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The cosmic microwave background (CMB) B-mode signal is potentially weaker than the diffuse Galactic foregrounds over most of the sky at any frequency. A common method of separating the CMB from these foregrounds is via pixel-based parametric-model fitting. There are not currently enough all-sky maps to fit anything more than the most simple models of the sky. By simulating the emission in seven representative pixels, we demonstrate that the inclusion of a 5 GHz data point allows for more complex models of low-frequency foregrounds to be fitted than at present. It is shown that the inclusion of the C-BASS data will significantly reduce the uncertainties in a number of key parameters in the modelling of both the galactic foregrounds and the CMB. The extra data allow estimates of the synchrotron spectral index to be constrained much more strongly than is presently possible, with corresponding improvements in the accuracy of the recovery of the CMB amplitude. However, we show that to place good limits on models of the synchrotron spectral curvature will require additional low-frequency data.
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We present multi-wavelength detections of nine candidate gravitationally-lensed dusty star-forming galaxies (DSFGs) selected at 218GHz (1.4mm) from the ACT equatorial survey. Among the brightest ACT sources, these represent the subset of the total ACT sample lying in Herschel SPIRE fields, and all nine of the 218GHz detections were found to have bright Herschel counterparts. By fitting their spectral energy distributions (SEDs) with a modified blackbody model with power-law temperature distribution, we find the sample has a median redshift of z = 4.1 - 1.0 + 1.1 (68 per cent confidence interval), as expected for 218GHz selection, and an apparent total infrared luminosity of log 10 ( µ L IR / L â ) = 13.86 - 0.30 + 0.33 , which suggests that they are either strongly lensed sources or unresolved collections of unlensed DSFGs. The effective apparent diameter of the sample is µ d = 4.2 - 1.0 + 1.7 kpc , further evidence of strong lensing or multiplicity, since the typical diameter of dusty star-forming galaxies is 1.0-2.5 kpc. We emphasize that the effective apparent diameter derives from SED modelling without the assumption of optically thin dust (as opposed to image morphology). We find that the sources have substantial optical depth. ( τ = 4.2 - 1.9 + 3.7 ) to dust around the peak in the modified blackbody spectrum (λ obs ⩽ 500µm), a result that is robust to model choice.
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We evaluate the modulation of cosmic microwave background polarization using a rapidly rotating, half-wave plate (HWP) on the Atacama B-Mode Search. After demodulating the time-ordered-data (TOD), we find a significant reduction of atmospheric fluctuations. The demodulated TOD is stable on time scales of 500-1000 s, corresponding to frequencies of 1-2 mHz. This facilitates recovery of cosmological information at large angular scales, which are typically available only from balloon-borne or satellite experiments. This technique also achieves a sensitive measurement of celestial polarization without differencing the TOD of paired detectors sensitive to two orthogonal linear polarizations. This is the first demonstration of the ability to remove atmospheric contamination at these levels from a ground-based platform using a rapidly rotating HWP.
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Pediatric cardiopulmonary bypass is still a challenge because of electrolyte disturbances and inflammation. Many investigations deal with different types of hemofiltration to reduce these potentially harmful side effects. We tested the hypothesis of whether bicarbonate-buffered hemofiltration of the priming solution minimizes electrolyte and acid-base disturbances during the initiation of cardiopulmonary bypass and whether bicarbonate-buffered hemofiltration performed during cardiopulmonary bypass could reduce cytokine levels. Twenty children younger than 2 years of age (mean age 166 +/- 191 days; mean weight 6.42 +/- 3.22 kg) scheduled for pediatric cardiac surgery with cardiopulmonary bypass were enrolled in this prospective clinical study. Cardiopulmonary bypass circuits were primed with a bicarbonate-buffered hemofiltration solution, gelatin and 1 unit of packed red blood cells. The priming was hemofiltered using an ultrahemofilter until approximately 1000 mL of ultrafiltrate was restored with the buffered solution. Further hemofiltration was performed throughout the whole bypass time, especially during rewarming. Blood gas analyses and inflammatory mediators were monitored during the operation. Blood gas analysis results after initiation of cardiopulmonary bypass and throughout the entire study remained within the physiologic ranges. Even potassium decreased from 4.0 +/- 0.3 to 3.4 +/- 0.4 mmol l(-1) after initiation of cardiopulmonary bypass. Plasma levels of tumor necrosis factor alpha decreased significantly (47 +/- 44 vs. 24 +/- 21 pg mL(-1)) whereas complement factor C3a (5.0 +/- 2.9 vs. 16.8 +/- 6.6 ng mL(-1)) and interleukin-6 (7.3 +/- 15.2 vs. 110 +/- 173 pg mL(-1)) increased despite hemofiltration. In conclusion, this study shows that bicarbonate-buffered ultrafiltration is an efficient, simple and safe method for performing hemofiltration, both of the priming solution and during the entire bypass time. The use of a physiological restitution solution prevents electrolyte and acid-base balance disturbances. The elimination of inflammatory mediators seems to be as effective as other ultrafiltration methods.
