RESUMEN

In some locations around the globe, the U concentrations may exceed WHO standards by 2-folds therefore, effective yet environmentally wise solutions to purify radioactive waters are of significant importance. Here, the optimized and fully controlled coal-fly-ash based Na-P1 zeolite functionalization by employing novel, biodegradable biosurfactant molecule - cocamidopropyl betaine (CAPB) is showcased. The zeolite's surface decoration renders three composites with varying amounts of introduced CAPB molecule (Na-P1 @ CAPB), with 0.44, 0.88, and 1.59-times External Cation Exchange Capacity (ECEC). Wet-chemistry experiments revealed extremely high U adsorption capacity (qmax = 137.1 mg U/g) unveiling preferential interactions of uranyl dimers with CAPB molecules coupled with ion-exchange between Na+ ions. Multimodal spectroscopic analyses, including Fourier-Transformed Infra-Red (FT-IR), X-ray Photoelectron (XPS), and X-ray Absorption Fine Structure (XAFS), showed the hexavalent oxidation state of U, and no secondary release of the CAPB molecule from the composite. The EXAFS signals fingerprint changes in the interatomic distances of adsorbed U, showing the impact of the O and N, heteroatoms present in the CAPB molecule on U binding mechanism. The presented research outcomes showcase the easy, scalable, optimized, and environmentally friendly synthesis of biofunctional zeolite effectively purifying the real-life U-bearing wastewaters from the vicinity of the Pribram deposit (Czech Republic).

RESUMEN

This study investigates the immobilization mechanisms of heavy metal ions in the C-S-H phase. Synthetic C-S-H phases were prepared via the precipitation method, incorporating five different ions (Pb(II), Cd(II), Ni(II), Zn(II), and Cr(III)). Structural analysis of the obtained material was conducted using vibrational spectroscopy (both FT-IR and Raman), X-ray photoelectron spectroscopy, and X-ray diffraction. Spectroscopic methods were primarily employed to evaluate the structural effects and polymerization degree of the resulting C-S-H phase. Morphological changes were characterized using scanning and transmission electron microscopy (SEM and TEM, respectively). Our findings reveal several mechanisms for immobilizing heavy metal cations: precipitation of insoluble compounds (particularly notable for Ni(II) and Cr(III)), replacement of Ca(II) ions within the silicate structure (evident in the crystallization of Ca(OH)2 in samples containing Cd(II), Ni(II), and Zn(II) in minimal quantities), and strong bonding of certain metals (such as Pb(II)) with the C-S-H phase structure. These insights contribute to understanding the potential applications of C-S-H phases in heavy metal immobilization.

RESUMEN

Uranium, a key member of the actinides series, is radioactive and may cause severe environmental hazards once discharged into the water due to high toxicity. Removal of uranium via adsorption by applying tailored, functional adsorbents is at the forefront of tackling such pollution. Here, we report the optimized functionalization of the powder coal fly-ash (CFA) derived Na-P1 synthetic zeolite to the form of granules by employing the biodegradable polymer-calcium alginate (CA) and their application to remove aqueous U. The optimized synthesis showed that granules are formed at the CA concentration equals to 0.5 % wt., and that application of 1% wt. solution renders the most effective U scavengers. The maximum U adsorption capacity (qmax) increases significantly after CA modification from 44.48 mgU/g for native, powder Na-P1 zeolite to 62.53 mg U/g and 76.70 mg U/g for 0.5 % wt. and 1 % wt. CA respectively. The U adsorption follows the Radlich-Peterson isotherm model, being the highest at acidic pH (pHeq∼4). The U adsorption kinetics reveals swift U uptake, reaching equilibrium after 2h for 1 % ZACB and 3 h for 0.5 % wt. ZACB following the pseudo-second-order (PSO) kinetic model. SEM-EDXS investigation elucidates that adsorbed U occurs onto materials as an inhomogenous, well-dispersed, and micrometer-scale aggregate. Further, XPS and µ-XRF spectroscopies complementarily confirmed the hexavalent oxidation state of adsorbed U and its altered distribution on ZACBs with varying CA concentrations. U distribution was probed "in-situ" onto materials while correlations between the major elements (Al, Si, Ca, U) contributing to U scavenging were calculated and compared. Finally, a real-life coal mine wastewater (CMW) polluted by 238U and 228,226Ra was successfully purified, satisfying WHO guidelines after treatment using ZACBs. These findings offer new insights on successful yet optimized Na-P1 zeolite modification using biodegradable polymer (Ca2+-exchanged alginate) aimed at efficient U removal, displaying a near-zero environmental impact.

