as the backbone layer is developed using a water-dissolvable polymer layer for surface planarization. A two-layered printing followed by surface treatment is demonstrated for the substrate preparation. As mechanical support but flexible, a thick and sparse thermoplastic polyurethane layer is printed. On its surface, a thin and dense poly(vinyl alcohol) (PVA) is then printed. A precise control of PVA reflow using a microwater droplet results in a flexible and extremely uniform substrate. A Cu-Ag nanowire network is directly 3D printed on the flexible substrate for the conducting layer, followed by ZnO for the photosensitive material. Unlike the planar two-dimensional printing that provides thin films, 3D printing allows the electrode to have a step height, which can be made like a dam to accommodate a thick film of ZnO. Photosensitivity as a function of various ZnO thickness values was investigated to establish an optimal thickness for UV response. The device was also tested in natural sunlight along with stability and reliability.
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Óxido de Zinco/química , Impressão Tridimensional , Raios UltravioletaRESUMO
AIM: The doses of darbepoetin alfa required to maintain target haemoglobin levels after s.c. or i.v. administration when recombinant human erythropoietin (rHuEpo) treatment was replaced by darbepoetin alfa treatment in haemodialysis (HD) patients were compared. METHODS: In this prospective, randomized, open-label study, 65 HD patients who were receiving stable SC doses of rHuEpo were switched to an equivalent dose of darbepoetin alfa at a reduced frequency by s.c. or i.v. administration. Patients were randomly assigned to the s.c. group (n = 32) or the i.v. group (n = 33). Darbepoetin alfa doses were titrated to maintain target haemoglobin levels of 8.0-11.0 g/dL for up to 24 weeks. A period of 20 weeks was used for dose titration and haemoglobin stabilization. This was followed by a 4 week evaluation period. RESULTS: The mean haemoglobin concentration during the evaluation period was similar in the s.c. and i.v. groups. The mean dose and mean weight-standardized dose of darbepoetin alfa during the evaluation period tended to be lower in the s.c. group than the i.v. group, although these differences were not statistically significant. The mean weekly darbepoetin alfa dose requirements during the evaluation period significantly decreased in both groups compared to the dose requirements at randomization. CONCLUSION: There is a possibility that s.c. administration of darbepoetin alfa is more efficacious than i.v. administration, but a definite benefit cannot be demonstrated with the current sample size. A bigger sample size is needed to confirm the result.
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Anemia/tratamento farmacológico , Eritropoetina/análogos & derivados , Diálise Renal , Darbepoetina alfa , Eritropoetina/administração & dosagem , Hemoglobinas/análise , Humanos , Injeções Intravenosas , Injeções Subcutâneas , Estudos ProspectivosRESUMO
Profiling the transcriptomes of individual cells with single-cell RNA sequencing (scRNA-seq) has been widely applied to provide a detailed molecular characterization of cellular heterogeneity within a population of cells. Despite recent technological advances of scRNA-seq, technical variability of gene expression in scRNA-seq is still much higher than that in bulk RNA-seq. Accounting for technical variability is therefore a prerequisite for correctly analyzing single-cell data. This chapter describes a computational pipeline for detecting highly variable genes exhibiting higher cell-to-cell variability than expected by technical noise. The basic pipeline using the scater and scran R/Bioconductor packages includes deconvolution-based normalization, fitting the mean-variance trend, testing for nonzero biological variability, and visualization with highly variable genes. An outline of the underlying theory of detecting highly variable genes is also presented. We illustrate how the pipeline works by using two case studies, one from mouse embryonic stem cells with external RNA spike-ins, and the other from mouse dentate gyrus cells without spike-ins.
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RNA/genética , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , Animais , Biologia Computacional/métodos , Giro Denteado/fisiologia , Perfilação da Expressão Gênica/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Camundongos , Células-Tronco Embrionárias Murinas/fisiologia , Transcriptoma/genéticaRESUMO
A single walled carbon nanotube (SWCNT) based γ ray detector is demonstrated without a conventional scintillation mechanism. The change in the conductance of a two terminal SWCNT resistor in response to γ ray exposure is exploited as a sensing mechanism. Radiation-induced ambient oxygen dissociation and subsequent adsorption of oxygen species on the SWCNT surface alter its electrical properties. The responses to the total dose and dose rate are investigated along with the sensing mechanism. The detector showed good sensitivity to γ ray and a capability to distinguish radiation dose rates ranging from 2.4 to 16.4 R/min.
