RESUMEN
The purpose of this pilot in vitro study was to evaluate the temperature increase in the pulp chamber of the teeth, during Er,Cr:YSGG bleaching, as well as to show which teeth are the most susceptible in terms of pulp temperature increase during laser-activated bleaching treatment. Although Er:YAG studies have been published on this subject, it is the first time Er,Cr:YSGG wavelength is tested. Fifteen teeth were tested--3 each of the following--(maxillary central incisors, lateral incisors, canines, premolars and mandibular incisors). The bleaching procedure comprised an Er,Cr:YSGG laser (2780 nm, Waterlase MD, Biolase, USA) and a yellow-coloured bleaching agent with a concentration of 38 % H2O2 (Power whitening, WHITEsmile GmbH, Germany). The tip used was a 6-mm long Z-type glass tip (MZ8) of a 800 µm diameter. Average output power was set to 1.25 W, pulse duration 700 µs (S-mode), whilst the pulse repetition rate was 10 Hz. The results showed that the most susceptible teeth in terms of pulp temperature increase were the lateral maxillary incisors and the mandibular incisors. The mean temperature increase on these teeth was 1.06 and 1.00 °C, respectively, on 60 s Er,Cr:YSGG-supported bleaching.
Asunto(s)
Cavidad Pulpar/efectos de la radiación , Láseres de Estado Sólido , Temperatura , Blanqueamiento de Dientes , Diente/efectos de la radiación , Cavidad Pulpar/efectos de los fármacos , Humanos , Proyectos Piloto , Diente/efectos de los fármacosRESUMEN
OBJECTIVE: The aim of this study was to evaluate the effectiveness of a radial firing tip of an Er,Cr:YSGG laser as an adjunct to a nonsurgical periodontal treatment. METHODS: Twelve patients with chronic or aggressive periodontitis were treated by conventional periodontal treatment using ultrasonic devices and hand instruments and, additionally, in two quadrants with an Er,Cr:YSGG laser. A new radial firing tip (RFPT 14-5, Biolase) was used with 1.5 W, 30 Hz, 11% air, 20% water, and pulse duration 140 µs. Microbiological smears were taken before treatment, one day after lasing, and three and six months after lasing. Pocket depths of all periodontal sites were measured before and six months after treatment. RESULTS: The total bacterial load of Prevotella intermedia, Tannerella forsythia, Treponema denticola, Fusobacterium nucleatum, Porphyromonas gingivalis, and Aggregatibacter actinomycetemcomitans inside the pocket was reduced significantly throughout the whole examination time. Greater pocket depth reductions were observed in all groups. There was a slight higher reduction of pocket depth in the lased group after six months. CONCLUSIONS: These results support the thesis that Er,Cr:YSGG laser supported periodontal treatment leads to a significant reduction of periopathogenes and thereby helps the maintenance of periodontal health.
Asunto(s)
Láseres de Estado Sólido/uso terapéutico , Terapia por Luz de Baja Intensidad/métodos , Bolsa Periodontal/microbiología , Bolsa Periodontal/radioterapia , Carga Bacteriana/métodos , Femenino , Humanos , Masculino , Bolsa Periodontal/diagnóstico , Proyectos PilotoRESUMEN
The purpose of this study was to evaluate the temperatures on the root surfaces during Nd:YAG laser irradiation in root canals using pulse durations of 180 and 320 µs. Thirty extracted human teeth were used in this study. The teeth were enlarged up to ISO 40 (multi-rooted) or up to ISO 60 (single-rooted) by conventional technique using K-files. Then the teeth were placed into a water bath with a constant temperature of 37 °C and then irradiated with an Nd:YAG laser having an output power of 1.5 W, a frequency of 15 Hz, using an optic fiber of 200 µm diameter. The temperature on the root surface was measured by means of attaching thermocouples in three areas (coronal, mesial, and apical regions) of the root canals. The thermographic study showed that the average temperature elevation for both pulse durations on the root surfaces was less than 9 °C. There was no significant difference in the observed temperatures in coronal and mesial areas. Though a higher increase of temperature was observed in the apical region when the pulse length of the Nd:YAG laser was 320 µs. The results of the study showed that the temperature rises during Nd:YAG laser irradiation with parameters used in this study minimal to cause damage on bone and periodontal tissues. Moreover, it was suggested that in order to have lower temperature in the apical region, an Nd:YAG laser with a pulse length of 180 µs is preferred than one with a pulse length of 320 µs.