Asunto(s)

Uranio , Zeolitas , Zeolitas/química , Intercambio Iónico , Polvos , Iones , Cinética , Sodio/química , Adsorción , Carbón Mineral , Polímeros , Concentración de Iones de Hidrógeno

RESUMEN

Density functional theory (DFT) calculations and x-ray diffraction techniques were employed to evaluate the value of the tilt angle in ferroelectric smectic C^{*} and antiferroelectric smectic C_{A}^{*} phases. Five homologues from the chiral series denoted as 3FmHPhF6(m=2,4,5,6,7), based on 4-(1-methylheptyloxycarbonyl) phenyl 4'-octyloxybiphenyl-4-carboxylate (MHPOBC), were studied. Two types of conformations for the nonchiral terminal chain (fully extended and gauche) and three types of deviation from the rodlike shape of the molecules (hockey stick, zigzag, and C shape) were computationally considered. The nonlinear shape of the molecules was accounted for by introducing a shape parameter δΘ. We observe that calculations of the tilt angle which consider the C-shaped structures, in both the fully extended or gauche conformations, lead to good agreement with the values of the tilt angle obtained from electro-optical measurements below the saturation temperature. The results allow us to conclude that such structures are adopted by molecules in the examined series of smectogens. Additionally, this study proves the presence of the standard orthogonal SmA^{*} phase for the homologues with m=6, 7, and the de Vries SmA^{*} phase for m=5.

RESUMEN

Lead halides in an asymmetric layered structure form memristive devices which are controlled by the electronic structure of the PbX2|metal interface. In this paper, we explain the mechanism that stands behind the I- V pinched hysteresis loop of the device and shortly present its synaptic-like plasticity (spike-timing-dependent plasticity and spike-rate-dependent plasticity) and nonvolatile memory effects. This memristive element was incorporated into a reservoir system, in particular, the echo-state network with delayed feedback, which exhibits brain-like recurrent behavior and demonstrates metaplasticity as one of the available learning mechanisms. It can serve as a classification system that classifies input signals according to their amplitude.

RESUMEN

The studies of low-temperature immobilization of bound water in Antarctic lichenized fungus Turgidosculum complicatulum were performed using 1H NMR and DSC over a wide range of thallus hydration. 1H NMR free induction decays were decomposed into a solid component well described by the Gaussian function and two exponentially decaying components coming from a tightly bound water and from a loosely bound water fraction. 1H NMR spectra revealed one averaged mobile proton signal component. 1H NMR measurements recorded in time and in frequency domain suggest the non-cooperative bound water immobilization in T. complicatulum thallus. The threshold of the hydration level estimated by 1H NMR analysis at which the cooperative bound water freezing was detected was Δm/m0 ≈ 0.39, whereas for DSC analysis was equal to Δm/m0 = 0.375. Main ice melting estimated from DSC measurements for zero hydration level of the sample starts at tm = -(19.29 ± 1.19)°C. However, DSC melting peak shows a composed form being a superposition of the main narrow peak (presumably melting of mycobiont areas) and a broad low-temperature shoulder (presumably melting of isolated photobiont cells). DSC traces recorded after two-hour incubation of T. complicatulum thallus at -20 °C suggest much lower threshold level of hydration at which the ice formation occurs (Δm/m0 = 0.0842). Presumably it is a result of diffusion induced migration of separated water molecules to ice microcrystallites already present in thallus, but still beyond the calorimeter resolution.

Asunto(s)

Ascomicetos/metabolismo , Chlorophyta/metabolismo , Frío , Microbiología del Agua , Agua/química , Regiones Antárticas , Rastreo Diferencial de Calorimetría , Congelación , Espectroscopía de Resonancia Magnética , Termodinámica