Asunto(s)
Equilibrio Ácido-Base/efectos de los fármacos , Bicarbonatos/farmacología , Puente Cardiopulmonar/métodos , Hemofiltración/métodos , Equilibrio Hidroelectrolítico/efectos de los fármacos , Glucemia , Citocinas/sangre , Hematócrito , Hemoglobinas/análisis , Humanos , Lactante , Ácido Láctico/sangre , Estudios ProspectivosRESUMEN
BACKGROUND: Apnoea of prematurity (AOP) is a common problem in preterm infants which can be treated with various modes of nasal continuous positive airway pressure (NCPAP) or nasal intermittent positive pressure ventilation (NIPPV). It is not known which mode of NCPAP or NIPPV is most effective for AOP. OBJECTIVE: To assess the effect of four NCPAP/NIPPV systems on the rate of bradycardias and desaturation events in very low birthweight infants. METHODS: Sixteen infants (mean gestational age at time of study 31 weeks, 10 males) with AOP were enrolled in a randomised controlled trial with a crossover design. The infants were allocated to receive nasal pressure support using four different modes for 6 h each: NIPPV via a conventional ventilator, NIPPV and NCPAP via a variable flow device, and NCPAP delivered via a constant flow underwater bubble system. The primary outcome was the cumulative event rate of bradycardias (< or =80 beats per minute) and desaturation events (< or =80% arterial oxygen saturation), which was obtained from cardio-respiratory recordings. RESULTS: The median event rate was 6.7 per hour with the conventional ventilator in NIPPV mode, and 2.8 and 4.4 per hour with the variable flow device in NCPAP and NIPPV mode, respectively (p value<0.03 for both compared to NIPPV/conventional ventilator). There was no significant difference between the NIPPV/conventional ventilator and the underwater bubble system. CONCLUSION: A variable flow NCPAP device may be more effective in treating AOP in preterm infants than a conventional ventilator in NIPPV mode. It remains unclear whether synchronised NIPPV would be even more effective.
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Apnea/terapia , Enfermedades del Prematuro/terapia , Recién Nacido de muy Bajo Peso , Respiración con Presión Positiva/métodos , Peso al Nacer , Presión de las Vías Aéreas Positiva Contínua , Estudios Cruzados , Femenino , Edad Gestacional , Humanos , Recién Nacido , Recien Nacido Prematuro , Ventilación con Presión Positiva Intermitente , Masculino , Resultado del TratamientoRESUMEN
Agents suppressing microglial activation are attracting attention as candidate drugs for neuroprotection in Parkinson s disease (PD): While different mechanisms including environmental toxins and genetic factors initiate neuronal damage in the substantia nigra and striatum in PD, there is unequivocal evidence that activation of neuroinflammatory cells aggravates this neurodegenerative process. It was shown that following an acute exposure to the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and other toxins the degenerative process continues for years in absence of the toxin. Reactive microglia has been observed in the substantia nigra of patients with PD, indicating that this inflammatory process might aggravate neurodegeneration. By releasing various kinds of noxious factors such as cytokines or proinflammatory molecules microglia may damage CNS cells. The stimuli triggering microgliosis in Parkinsonian syndromes are unknown so far: However, analysis of neuronal loss in PD patients shows that it is not uniform but that neurons containing neuromelanin (NM) are predominantly involved. We hypothesized that extraneuronal melanin might trigger microgliosis, microglial chemotaxis and microglial activation in PD with subsequent release of neurotoxic mediators. The addition of human NM to microglial cell cultures induced positive chemotactic effects, activated the pro-inflammatory transcription factor nuclear factor kappa B (NF-kappaB) via phosphorylation and degradation of the inhibitor protein kappaB (IkappaB), and led to an upregulation of TNF-alpha, IL-6 and NO. These findings demonstrate a crucial role of NM in the pathogenesis of Parkinson's disease by augmentation of microglial activation, leading to a vicious cycle of neuronal death, exposure of additional neuromelanin and chronification of inflammation. Antiinflammatory drugs may be one of the new approaches in the treatment of PD.