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Raios gama , Nanotubos de Carbono/química , Radiometria/métodos , Adsorção , Técnicas Eletroquímicas/instrumentação , Técnicas Eletroquímicas/métodos , Microcomputadores , Ozônio/química , Radiometria/instrumentaçãoRESUMO
A fabric-compatible UV sensor is presented using a cellulose-based thread coated with single-wall carbon nanotube ink. Two-terminal resistive responses of the thread were measured upon exposure to UV, and the effects of intensity, wavelength, and on/off cycling were studied. The sensor was tested in the field under direct sunlight, demonstrating practical usability for a wearable/flexible UV sensor system. The results here confirm the potential for an inexpensive wearable sensor in contrast to the conventional rigid and bulky solid-state detectors.
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Successful transition to commercialization and practical implementation of nanotechnology innovations may very well need device designs that are tolerant to the inherent variations and imperfections in all nanomaterials including carbon nanotubes, graphene, and others. As an example, a single-walled carbon nanotube network based gas sensor is promising for a wide range of applications such as environment, industry, and biomedical and wearable devices due to its high sensitivity, fast response, and low power consumption. However, a long-standing issue has been the production of extremely high purity semiconducting nanotubes, thereby contributing to the delay in the market adoption of those sensors. Inclusion of even less than 0.1% of metallic nanotubes, which is inevitable, is found to result in a significant deterioration of sensor-to-sensor uniformity. Acknowledging the coexistence of metallic and semiconducting nanotubes as well as all other possible imperfections, we herein present a novel variation-tolerant sensor design where the sensor response is defined by a statistical Gaussian measure in contrast to a traditional deterministic approach. The single input and multiple output data are attained using multiport electrodes fabricated over a relatively large area single nanotube ensemble. The data processing protocol discards outlier data points and the origin of the outliers is investigated. Both the experimental demonstration and complementary analytical modeling support the hypothesis that the statistical analysis of the device can strengthen the credibility of the sensor constructed using nanomaterials with any imperfections. The proposed strategy can also be applied to physical, radiation, and biosensors as well as other electronic devices.
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Técnicas Eletroquímicas/métodos , Nanotubos de Carbono/química , Amônia/análise , Interpretação Estatística de Dados , Técnicas Eletroquímicas/instrumentação , Eletrodos , Gases/análiseRESUMO
BACKGROUND: Leukoaraiosis is a term used to define the abnormal appearance of subcortical white matter of the brain by means of neuroimaging and is regarded as an intermediate surrogate of stroke. The goal of this study is to identify the prevalence of leukoaraiosis and analyze predictors of risk of leukoaraiosis. STUDY DESIGN: Cross-sectional study. SETTING & PARTICIPANTS: 57 peritoneal dialysis (PD) patients without diabetes treated in 3 academic medical-associated dialysis units who did not have a history of cerebrovascular disease or neurological symptoms compared with a convenience sample of 57 age- and sex-matched hypertensive control subjects with normal renal function. PREDICTOR: End-stage renal disease treated by PD compared with hypertension, adjusted for clinical and laboratory characteristics. OUTCOME & MEASUREMENT: Hyperintense areas on magnetic resonance imaging T2 high-signal intensity scoring system. RESULTS: The prevalence of leukoaraiosis was significantly greater in patients on PD therapy than controls (68.4% versus 17.5%; P < 0.001). High T2 signal intensity score in patients on PD therapy compared with controls was significantly higher in the anterior circulation of the brain, relatively sparing the posterior fossa. End-stage renal disease, age, and poor control of blood pressure were significant independent predictors of leukoaraiosis. LIMITATIONS: There is the possibility that biases regarding the selection of enrolled patients had an influence on a study result. CONCLUSIONS: Cerebral magnetic resonance imaging of PD patients without evidence of cerebrovascular disease showed a high prevalence of leukoaraiosis in the anterior circulation of the brain. Old age, poorly controlled hypertension, and the PD procedure itself and/or end-stage renal disease seem to be associated with the presence of leukoaraiosis.