RESUMEN

BACKGROUND: Psoriasis has been shown to increase cardiovascular risk, and a contributor to this might be enhanced myocardial fibrosis promoted by the disease-associated pro-inflammatory milieu. OBJECTIVE: We sought to investigate the relationship of galectin-3 (Gal-3) - a recognized mediator of fibrosis with inflammatory activation and left ventricular (LV) systolic and diastolic function in patients with psoriasis. METHODS: We enrolled 102 psoriatic patients (mean age: 52.5 ± 12.6 years). Sixty-five age- and sex-matched healthy subjects served as controls. Echocardiographic assessment of myocardial function included estimation of LV longitudinal systolic deformation (GLS) and diastolic indices: tissue e' velocity and E/e' ratio. Laboratory measurements encompassed blood Gal-3, creatinine, glucose, insulin, CRP and erythrocyte sedimentation rate (ESR). RESULTS: Patients with psoriasis were characterized by elevated Gal-3 (12.3 [9.3-13.4] vs. 6.3 [5.5-9.4] ng/mL in healthy controls, P < 0.001), ESR (17.0 [11.0-29.0] vs. 8.5 [6.0-13.0] mm, respectively, P < 0.001) and CRP (3.1 [1.7-10.6] vs. 1.9 [1.5-4.0] mg/L, respectively, P < 0.001), and reduced GLS (19.9 ± 3.7 vs. 22.0 ± 3.0%, respectively, P < 0.001). Progressive deterioration of GLS was demonstrated across Gal-3 tertiles. Significant associations between GLS and age (beta = -0.21, P < 0.04), Gal-3 (beta = -0.27, P < 0.01), CRP (beta = -0.22, P < 0.03), ESR (beta = -0.25, P < 0.01), waist circumference (beta = -0.22, P < 0.03) and waist-to-hip ratio (beta = -0.20, P < 0.05) were found. Stepwise multiple regression analysis revealed that the independent determinants of GLS in psoriatic patients were Gal-3 (beta = -0.24, P < 0.01) and ESR (beta = -0.21, P < 0.03). Regression-based mediation analysis demonstrated that the relationship between ESR and GLS was partially mediated by Gal-3. CONCLUSIONS: Subclinical left ventricular systolic dysfunction in psoriasis, as evidenced by reduced GLS, is linked with the inflammatory upregulation, and enhanced profibrotic activity (as reflected by elevated serum Gal-3) may be involved in this process. These putative mechanisms may be responsible for the observed higher incidence of heart failure in this disease condition and should be considered as a potential target for preventive and therapeutic measures.

Asunto(s)

Galectina 3/sangre , Psoriasis/sangre , Disfunción Ventricular Izquierda/sangre , Disfunción Ventricular Izquierda/diagnóstico por imagen , Adulto , Anciano , Enfermedades Asintomáticas , Proteínas Sanguíneas , Sedimentación Sanguínea , Proteína C-Reactiva/metabolismo , Estudios de Casos y Controles , Diástole , Ecocardiografía , Femenino , Galectinas , Humanos , Masculino , Persona de Mediana Edad , Sístole , Disfunción Ventricular Izquierda/fisiopatología , Circunferencia de la Cintura , Relación Cintura-Estatura

RESUMEN

The purpose of this paper is to review recent developments on polyurethanes aimed at the design, synthesis, modifications, and biological properties in the field of bone tissue engineering. Different polyurethane systems are presented and discussed in terms of biodegradation, biocompatibility and bioactivity. A comprehensive discussion is provided of the influence of hard to soft segments ratio, catalysts, stiffness and hydrophilicity of polyurethanes. Interaction with various cells, behavior in vivo and current strategies in enhancing bioactivity of polyurethanes are described. The discussion on the incorporation of biomolecules and growth factors, surface modifications, and obtaining polyurethane-ceramics composites strategies is held. The main emphasis is placed on the progress of polyurethane applications in bone regeneration, including bone void fillers, shape memory scaffolds, and drug carrier.

Asunto(s)

Poliuretanos/química , Regeneración Ósea , Cerámica , Porosidad , Ingeniería de Tejidos

RESUMEN

A new method for direct covalent immobilization of protein molecules (including antibodies) on organic polymers with plasma-induced random micronanoscale topography and stable-in-time chemical functionality is presented. This is achieved using a short (1-5 min) plasma etching and simultaneous micronanotexturing process, followed by a fast thermal annealing step, which induces accelerated hydrophobic recovery while preserving important chemical functionality created by the plasma. Surface-bound biomolecules resist harsh washing with sodium dodecyl sulfate and other detergents even at elevated temperatures, losing less than 40% of the biomolecules bound even at the harshest washing conditions. X-ray photoelectron spectroscopy, secondary-ion mass spectrometry, and electron paramagnetic resonance are used to unveil the chemical modification of the plasma-treated and stabilized surfaces. The nanotextured and chemically stabilized surfaces are used as substrates for the development of immunochemical assays for the sensitive detection of C-reactive protein and salmonella lipopolysaccharides through immobilization of the respective analyte-specific antibodies onto them. Such substrates are stable for a period of 1 year with ambient storage.