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Antiinflamatorios/farmacología , Inflamación/tratamiento farmacológico , Melaninas/metabolismo , Microglía/efectos de los fármacos , Enfermedades Neurodegenerativas/tratamiento farmacológico , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Sustancia Negra/efectos de los fármacos , Animales , Antiinflamatorios/uso terapéutico , Antiparkinsonianos/farmacología , Muerte Celular/efectos de los fármacos , Citocinas/metabolismo , Humanos , Inflamación/metabolismo , Inflamación/patología , Mediadores de Inflamación/metabolismo , Microglía/metabolismo , Microglía/patología , Enfermedades Neurodegenerativas/metabolismo , Enfermedades Neurodegenerativas/patología , Neuronas/patología , Fármacos Neuroprotectores/uso terapéutico , Enfermedad de Parkinson/tratamiento farmacológico , Sustancia Negra/metabolismo , Sustancia Negra/patologíaRESUMEN
Polarization observations of the cosmic microwave background with the Cosmic Background Imager from September 2002 to May 2004 provide a significant detection of the E-mode polarization and reveal an angular power spectrum of polarized emission showing peaks and valleys that are shifted in phase by half a cycle relative to those of the total intensity spectrum. This key agreement between the phase of the observed polarization spectrum and that predicted on the basis of the total intensity spectrum provides support for the standard model of cosmology, in which dark matter and dark energy are the dominant constituents, the geometry is close to flat, and primordial density fluctuations are predominantly adiabatic with a matter power spectrum commensurate with inflationary cosmological models.
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Increased binding of a ligand for the peripheral benzodiazepine binding receptor is currently used in PET studies as an in vivo measurement of inflammation in diseases like multiple sclerosis and Alzheimer's disease. Although peripheral-type benzodiazepin receptors (PBRs) are abundant in many cell types and expressed in the CNS physiologically only at low levels, previous reports suggest that after experimental lesions in animal models and in human neurodegenerative/-inflammatory diseases upregulated PBR expression with increased binding of its ligand PK11195 is confined mainly to activated microglia in vivo/in situ. Because the functional role of the PBR is unknown, we confirm by immunohistochemistry and PCR (I) that this receptor is expressed on microglia in vitro and (II) that benzodiazepines modulate proliferation of microglial cells and the release of the inflammatory molecules nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha) in cell culture supernatants of primary rat microglia. Compared to lipopolysaccharide-activated controls the release of NO was markedly decreased in cultures treated with benzodiazepines (clonazepam, midazolam, diazepam) and the PBR ligand PK11195. Moreover, release of TNF-alpha and proliferation was significantly inhibited in the benzodiazepine-treated groups. These findings link the in vivo data of elevated PBR levels in neurodegenerative/-inflammatory diseases to a functional role and opens up possible therapeutic intervention targeting the PBR in microglia.
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Encefalitis/metabolismo , Gliosis/metabolismo , Microglía/metabolismo , Enfermedades Neurodegenerativas/metabolismo , Receptores de GABA-A/metabolismo , Animales , Animales Recién Nacidos , Benzodiazepinas/farmacología , Sitios de Unión/efectos de los fármacos , Sitios de Unión/fisiología , División Celular/efectos de los fármacos , División Celular/fisiología , Células Cultivadas , Encefalitis/tratamiento farmacológico , Encefalitis/fisiopatología , Gliosis/patología , Gliosis/fisiopatología , Mediadores de Inflamación/metabolismo , Isoquinolinas/farmacología , Ligandos , Microglía/efectos de los fármacos , Microglía/patología , Mielitis/metabolismo , Mielitis/fisiopatología , Enfermedades Neurodegenerativas/tratamiento farmacológico , Enfermedades Neurodegenerativas/fisiopatología , Óxido Nítrico/metabolismo , Ratas , Ratas Wistar , Factor de Necrosis Tumoral alfa/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/fisiologíaRESUMEN
Autopsy studies and animal experiments suggest that microglial inflammation contributes to the pathogenesis of amyotrophic lateral sclerosis (ALS). Monocyte-chemoattractant protein (MCP-1) might play an important role in microglial recruitment. We studied MCP-1 levels in sera and cerebrospinal fluid of 29 ALS patients and compared the results with 11 control patients with tension headache. The MCP-1 level was determined using enzyme-linked immunosorbent assays (ELISA). A significant increase in cerebrospinal fluid MCP-1 level but not serum level was seen in the patients with ALS compared to the control subjects. These results suggest that cerebrospinal fluid MCP-1 activity may be a sensitive marker for neuroinflammation in ALS useful for monitoring treatment trials in ALS.