Asunto(s)
Láseres de Estado Sólido/uso terapéutico , Terapia por Luz de Baja Intensidad , Preparación del Conducto Radicular/métodos , Raíz del Diente/efectos de la radiación , Calor/efectos adversos , Humanos , Láseres de Estado Sólido/efectos adversos , Terapia por Luz de Baja Intensidad/efectos adversos , Preparación del Conducto Radicular/efectos adversos , Raíz del Diente/lesiones , Raíz del Diente/fisiologíaRESUMEN
STUDY DESIGN: This was an experimental study. OBJECTIVES: White matter sparing influences locomotor recovery after traumatic spinal cord injury (SCI). The objective of the present post-mortem magnetic resonance imaging (MRI) investigation was to assess the potential of a simple inversion recovery (IR) sequence in combination with high-resolution proton density (PD) images to selectively depict spared white matter after experimental SCI in the rat. SETTING: This study was conducted at University of Liège and Centre Hospitalier Universitaire, Liège, Belgium and Hasselt University, Diepenbeek, Belgium. METHODS: Post-mortem 9.4 tesla (T) MRI was obtained from five excised rat spines 2 months after compressive SCI. The locomotor recovery had been followed weekly using the standardized Basso-Beattie-Bresnahan scale. IR MRI was used to depict normal white matter as very hypo-intense. Preserved white matter, cord atrophy and lesion volume were assessed, and histology was used to confirm MRI data. RESULTS: MRI showed lesion severity and white matter sparing in accordance with the degree of locomotor recovery. IR MRI enhanced detection of spared and injured white matter by selectively altering the signal of spared white matter. Even subtle white matter changes could be detected, increasing diagnostic accuracy as compared to PD alone. MRI accuracy was confirmed by histology. CONCLUSION: High-resolution IR-supported PD MRI provides useful micro-anatomical information about white matter damage and sparing in the post-mortem assessment of chronic rat SCI.
Asunto(s)
Fibras Nerviosas Mielínicas/patología , Recuperación de la Función/fisiología , Traumatismos de la Médula Espinal/patología , Médula Espinal/patología , Animales , Atrofia , Evaluación de la Discapacidad , Modelos Animales de Enfermedad , Imagen por Resonancia Magnética/métodos , Vías Nerviosas/lesiones , Vías Nerviosas/patología , Protones , Ratas , Traumatismos de la Médula Espinal/diagnóstico , Traumatismos de la Médula Espinal/mortalidadRESUMEN
Although CO(2) laser irradiation can decrease enamel demineralisation, it has still not been clarified which laser wavelength and which irradiation conditions represent the optimum parameters for application as preventive treatment. The aim of the present explorative study was to find low-fluence CO(2) laser (lambda = 10.6 microm) parameters resulting in a maximum caries-preventive effect with the least thermal damage. Different laser parameters were systematically evaluated in 3 steps. In the first experiment, 5 fluences of 0.1, 0.3, 0.4, 0.5 and 0.6 J/cm(2), combined with high repetition rates and 10 micros pulse duration, were chosen for the experiments. In a second experiment, the influence of different pulse durations (5, 10, 20, 30 and 50 micros) on the demineralisation of dental enamel was assessed. Finally, 3 different irradiation times (2, 5 and 9 s) were tested in a third experiment. In total, 276 bovine enamel blocks were used for the experiments. An 8-day pH-cycling regime was performed after the laser treatment. Demineralisation was assessed by lesion depth measurements with a polarised light microscope, and morphological changes were assessed with a scanning electron microscope. Irradiation with 0.3 J/cm(2), 5 micros, 226 Hz for 9 s (2,036 overlapping pulses) increased caries resistance by up to 81% compared to the control and was even significantly better than fluoride application (25%, p < 0.0001). Scanning electron microscopy examination did not reveal any obvious damage caused by the laser irradiation.