Asunto(s)

Anticuerpos/química , Técnicas Biosensibles/instrumentación , Inmunoensayo/instrumentación , Membranas Artificiales , Impresión Molecular/métodos , Nanopartículas/química , Adsorción , Anticuerpos/inmunología , Diseño de Equipo , Análisis de Falla de Equipo , Ensayo de Materiales , Nanopartículas/ultraestructura , Gases em Plasma/química , Unión Proteica , Propiedades de Superficie

RESUMEN

The aim of tissue engineering is the fabrication of three-dimensional scaffolds that can be used for the reconstruction and regeneration of damaged or deformed tissues and organs. A wide variety of techniques have been developed to create either fibrous or porous scaffolds from polymers, metals, composite materials and ceramics. However, the most promising materials are biodegradable polymers due to their comprehensive mechanical properties, ability to control the rate of degradation and similarities to natural tissue structures. Polyurethanes (PUs) are attractive candidates for scaffold fabrication, since they are biocompatible, and have excellent mechanical properties and mechanical flexibility. PU can be applied to various methods of porous scaffold fabrication, among which are solvent casting/particulate leaching, thermally induced phase separation, gas foaming, emulsion freeze-drying and melt moulding. Scaffold properties obtained by these techniques, including pore size, interconnectivity and total porosity, all depend on the thermal processing parameters, and the porogen agent and solvents used. In this review, various polyurethane systems for scaffolds are discussed, as well as methods of fabrication, including the latest developments, and their advantages and disadvantages.

Asunto(s)

Plásticos Biodegradables , Poliuretanos , Andamios del Tejido/química , Animales , Plásticos Biodegradables/química , Plásticos Biodegradables/farmacología , Humanos , Poliuretanos/química , Poliuretanos/farmacología , Porosidad

RESUMEN

Protein interactions with surfaces of promising conducting polymers are critical for development of bioapplications. Surfaces of spin-cast and postbaked poly(3-alkylthiophenes), regiorandom P3BT, and regioregular RP3HT are examined prior to and after adsorption of model protein, bovine serum albumin, with time-of-flight secondary ion mass spectrometry, atomic force microscopy, and X-ray photoelectron spectroscopy. The multivariate method of principal component analysis applied to ToF-SIMS data maximizes information on subtle differences in surface chemistry: PCA reveals alkyl side chains and conjugated backbones, exposed for RP3HT and P3BT, respectively. Phase imaging AFM shows semicrystalline microstructure of RP3HT and amorphous morphology of P3BT films. A cellular-like pattern of proteins adsorbed on RP3HT develops with coverage to more uniform overlayer, observed always on P3BT. The amount of adsorbed protein, determined by XPS as a function of BSA concentration (up to 10 mg/mL), is ∼21% lower for RP3HT than P3BT (up to 1.1 mg/m(2)). Although PCA differentiates protein from polythiophene, relative protein surface composition evaluated from ToF-SIMS saturates rather than increases with amount of adsorbed BSA from XPS. This reflects ToF-SIMS sensitivity to outermost layer of proteins, enabling multivariate analysis of protein conformation or orientation. PCA distinguishes between amino acids characteristic for external regions of BSA adsorbed to P3BT and RP3HT. These amino acids are identified for P3BT and RP3HT as hydrophilic and hydrophobic, respectively, by relative hydrophobicity of amino acid side chains. Alternative identification with BSA domains fails, pointing to substrate-induced changes in conformation and degree of denaturation rather than orientation of adsorbed protein.

Asunto(s)

Membranas Artificiales , Polímeros/química , Albúmina Sérica Bovina/química , Tiofenos/química , Adsorción , Animales , Bovinos , Estructura Terciaria de Proteína , Propiedades de Superficie

RESUMEN

It is well known that the process of plant cell differentiation depends on the symplasmic isolation of cells. Before starting the differentiation programme, the individual cell or group of cells should restrict symplasmic communication with neighbouring cells. We tested the symplasmic communication between epidermal cells in the different root zones of parental barley plants Hordeum vulgare L., cv. 'Karat' with normal root hair development, and two root hairless mutants (rhl1.a and rhl1.b). The results clearly show that symplasmic communication was limited during root hair differentiation in the parental variety, whereas in both root hairless mutants epidermal cells were still symplasmically connected in the corresponding root zone. This paper is the first report on the role of symplasmic isolation in barley root cell differentiation, and additionally shows that a disturbance in the restriction of symplasmic communication is present in root hairless mutants.