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Esclerosis Amiotrófica Lateral/líquido cefalorraquídeo , Esclerosis Amiotrófica Lateral/patología , Proteínas del Líquido Cefalorraquídeo/biosíntesis , Quimiocina CCL2/biosíntesis , Quimiocina CCL2/líquido cefalorraquídeo , Microglía/metabolismo , Microglía/patología , Edad de Inicio , Anciano , Esclerosis Amiotrófica Lateral/inmunología , Análisis de Varianza , Movimiento Celular/inmunología , Proteínas del Líquido Cefalorraquídeo/sangre , Proteínas del Líquido Cefalorraquídeo/líquido cefalorraquídeo , Quimiocina CCL2/sangre , Humanos , Microglía/inmunología , Persona de Mediana Edad , Análisis de Regresión , Estadísticas no Paramétricas , Regulación hacia Arriba/inmunologíaRESUMEN
PURPOSE: To report the detection of non-compacted ventricular myocardium (NCVM) with MRI compared to echocardiography in 8 patients. MATERIAL AND METHODS: Non-compaction of the ventricular myocardium is a congenital disorder characterized by an altered structure of the myocardial wall resulting from an intrauterine arrest in endomyocardial embryogenesis. The morphological findings consist of a prominent meshwork of multiple myocardial trabeculations and deep intertrabecular recesses, communicating with the left ventricular cavity. 8 consecutive patients (mean age 7.3 years) with clinical and echocardiographic signs of NCVM were examined by MRI (1.5 T, Vision, Siemens) in short axis and 2- and 4-chamber views, using T (1)-weighted TSE and Cine-GRE in 6 patients and true FISP sequences in 2 patients. MRI and echocardiography were evaluated for visibility, signs of NCVM and involvement of myocardial wall segments. Thickness was measured for non-compacted and compacted myocardium and the non-compacted to compacted (N/C) ratio calculated. RESULTS: MRI diagnosed 6 of 8 patients of having NCVM. Myocardial thickness as measured by echocardiography and MRI showed a good correlation in compacted myocardium (r = 8.82) and no correlation in non-compacted myocardium (r = 0.4). In 2 cases, non-compacted myocardium was detected but echocardiography did not reach the N/C ratio > 2 as required to diagnose NCVM in accordance with the criteria found in the literature. Both patients were also misdiagnosed by MRI performed with Cine-GRE. MRI reached a N/C ratio > 2 in only three patients. Newer TruFisp sequences showed no definite advantages. Extent of non-compaction could be visualized correctly with MRI. CONCLUSION: Echocardiography is the method of choice to detect NCVM. MRI can be an alternative in some cases. The diagnosis of NCVM should not be considered until N/C ratio is over 2.
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Ecocardiografía , Cardiopatías Congénitas/diagnóstico por imagen , Cardiopatías Congénitas/diagnóstico , Imagen por Resonancia Magnética , Adolescente , Factores de Edad , Anciano , Niño , Preescolar , Electrocardiografía , Frecuencia Cardíaca , Humanos , Lactante , Imagen por Resonancia Magnética/métodos , Imagen por Resonancia Cinemagnética/métodos , Disfunción Ventricular Izquierda/diagnóstico , Disfunción Ventricular Izquierda/etiologíaRESUMEN
The cerebellar external granular layer (EGL) is an unusually long-lasting neural proliferative zone positioned immediately beneath the pial surface. Its position and stability critically depend on meningeal cells, as their selective destruction leads to its rapid dispersal, creating massive cortical ectopia. Similar ectopias have recently been described as a side effect of deficiency for stromal cell-derived factor 1 (SDF-1), a chemoattractant for haematopoietic precursor cell migration. Here we show that SDF-1 is present in meningeal cells in vivo and in vitro, where it is secreted in functionally relevant concentrations into the medium. Correspondingly, the SDF-1 receptor (termed CXCR4) can be demonstrated on stem cells of the external granular layer, but is absent on postmitotic cells commencing their final inward migration. We show that SDF-1 is concentrated by heparan sulphate proteoglycans highly expressed in the EGL in a laminar fashion, which thus might act to locally restrict SDF-1 action to the EGL in a kind of step gradient. In vitro, SDF-1 chemotactically attracts neuronal cells isolated from the external, but not from the internal granular layer, in a Boyden chamber assay in concentrations found in meningeal cell-conditioned medium. Selective removal of SDF-1 from conditioned media by immunoprecipitation abolishes their chemoattractive action, which can be reconstituted again by the addition of recombinant SDF-1. Meningeal cells are thus an important source for the expression of SDF-1 during brain development, which--comparable to its role in haematopoiesis--appears to be a key factor attracting precursor cells to their proliferative compartment.