Asunto(s)
Susceptibilidad a Caries Dentarias/efectos de la radiación , Caries Dental/prevención & control , Esmalte Dental/efectos de la radiación , Dureza/efectos de la radiación , Láseres de Gas/uso terapéutico , Animales , Bovinos , Estudios Transversales , Terapia por Láser/instrumentación , Terapia por Láser/métodos , Modelos Lineales , Estadísticas no Paramétricas , Desmineralización Dental/prevención & control , Desmineralización Dental/radioterapiaRESUMEN
OBJECTIVE: The aim of this in vitro study was to investigate the antibacterial depth effect of continuous wave laser irradiation with a wavelength of 980 nm in the root canal wall dentin of bovine teeth. BACKGROUND DATA: The long-term success of an endodontic therapy often fails due to remaining bacteria in the root canal or dentin tubules, which cannot be sufficiently eliminated through the classical root canal preparation technique nor through rinsing solutions. MATERIALS AND METHODS: A total of 102 slices of bovine root dentin of different thicknesses (100, 300 and 500 micro m) were prepared. The samples were inoculated from one side with 5 micro L of an enterococcus faecalis suspension of defined concentration. Four slices per slice thickness served as a control group; the rest of the 30 slices per thickness were subjected to laser irradiation - 10 each of these slices were irradiated with distal outputs of 1.75, 2.3, and 2.8 Watts (W). After drying them for 30 sec, the back of the inoculated dentin slice was irradiated for 32 seconds with a 200- micro m fiber optical waveguide under constant movement of the fibers. The remaining bacteria were then detached in NaCl under vibration. The eluate produced by this was - taking account of the degree of dilution - plated out on sheep blood agar plates. After 24 h of incubation, the grown bacterial colonies were able to be counted out and evaluated. By doing so, they were compared with the non-irradiated, but otherwise identically treated control group. RESULTS: With a slice thickness of 100 micro m, the 980-nm diode laser achieved a maximum bacterial reduction of 95% at 1.75 W, 96% at 2.3 W, and 97% at 2.8 W. With a slice thickness of 300 micro m, a maximum of 77% of the bacteria was destroyed at 1.75 W, 87% at 2.3 W, and 89% at 2.8 W. The maximum bacterial reduction with a slice thickness of 500 micro m was 57% at 1.75 W, 66% at 2.3 W, and 86% at 2.8 W. CONCLUSION: The results of this research show that the 980-nm diode laser can eliminate bacteria that have immigrated deep into the dentin, thus being able to increase the success rate in endodontic therapy.
Asunto(s)
Bacterias/efectos de la radiación , Cavidad Pulpar/microbiología , Dentina/microbiología , Rayos Láser , Animales , Bovinos , Cavidad Pulpar/efectos de la radiación , Dentina/efectos de la radiación , Enterococcus faecalis/efectos de la radiación , Técnicas In VitroRESUMEN
Axons undergo Wallerian degeneration distal to a point of injury. Experimental investigations have documented many of the cellular and molecular events that underlie this behaviour. Since relatively little is known about such events in human CNS pathologies and current experimental intervention strategies indicate the possibility of significant axon regeneration along the original degenerated fibre tract, we performed an immunohistochemical investigation of the dynamics of Wallerian degeneration in post mortem spinal cords of patients who died 2 days to 30 years after either cerebral infarction or traumatic spinal cord injury. Neurofilament (NF) staining demonstrated a spatio-temporal pattern of axonal loss within degenerating descending nerve fibre tracts that could be detected close to the lesion as early as 12 days after injury and progressed to an almost complete loss of NF immunoreactivity at survival times of 1 year and longer. Immunohistochemistry for glial fibrillary acidic protein revealed a late astrocytic reaction starting at 4 months after injury in the degenerating tracts, leading to the long-term deposition of a dense astrocytic scar. These events were accompanied by the gradual reduction of myelin basic protein in affected nerve fibre tracts, leading to almost complete loss by 3 years after injury. Since the extracellular matrix molecule chondroitin sulphate proteoglycan (CSPG) is known to be strongly inhibitory for axonal regeneration and to be a major component of gliotic scar tissues, we investigated the possible deposition of CSPG within the degenerating nerve fibre tracts. Apart from a local up-regulation close to the lesion site, our results show no enhanced CSPG expression within degenerated tracts at any survival time. This suggests that despite the apparent lack of CSPG in Wallerian degeneration, the slow reduction of CNS myelin and the long-term deposition of a dense astrocytic scar may present an environment that is non-supportive for axon regrowth.