Asunto(s)

Diferenciación Celular , Hordeum/citología , Epidermis de la Planta/citología , Proteínas de Plantas/metabolismo , Raíces de Plantas/citología , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Diferenciación Celular/genética , Proteínas de Unión al ADN , Genes de Plantas , Hordeum/genética , Hordeum/crecimiento & desarrollo , Mutación , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Permeabilidad , Epidermis de la Planta/crecimiento & desarrollo , Proteínas de Plantas/genética , Raíces de Plantas/crecimiento & desarrollo

RESUMEN

To provide complete characterization of immunoassay on silicon biosensor surfaces, atomic force microscopy, (angle-resolved) X-ray photoelectron spectroscopy and time-of-flight secondary ion mass spectrometry were applied to examine Si(3)N(4) surfaces modified with (3-aminopropyl)triethoxysilane, coated with gamma globulins (IgG), blocked with bovine serum albumin and then reacted with anti-IgG antibody for two complementary pairs (rabbit and mouse IgG) at various concentrations (from 0.3 nM to 330 nM). Protein coverage, as reflected in (amine to total N1s) XPS signal ratio and determined from ARXPS, decreases slightly due to blocking and then increases monotonically for anti-IgG antibody concentrations higher than 1 nM. AFM images reveal hardly any change of lateral nanostructure due to blocking but response to antibody solutions, based on both the mean size (from autocorrelation) and dominant spacing (from Fourier analysis) of surface features, similar to that given by ARXPS. AFM height histograms provided information about the vertical nanostructure and the parameters of height distribution (average height, spread - roughness and skewness) were distinctly influenced by coating, blocking and immunoreaction. Average protein layer thickness values determined based on protein structure (molecular weight, dimensions) and surface coverage provided from ARXPS were in accord with average height of protein layer determined from AFM. TOF-SIMS analysis indicated that BSA blocks free surface sites and in addition replaces some already adsorbed IgGs.

Asunto(s)

Inmunoensayo/métodos , Nanoestructuras/química , Albúmina Sérica Bovina/análisis , Silicio/química , Animales , Bovinos , Ratones , Microscopía de Fuerza Atómica , Espectroscopía de Fotoelectrones , Conejos , Propiedades de Superficie

RESUMEN

The mechanisms of malignant cell transformation mediated by the oncogenic anaplastic lymphoma kinase (ALK) tyrosine kinase remain only partially understood. In this study, we report that T-cell lymphoma (TCL) cells carrying the nucleophosmin (NPM)/ALK fusion protein (ALK+ TCL) strongly express hypoxia-induced factor 1α (HIF1α) mRNA, even under normoxic conditions, and markedly upregulate HIF1α protein expression under hypoxia. HIF1α expression is strictly dependent on the expression and enzymatic activity of NPM/ALK, as shown in BaF3 cells transfected with wild-type NPM/ALK and kinase-inactive NPM/ALK K210R mutant and by the inhibition of the NPM/ALK function in ALK+ TCL cells by a small-molecule ALK inhibitor. NPM/ALK induces HIF1α expression by upregulating its gene transcription through its key signal transmitter signal transducer and activator of transcription 3 (STAT3), which binds to the HIF1α gene promoter as shown by the chromatin immunoprecipitation assay and is required for HIF1α gene expression as demonstrated by its small interfering RNA-mediated depletion. In turn, depletion of HIF1α increases mammalian target of rapamycin complex 1 activation, cell growth and proliferation and decreases vascular endothelial growth factor synthesis. These results identify a novel cell-transforming property of NPM/ALK, namely its ability to induce the expression of HIF1α, a protein with an important role in carcinogenesis. These results also provide another rationale to therapeutically target NPM/ALK and STAT3 in ALK+ TCL.