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Cerebelo/fisiología , Quimiocinas CXC/metabolismo , Factores Quimiotácticos/metabolismo , Meninges/metabolismo , Neuronas/metabolismo , Células Madre/metabolismo , Animales , Animales Recién Nacidos , Células Cultivadas , Cerebelo/química , Cerebelo/citología , Quimiocina CXCL12 , Quimiocinas CXC/análisis , Quimiocinas CXC/biosíntesis , Factores Quimiotácticos/análisis , Factores Quimiotácticos/biosíntesis , Embrión de Mamíferos , Femenino , Masculino , Meninges/química , Meninges/citología , Neuronas/química , Ratas , Ratas Wistar , Células Madre/químicaRESUMEN
To determine the possible contribution of glial cells via oxidative stress/cytokine secretion in the pathogenesis of Parkinson's disease (PD), Alzheimer disease (AD), amyotrophic lateral sclerosis (ALS) or multiple sclerosis (MS) the concentration of nitric oxide (NO) (by the Griess method) and Interleukin-6 (IL-6) (by enzyme-linked immunosorbent assay) were measured in resting rat microglial and astrocytic cell culture supernatants stimulated by cerebrospinal fluid (CSF) (dilution 1:4, 1:10) from patients with the aforementioned diseases. Neither the concentration of NO (optical density at 450 nm: control, 0.036+/-0.006; MS, 0.034+/-0.008; AD, 0.031+/-0.006; PD, 0.02+/-0.01; lipopolysaccharide (LPS), 0.26+/-0.018) nor the amount of IL-6 (ng/ml: control, 0.112+/-0.026; PD, 0.12+/-0.027; MS, 0.123+/-0.008; ALS, 0.137+/-0.01; LPS, 1.81+/-0.11) differed in any disease group from those of unaffected controls. These findings suggest that the stimuli for inflammatory activation of glia are quite localized and not present in sufficient concentrations in the CSF of affected patients.
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Citocinas/inmunología , Encefalitis/líquido cefalorraquídeo , Gliosis/líquido cefalorraquídeo , Enfermedades Neurodegenerativas/líquido cefalorraquídeo , Neuroglía/inmunología , Óxido Nítrico/inmunología , Estrés Oxidativo/inmunología , Astrocitos/efectos de los fármacos , Astrocitos/inmunología , Astrocitos/metabolismo , Comunicación Autocrina/efectos de los fármacos , Comunicación Autocrina/inmunología , Células Cultivadas/efectos de los fármacos , Células Cultivadas/inmunología , Células Cultivadas/metabolismo , Proteínas del Líquido Cefalorraquídeo/inmunología , Proteínas del Líquido Cefalorraquídeo/metabolismo , Proteínas del Líquido Cefalorraquídeo/farmacología , Citocinas/metabolismo , Citocinas/farmacología , Encefalitis/inmunología , Encefalitis/fisiopatología , Gliosis/inducido químicamente , Gliosis/inmunología , Humanos , Interferón gamma/inmunología , Interferón gamma/metabolismo , Interferón gamma/farmacología , Interleucina-1/inmunología , Interleucina-1/metabolismo , Interleucina-1/farmacología , Interleucina-6/inmunología , Interleucina-6/metabolismo , Interleucina-6/farmacología , Lipopolisacáridos/farmacología , Microglía/efectos de los fármacos , Microglía/inmunología , Microglía/metabolismo , Enfermedades Neurodegenerativas/inmunología , Enfermedades Neurodegenerativas/fisiopatología , Neuroglía/efectos de los fármacos , Neuroglía/metabolismo , Óxido Nítrico/metabolismo , Factor de Necrosis Tumoral alfa/inmunología , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
Microglia are the principal immune cells in the central nervous system (CNS), characterized by a highly specific morphology and unusual antigenic phenotype. An increasing number of studies have focused on the role of microglia in the pathogenesis of neurodegenerative diseases. To elucidate the function of microglial cells under several neuropathological conditions, we have studied and established a cell culture model that allows us to cultivate microglial cells in their inactive, resting (ramified) phenotype. In the first part of this work, we describe the interaction of microglia cells with their epithelial (astrocytic) microenvironment. The second part reviews experiments with microglia cell cultures to elucidate underlying signalling pathways and summarizes recent advances of our knowledge in microglial molecular pathways that may ultimately lead to neurodegeneration.