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Encéfalo/patologia , Falência Renal Crônica/patologia , Leucoaraiose/patologia , Diálise Peritoneal/efeitos adversos , Adulto , Idoso , Artérias Carótidas/diagnóstico por imagem , Estudos Transversais , Feminino , Humanos , Hipertensão/complicações , Hipertensão/patologia , Falência Renal Crônica/sangue , Falência Renal Crônica/complicações , Falência Renal Crônica/terapia , Leucoaraiose/epidemiologia , Leucoaraiose/etiologia , Modelos Logísticos , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Prevalência , Fatores de Risco , UltrassonografiaRESUMO
Carbon nanotube (CNT) is a promising candidate as a sensor material for the sensitive detection of gases/vapors, biomarkers, and even some radiation, as all these external variables affect the resistance and other properties of nanotubes, which forms the basis for sensing. Ultraviolet (UV) radiation does not impact the nanotube properties given the substantial mismatch of bandgaps and therefore, CNTs have never been considered for UV sensing, unlike the popular ZnO and other oxide nanwires. It is well-known that UV assists the adsorption/desorption characteristics of oxygen on carbon nanotubes, which changes the nanotube resistance. Here, we demonstrate a novel sensor structure encapsulated with an air pocket, where the confined air is responsible for the UV sensing mechanism and assures sensor stability and repeatability over time. In addition to the protection from any contamination, the air pocket encapsulated sensor offers negligible baseline drift and fast recovery compared to previously reported sensors. The air pocket isolated from the outside environment can act as a stationary oxygen reservoir, resulting in consistent sensor characteristics. Furthermore, this sensor can be used even in liquid environments.
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Ar , Técnicas de Química Analítica/instrumentação , Nanotubos de Carbono/química , Raios Ultravioleta , CápsulasRESUMO
The denitrification of nitrate (NO3(-)) by mixed cultures in the presence of zero-valent iron [Fe(0)] and biochar was investigated through a series of batch experiments. It was hypothesized that biochar may provide microbes with additional electrons to enhance the anaerobic biotransformation of nitrate in the presence of Fe(0) by facilitating electron transfer. When compared to the anaerobic transformation of nitrate by microbes in the presence of Fe(0) alone, the presence of biochar significantly enhanced anaerobic denitrification by microbes with Fe(0). Graphite also promoted the anaerobic microbial transformation of nitrate with Fe(0), and it was speculated that electron-conducting graphene moieties were responsible for the improvement. The results obtained in this work suggest that nitrate can be effectively denitrified by microbes with Fe(0) and biochar in natural and engineered systems.
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Bactérias/metabolismo , Carvão Vegetal/farmacologia , Desnitrificação/efeitos dos fármacos , Ferro/farmacologia , Nitratos/metabolismo , Bactérias/efeitos dos fármacos , Biodegradação Ambiental/efeitos dos fármacos , TemperaturaRESUMO
BACKGROUND AND PURPOSE: Hepatitis C virus (HCV) infection is responsible for various chronic inflammatory liver diseases. Here, we have identified a naturally occurring compound with anti-HCV activity and have elucidated its mode of antiviral action. EXPERIMENTAL APPROACH: Luciferase reporter and real-time RT-PCR assays were used to measure HCV replication. Western blot, fluorescence-labelled HCV replicons and infectious clones were employed to quantitate expression levels of viral proteins. Resistant HCV mutant mapping, in vitro NS3 protease, helicase, NS5B polymerase and drug affinity responsive target stability assays were also used to study the antiviral mechanism. KEY RESULTS: A resveratrol tetramer, vitisin B from grapevine root extract showed high potency against HCV replication (EC50 = 6 nM) with relatively low cytotoxicity (EC50 >10 µM). Combined treatment of vitisin B with an NS5B polymerase inhibitor (sofosbuvir) exhibited a synergistic or at least additive antiviral activity. Analysis of a number of vitisin B-resistant HCV variants suggested an NS3 helicase as its potential target. We confirmed a direct binding between vitisin B and a purified NS3 helicase in vitro. Vitisin B was a potent inhibitor of a HCV NS3 helicase (IC50 = 3 nM). In vivo, Finally, we observed a preferred tissue distribution of vitisin B in the liver after i.p. injection in rats, at clinically attainable concentrations. Conclusion and Implications Vitisin B is one of the most potent HCV helicase inhibitors identified so far. Vitisin B is thus a prime candidate to be developed as the first HCV drug derived from natural products.