Asunto(s)

Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Proteínas Nucleares/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Fosfotransferasas/metabolismo , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/metabolismo , Proliferación Celular , Transformación Celular Neoplásica , Regulación Neoplásica de la Expresión Génica , Humanos , Hipoxia , Linfoma de Células T/genética , Linfoma de Células T/metabolismo , Linfoma de Células T/patología , Nucleofosmina , Proteínas de Fusión Oncogénica/antagonistas & inhibidores , Proteínas de Fusión Oncogénica/genética , Proteínas Tirosina Quinasas/genética , ARN Mensajero/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Transducción de Señal/fisiología

RESUMEN

BACKGROUND: Coronary heart disease patients and end-stage renal disease patients have been documented to have an increased amount of coronary artery calcifications (CAC). PURPOSE: To evaluate the distribution of CAC and its influence on interscan variability of measurement in end-stage renal disease and coronary heart disease patients, proven to have calcifications. MATERIAL AND METHODS: 69 patients having CAC, including 34 with coronary heart disease and 35 with end-stage renal disease, were scanned twice with multidetector-row computed tomography (MDCT). Amount of CAC was determined as the number of calcified lesions (CN), total calcium score (CS), calcium volume (CV), and calcium mass (CM). Distribution of CAC was evaluated on a per-patient basis as the median CS and CM of a single lesion. Density of the calcifications was calculated as the patient's CM divided by CV. RESULTS: The overall median CS was 457.2, and the median CM was 75.6 mg. There were no significant differences in the number of calcified lesions, CS, or CM between the two groups. Both CS and CM of a single lesion, as well as the mean calcium density were lower in renal disease patients (P<0.05) than in coronary heart disease subjects. The relative interscan variability of coronary calcium measurement was higher in the renal disease group (P<0.05). There was a negative correlation between the calcium concentration and the relative interscan variability. CONCLUSION: The results indicate that the coronary calcium distribution influences the measurement interscan reproducibility, and the distribution may differ between end-stage renal disease patients and coronary heart disease patients, reflecting the dissimilar nature of coronary calcifications in those groups.

Asunto(s)

Angiografía Coronaria/estadística & datos numéricos , Enfermedad de la Arteria Coronaria/diagnóstico por imagen , Procesamiento de Imagen Asistido por Computador/estadística & datos numéricos , Fallo Renal Crónico/diagnóstico por imagen , Tomografía Computarizada Espiral/estadística & datos numéricos , Adulto , Análisis de Varianza , Calcio/metabolismo , Progresión de la Enfermedad , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Reproducibilidad de los Resultados , Programas Informáticos

RESUMEN

The mechanisms of cell transformation mediated by the nucleophosmin (NPM)/anaplastic lymphoma kinase (ALK) tyrosine kinase are only partially understood. Here, we report that cell lines and native tissues derived from the NPM/ALK-expressing T-cell lymphoma display persistent activation of mammalian target of rapamycin (mTOR) as determined by phosphorylation of mTOR targets S6rp and 4E-binding protein 1 (4E-BP1). The mTOR activation is serum growth factor-independent but nutrient-dependent. It is also dependent on the expression and enzymatic activity of NPM/ALK as demonstrated by cell transfection with wild-type and functionally deficient NPM/ALK, small interfering RNA (siRNA)-mediated NPM/ALK depletion and kinase activity suppression using the inhibitor WHI-P154. The NPM/ALK-induced mTOR activation is transduced through the mitogen-induced extracellular kinase (MEK)/extracellular signal-regulated kinase (ERK) signaling pathway and, to a much lesser degree, through the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway. Accordingly, whereas the low-dose PI3K inhibitor wortmannin and Akt inhibitor III profoundly inhibited Akt phosphorylation, they had a very modest effect on S6rp and 4E-BP1 phosphorylation. In turn, MEK inhibitors U0126 and PD98059 and siRNA-mediated depletion of either ERK1 or ERK2 inhibited S6rp phosphorylation much more effectively. Finally, the mTOR inhibitor rapamycin markedly decreased proliferation and increased the apoptotic rate of ALK+TCL cells. These findings identify mTOR as a novel key target of NPM/ALK and suggest that mTOR inhibitors may prove effective in therapy of ALK-induced malignancies.

Asunto(s)

Proteínas Nucleares/metabolismo , Proteínas Quinasas/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Transducción de Señal/efectos de los fármacos , Sirolimus/farmacología , Quinasa de Linfoma Anaplásico , Animales , Western Blotting , Línea Celular , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Quinasas MAP Reguladas por Señal Extracelular/genética , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Inmunohistoquímica , Linfoma de Células T/genética , Linfoma de Células T/metabolismo , Linfoma de Células T/patología , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Proteínas Quinasas Activadas por Mitógenos/genética , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Modelos Biológicos , Proteínas Nucleares/genética , Nucleofosmina , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatidilinositol 3-Quinasas/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3 , Inhibidores de Proteínas Quinasas , Proteínas Quinasas/genética , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/genética , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Quinazolinas/farmacología , ARN Interferente Pequeño/genética , Proteínas Tirosina Quinasas Receptoras , Serina-Treonina Quinasas TOR , Transfección