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Microglía/fisiología , Animales , Astrocitos/fisiología , Técnicas de Cultivo de Célula , Movimiento Celular , Células Cultivadas , Células Epiteliales/fisiología , Regulación de la Expresión Génica , Humanos , Mediadores de Inflamación , Microglía/inmunología , Microglía/ultraestructura , Modelos Neurológicos , FN-kappa B/fisiología , Fenotipo , Transducción de Señal , Transcripción GenéticaRESUMEN
Microglial cells in the healthy adult brain possess a characteristic ramified morphology with multiple branched processes, small somata and down-regulated inflammatory properties. In contrast, microglial cells isolated from new-born rat brain inevitably show a non-ramified amoeboid phenotype, which is observed in vivo after pathologic activation or during development. To identify factors that control microglial morphology we investigated the effects of purines alone or in combination with astrocyte-conditioned medium (ACM). Under optimized culture conditions postnatal rat microglial cells developed an amoeboid to ovoid phenotype. Addition of 0.6-1 mM ATP or adenosine induced the outgrowth of numerous processes after 2-3 days that could be observed also in the presence of ACM as previously reported. Culture in ACM plus ATP or adenosine yielded an optimized ramified phenotype. ATP or adenosine, but not ACM alone, also prevented the formation of a flat, amoeboid morphology induced by lipopolysaccharide (LPS); however, at 0.6-1 mM they did not reduce the initial LPS-induced activation of the transcription factor NF-kappaB. By using specific agonists or antagonists the morphological transformations could not be confined to a distinct purinoreceptor subtype, but appeared to be mediated by long-term presence of adenosine in the medium to which phosphorylated purines were rapidly hydrolyzed by microglial cells. Since ACM did not contain sufficient concentrations of ATP or adenosine, purines are not the only ramification-inducing factors present in ACM; however, they are a valuable tool to induce microglial ramification in vitro.
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Adenosina Trifosfato/metabolismo , Adenosina/metabolismo , Microglía/metabolismo , Adenosina/farmacología , Adenosina Trifosfato/farmacología , Animales , Animales Recién Nacidos , Astrocitos/citología , Astrocitos/metabolismo , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Medios de Cultivo Condicionados/farmacología , Relación Dosis-Respuesta a Droga , Lipopolisacáridos/farmacología , Microglía/citología , Microglía/efectos de los fármacos , FN-kappa B/biosíntesis , Fenotipo , Agonistas Purinérgicos , Antagonistas Purinérgicos , Purinas/metabolismo , Ratas , Ratas Wistar , Receptores Purinérgicos/metabolismoRESUMEN
The ftsL gene is required for the initiation of cell division in a broad range of bacteria. Bacillus subtilis ftsL encodes a 13-kDa protein with a membrane-spanning domain near its N terminus. The external C-terminal domain has features of an alpha-helical leucine zipper, which is likely to be involved in the heterodimerization with another division protein, DivIC. To determine what residues are important for FtsL function, we used both random and site-directed mutagenesis. Unexpectedly, all chemically induced mutations fell into two clear classes, those either weakening the ribosome-binding site or producing a stop codon. It appears that the random mutagenesis was efficient, so many missense mutations must have been generated but with no phenotypic effect. Substitutions affecting hydrophobic residues in the putative coiled-coil domain, introduced by site-directed mutagenesis, also gave no observable phenotype except for insertion of a helix-breaking proline residue, which destroyed FtsL function. ftsL homologues cloned from three diverse Bacillus species, Bacillus licheniformis, Bacillus badius, and Bacillus circulans, could complement an ftsL null mutation in B. subtilis, even though up to 66% of the amino acid residues of the predicted proteins were different from B. subtilis FtsL. However, the ftsL gene from Staphylococcus aureus (whose product has 73% of its amino acids different from those of the B. subtilis ftsL product) was not functional. We conclude that FtsL is a highly malleable protein that can accommodate a large number of sequence changes without loss of function.