RESUMEN

The mechanisms of cell transformation mediated by the highly oncogenic, chimeric NPM/ALK tyrosine kinase remain only partially understood. Here we report that cell lines and native tissues derived from the NPM/ALK-expressing T-cell lymphoma (ALK+ TCL) display phosphorylation of the extracellular signal-regulated protein kinase (ERK) 1/2 complex. Transfection of BaF3 cells with NPM/ALK induces phosphorylation of EKR1/2 and of its direct activator mitogen-induced extracellular kinase (MEK) 1/2. Depletion of NPM/ALK by small interfering RNA (siRNA) or its inhibition by WHI-154 abrogates the MEK1/2 and ERK1/2 phosphorylation. The NPM/ALK-induced MEK/ERK activation is independent of c-Raf as evidenced by the lack of MEK1/2 and ERK1/2 phosphorylation upon c-Raf inactivation by two different inhibitors, RI and ZM336372, and by its siRNA-mediated depletion. In contrast, ERK1/2 activation is strictly MEK1/2 dependent as shown by suppression of the ERK1/2 phosphorylation by the MEK1/2 inhibitor U0126. The U0126-mediated inhibition of ERK1/2 activation impaired proliferation and viability of the ALK+ TCL cells and expression of antiapoptotic factor Bcl-xL and cell cycle-promoting CDK4 and phospho-RB. Finally, siRNA-mediated depletion of both ERK1 and ERK2 inhibited cell proliferation, whereas depletion of ERK 1 (but not ERK2) markedly increased cell apoptosis. These findings identify MEK/ERK as a new signaling pathway activated by NPM/ALK and indicate that the pathway represents a novel therapeutic target in the ALK-induced malignancies.

Asunto(s)

MAP Quinasa Quinasa 1/metabolismo , MAP Quinasa Quinasa 2/metabolismo , Sistema de Señalización de MAP Quinasas , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-raf/metabolismo , Proliferación Celular , Células Cultivadas , Activación Enzimática , Eliminación de Gen , Regulación de la Expresión Génica , Humanos , Proteína Quinasa 1 Activada por Mitógenos/genética , Proteína Quinasa 3 Activada por Mitógenos/genética , Proteínas Tirosina Quinasas/genética

RESUMEN

To understand better T-cell lymphomagenesis, we examined promoter CpG methylation and mRNA expression of closely related genes encoding p16, p15, and p14 tumor suppressor genes in cultured malignant T-cells that were derived from cutaneous, adult type, and anaplastic lymphoma kinase (ALK)-expressing T-cell lymphomas. p16 gene was epigenetically silenced in all but one of the 10 malignant T-cell lines examined, p15 gene silenced in roughly half of the lines, and p14 was the least frequently affected. Extensive methylation of the p16 promoter was seen in six out of 10 cutaneous T-cell lymphoma patient samples and corresponded with lack of p16 protein expression in the cases examined. Treatment of cultured T-cells with the DNA methyltransferase inhibitor, 5-aza-2-deoxy-cytidine, resulted in reversal of the p16 gene silencing. However, expression of p16 protein was delayed in relationship to p16 promoter demethylation and required up to 3 weeks to occur, seemingly reflecting late activation of the p16 gene. These findings indicate that epigenetic silencing affects in T-cell malignancies, often simultaneously, several tumor suppressor genes that impact on key cell functions. The observed differential silencing of p16 and p14, and to a lesser degree p15 gene, indicates that the silencing is governed by precise, promoter region-specific mechanisms. The study provides also further rationale for treatment of at least some types of T-cell lymphomas with DNA methyltransferase inhibitors to target the epigenetically silenced tumor suppressor genes.