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Bacillus subtilis/genética , Proteínas Bacterianas/genética , Proteínas de Ciclo Celular , Proteínas de Escherichia coli , Genes Bacterianos , Leucina Zippers , Proteínas de la Membrana/genética , Alelos , Secuencia de Aminoácidos , Bacillus , Bacillus subtilis/citología , Bacillus subtilis/fisiología , Proteínas Bacterianas/fisiología , Secuencia de Bases , División Celular , Clonación Molecular , ADN Bacteriano , Prueba de Complementación Genética , Proteínas de la Membrana/fisiología , Datos de Secuencia Molecular , Mutagénesis , Homología de Secuencia de Aminoácido , Relación Estructura-ActividadRESUMEN
FtsL is a small bitopic membrane protein required for vegetative cell division and sporulation in Bacillus subtilis. We investigated its localization by fluorescence microscopy using a green fluorescent protein (GFP) fusion. GFP-FtsL was localized at mid-cell in vegetative cells and at the asymmetric septum in sporulating cells. We also show that FtsL forms a ring-like structure at the division site and that it remains localized at mid-cell during the whole septation process. By yeast two-hybrid analysis and non-denaturing polyacrylamide gel electrophoresis (PAGE) with purified proteins, FtsL was found to interact with another membrane-bound division protein, the FtsL-like DivIC protein.
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Bacillus subtilis/metabolismo , Proteínas Bacterianas/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Escherichia coli , Proteínas de la Membrana/metabolismo , Animales , Bacillus subtilis/genética , Bacillus subtilis/fisiología , Proteínas Bacterianas/genética , División Celular , Proteínas Fluorescentes Verdes , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Proteínas de la Membrana/genética , Conejos , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Esporas BacterianasRESUMEN
OBJECTIVES: We sought to validate recently proposed risk adjustment models for in-hospital percutaneous transluminal coronary angioplasty (PTCA) mortality on an independent data set of high risk patients undergoing PTCA. BACKGROUND: Risk adjustment models for PTCA mortality have recently been reported, but external validation on independent data sets and on high risk patient groups is lacking. METHODS: Between July 1, 1994 and June 1, 1996, 1,476 consecutive procedures were performed on a high risk patient group characterized by a high incidence of cardiogenic shock (3.3%) and acute myocardial infarction (14.3%). Predictors of in-hospital mortality were identified using multivariate logistic regression analysis. Two external models of in-hospital mortality, one developed by the Northern New England Cardiovascular Disease Study Group (model NNE) and the other by the Cleveland Clinic (model CC), were compared using receiver operating characteristic (ROC) curve analysis. RESULTS: In this patient group, an overall in-hospital mortality rate of 3.4% was observed. Multivariate regression analysis identified risk factors for death in the hospital that were similar to the risk factors identified by the two external models. When fitted to the data set, both external models had an area under the ROC curve >0.85, indicating overall excellent model discrimination, and both models were accurate in predicting mortality in different patient subgroups. There was a trend toward a greater ability to predict mortality for model NNE as compared with model CC, but the difference was not significant. CONCLUSIONS: Predictive models for PTCA mortality yield comparable results when applied to patient groups other than the one on which the original model was developed. The accuracy of the two models tested in adjusting for the relatively high mortality rate observed in this patient group supports their application in quality assessment or quality improvement efforts.
Asunto(s)
Angioplastia Coronaria con Balón/mortalidad , Enfermedad Coronaria/mortalidad , Mortalidad Hospitalaria , Ajuste de Riesgo/estadística & datos numéricos , Anciano , Angioplastia Coronaria con Balón/estadística & datos numéricos , Enfermedad Coronaria/terapia , Grupos Diagnósticos Relacionados/estadística & datos numéricos , Femenino , Humanos , Modelos Logísticos , Masculino , Michigan/epidemiología , Persona de Mediana Edad , Oportunidad Relativa , Pronóstico , Curva ROC , Reproducibilidad de los Resultados , Factores de RiesgoRESUMEN
Component D (HgdAB) of 2-hydroxyglutaryl-CoA dehydratase from Clostridium symbiosum was purified to homogeneity. It is able to use component A from Acidaminococcus fermentans (HgdC) to initiate catalysis together with ATP, Mg2+ and a strong reducing agent such as Ti(III)citrate. Component D from C. symbiosum has a 6 x higher specific activity compared with that from A. fermentans and contains a second [4Fe-4S] cluster but the same amount of riboflavin 5'-phosphate (1.0 per heterodimeric enzyme, m = 100 kDa). Mössbauer spectroscopy revealed symmetric cube-type structures of the two [4Fe-4S]2+ clusters. EPR spectroscopy showed the resistance of the clusters to reducing agents, but detected a sharp signal at g = 2. 004 probably due to a stabilized flavin semiquinone. Three genes from C. symbiosum coding for components D (hgdA and hgdB) and A (hgdC) were cloned and sequenced. Primer extension experiments indicated that the genes are transcribed in the order hgdCAB from an operon only half the size of that from A. fermentans. Sequence comparisons detected a close relationship to the dehydratase system from A. fermentans and HgdA from Fusobacterium nucleatum, as well as to putative proteins of unknown function from Archaeoglobus fulgidus. Lower, but significant, identities were found with putative enzymes from several methanogenic Archaea and Escherichia coli, as well as with the mechanistically related benzoyl-CoA reductases from the Proteobacteria Rhodopseudomonas palustris and Thauera aromatica.
Asunto(s)
Clostridium/enzimología , Hidroliasas/aislamiento & purificación , Proteínas Hierro-Azufre/aislamiento & purificación , Secuencia de Aminoácidos , Archaea/enzimología , Bacterias/enzimología , Clonación Molecular , Espectroscopía de Resonancia por Spin del Electrón , Mononucleótido de Flavina/aislamiento & purificación , Genes Bacterianos , Hidroliasas/genética , Proteínas Hierro-Azufre/genética , Modelos Químicos , Datos de Secuencia Molecular , Operón , Proteobacteria/enzimología , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Espectroscopía de MossbauerRESUMEN
Animal models of tremor have been widely used in experimental neurology, because they are an indispensable requirement for understanding the pathophysiology of human tremor disorders and the development of new therapeutic agents. This review focuses on three approaches to produce tremor in animals (application of tremorgenic drugs, experimental central nervous system lesions, study of genetic mutants) and their use in simulating tremor syndromes of humans. Whereas harmaline induces a postural/kinetic tremor in animals that shares some features with human essential tremor/enhanced physiological tremor, MPTP tremor is the best model available for rest tremor in people. The tremor following experimental lesion of the ventromedial tegmentum in primates closely resembles Holmes tremor in humans, whereas cerebellar intention tremor is mimicked by cooling of the lateral cerebellar nuclei. The "campus syndrome," discovered in a breed of Pietrain pigs, might be a useful model of human orthostatic tremor. However, no animal model has yet been generated that exactly recreates all features of any of the known tremor disorders in humans. Problems encountered when comparing tremor in animals and humans include differing tremor frequencies and the uncertainty, if specific transmitter abnormalities/central nervous system lesions seen in animal tremor models are characteristic for their human counterparts. The search for adequate tremor models continues.
Asunto(s)
Modelos Animales de Enfermedad , Temblor , Animales , Enfermedades del Sistema Nervioso Central/inducido químicamente , Enfermedades del Sistema Nervioso Central/genética , Harmalina , Humanos , Mamíferos , Especificidad de la Especie , Temblor/clasificación , Temblor/etiologíaRESUMEN
We have previously shown that (i) the ramified phenotype and (ii) the microglia-specific pattern of membrane currents are induced not only in microglia, but also in monocytes and macrophages if they are cultured in the presence of astrocytes. These findings indicated that microglia are not a separate type of cell of the myelomonocytic lineage, but are induced to take on their unique characteristics by astrocytes. Recently, it was discovered that the antibody 5-D-4 selectively stains ramified microglia in situ. We therefore studied the influence of astrocytes and other epithelial cells on the expression of the keratan sulfate epitope recognized by 5-D-4 in microglia and other myelomonocytic cells. Our findings show that this antigen is exclusively expressed in microglia only if they are induced to ramify by coculture with either astrocytes or epithelial cells. By contrast monocytes and macrophages, even if induced to take on the ramified phenotype do not stain positive with 5-D-4. These findings indicate (i) that 5-D-4 is a specific marker for ramified microglia in vitro, and (ii) that microglia are a separate class of myelomonocytic cells, distinct from monocytes and macrophages.