Asunto(s)

Epigénesis Genética , Regulación Leucémica de la Expresión Génica , Silenciador del Gen , Linfoma Cutáneo de Células T/metabolismo , Neoplasias Cutáneas/metabolismo , Proteína p14ARF Supresora de Tumor/biosíntesis , Adulto , Quinasa de Linfoma Anaplásico , Azacitidina/análogos & derivados , Azacitidina/farmacología , Línea Celular Tumoral , Inhibidor p15 de las Quinasas Dependientes de la Ciclina/biosíntesis , Metilación de ADN/efectos de los fármacos , Metilasas de Modificación del ADN/antagonistas & inhibidores , Metilasas de Modificación del ADN/metabolismo , Decitabina , Inhibidores Enzimáticos/farmacología , Epigénesis Genética/efectos de los fármacos , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Silenciador del Gen/efectos de los fármacos , Humanos , Linfoma Cutáneo de Células T/tratamiento farmacológico , Linfoma Cutáneo de Células T/patología , Regiones Promotoras Genéticas , Biosíntesis de Proteínas/efectos de los fármacos , Proteínas Tirosina Quinasas/biosíntesis , Proteínas Tirosina Quinasas Receptoras , Neoplasias Cutáneas/tratamiento farmacológico , Factores de Tiempo

RESUMEN

BACKGROUND: Given that vagal afferents project to brainstem regions that promote alertness, the authors tested the hypothesis that vagus nerve stimulation (VNS) would improve daytime sleepiness in patients with epilepsy. METHODS: Sixteen subjects with medically refractory seizures underwent polysomnography and multiple sleep latency tests (MSLT) and completed the Epworth Sleepiness Scale (ESS), a measure of subjective daytime sleepiness, before and after 3 months of VNS. Most subjects (>80%) were maintained on constant doses of antiepileptic medications. RESULTS: In the 15 subjects who completed baseline and treatment MSLT, the mean sleep latency (MSL) improved from 6.4 +/- 4.1 minutes to 9.8 +/- 5.8 minutes (+/- SD; p = 0.033), indicating reduced daytime sleepiness. All subjects with stimulus intensities of < or =1.5 mA showed improved MSL. In the 16 subjects who completed baseline and treatment ESS, the mean ESS score decreased from 7.2 +/- 4.4 to 5.6 +/- 4.5 points (p = 0.049). Improvements in MSLT and ESS were not correlated with reduction in seizure frequency. Sleep-onset REM periods occurred more frequently in treatment naps as compared to baseline naps (p < 0.008; Cochran-Mantel-Haenszel test). The amount of REM sleep or other sleep stages recorded on overnight polysomnography did not change with VNS treatment. CONCLUSIONS: Treatment with VNS at low stimulus intensities improves daytime sleepiness, even in subjects without reductions in seizure frequency. Daytime REM sleep is enhanced with VNS. These findings support the role of VNS in activating cholinergic and other brain regions that promote alertness.

Asunto(s)

Epilepsia/terapia , Privación de Sueño/terapia , Estimulación Eléctrica Transcutánea del Nervio , Nervio Vago , Adulto , Epilepsia/complicaciones , Femenino , Humanos , Masculino , Persona de Mediana Edad , Polisomnografía/métodos , Privación de Sueño/complicaciones , Sueño REM/fisiología , Estimulación Eléctrica Transcutánea del Nervio/métodos , Nervio Vago/fisiología

RESUMEN

BACKGROUND: Vagus nerve stimulation (VNS) is associated with respiratory effects such as hoarseness, dyspnea, and laryngeal irritation. The effects of VNS on sleep-related breathing in humans have not been reported previously. METHODS: Four epilepsy patients underwent polysomnography (PSG) before and after 3 months of treatment with VNS. Two of the four patients also underwent follow-up PSG to assess the effects of changing stimulus parameters on sleep-related breathing. RESULTS: All patients showed consistent sleep-related decreases in airflow and effort coinciding with VNS activation, although most events did not meet laboratory criteria for apneas or hypopneas. Apneas and hypopneas were more frequent during VNS activation than during nonactivation. Apnea-hypopnea index (AHI) for three subjects during VNS treatment PSG was <5 apneas and hypopneas/hour. In one patient with obstructive sleep apnea (OSA) before VNS treatment, AHI rose from 4 (pretreatment) to 11.3 (treatment). In this patient and in another patient without clinically significant OSA, lowering stimulus frequency, but not stimulus intensity, pulse width, or on-time, ameliorated VNS-related apneas and hypopneas. CONCLUSIONS: VNS is associated with adverse changes in respiration during sleep. In patients without preexisting OSA, this VNS effect is probably not clinically significant. In patients with preexisting OSA, VNS should be administered with care. Lowering VNS stimulus frequency or prolonging off-time may prevent exacerbation of OSA.

Asunto(s)

Epilepsia/fisiopatología , Sueño/fisiología , Nervio Vago/fisiopatología , Adulto , Estimulación Eléctrica , Femenino , Humanos , Masculino , Persona de Mediana Edad , Polisomnografía