Asunto(s)
Astrocitos/patología , Infarto Cerebral/patología , Vaina de Mielina/patología , Traumatismos de la Médula Espinal/patología , Degeneración Walleriana/patología , Adulto , Anciano , Anciano de 80 o más Años , Infarto Cerebral/metabolismo , Proteoglicanos Tipo Condroitín Sulfato/metabolismo , Cicatriz/metabolismo , Cicatriz/patología , Humanos , Persona de Mediana Edad , Proteínas de Neurofilamentos/metabolismo , Traumatismos de la Médula Espinal/metabolismo , Factores de Tiempo , Regulación hacia Arriba , Degeneración Walleriana/metabolismoRESUMEN
The analgesic effect of vagus nerve stimulation (VNS) has not yet been demonstrated in animals with the devices used in the clinic. We studied in awake rats the effects of two VNS protocols on the hind paw hot water test and compared the results with those previously obtained in the oro-facial formalin test. A stringent duty cycle (20 s on/18 s off) increased heat pain tolerance in both hind paws (average 188%) after 2 h of stimulation. VNS with parameters used in epilepsy (30 s on/5 min off) decreased heat tolerance after 2 h, but produced a significant antinociceptive effect after days of stimulation. VNS may thus be useful in pain disorders, even with the less stringent protocol.
Asunto(s)
Terapia por Estimulación Eléctrica/métodos , Manejo del Dolor , Umbral del Dolor/fisiología , Nervio Vago/fisiología , Animales , Encéfalo/fisiología , Vías Eferentes/fisiología , Calor/efectos adversos , Masculino , Inhibición Neural/fisiología , Dolor/fisiopatología , Dimensión del Dolor , Ratas , Ratas Wistar , Tiempo de Reacción/fisiología , Tractos Espinotalámicos/fisiologíaRESUMEN
Uniform dosimetry is a prerequisite for reproducible laser applications in research and practice. The light-tissue interaction is dependent on the absorbed energy (J) per unit of time (tau) in the case of pulsed lasers, and on the absorbed power (W) per unit of volume (e.g. mm3) in the case of continuous-wave (cw) lasers, and thus directly dependent on the energy distribution within the laser beam. Consequently, precise knowledge of the spatial beam profile, and of the pulse duration and treatment time, is indispensable. The objective of this paper was a theoretical study of the impact of different mode profiles on energy distribution in the beam. Also examined was the question of the influence of changes in the laser parameters on the mode structure. Three erbium:YAG lasers (lambda=2.94 microm) were used for this purpose. The transversal mode structure of the lasers was observed by irradiating thermal paper and verified by means of calculations. The effect induced in the mode profile by changing the pulse energy and pulse repetition rate was investigated. The results of the tests show that changes in the laser parameters result in jumps in the transversal modes and associated energy distributions in the beam. The experiments confirm that simply changing the transversal modes has a substantial effect on the threshold energy required for the ablation of dental enamel (50 mJ with TEM00, 22.6 mJ with TEM31). In practice, inhomogeneity makes it impossible to determine the irradiated area in order to calculate the energy or power density. In addition, the energy distribution in the beam changes as a result of variation of the laser output energy and the pulse repetition rate. Consequently, simply measuring the beam diameter yields a totally incorrect result for the applied flux density when using a beam profile with a relatively high mode.
Asunto(s)
Esmalte Dental/efectos de la radiación , Rayos Láser , Esmalte Dental/ultraestructura , Relación Dosis-Respuesta en la Radiación , Erbio , Humanos , Técnicas In Vitro , Distribución Normal , Radiometría , Dispersión de RadiaciónRESUMEN
The scientific investigation of fundamental problems plays a decisive role in understanding the mode of action and the consequences of the use of lasers on biological material. One of these fundamental aspects is the investigation of the ablation threshold of various laser wavelengths in dental enamel. Knowledge of the relationships and influencing factors in the laser ablation of hard tooth tissue constitutes the basis for use in patients and the introduction of new indications. The present paper examines the ablation threshold of an Er:YAG laser (lambda=2.94 micro m) and an Er:YSGG laser (lambda=2.79 micro m) in human dental enamel. To this end, 130 enamel samples were taken from wisdom teeth and treated with increasing energy densities of 2-40 J/cm(2). The sample material was mounted and irradiated on an automated linear micropositioner. Treatment was performed with a pulse duration of tau(P(FWHM)) approximately 150 micro s and a pulse repetition rate of 5 Hz for both wavelengths. The repetition rate of the laser and the feed rate of the micropositioner resulted in overlapping of the single pulses. The surface changes were assessed by means of reflected light and scanning electron microscopy. On the basis of the results, it was possible to identify an energy density range as the ablation threshold for both the Er:YAG and the Er:YSGG laser. With the Er:YAG laser, the transition was found in an energy density range of 9-11 J/cm(2). The range for the Er:YSGG laser was slightly higher at 10-14 J/cm(2).
Asunto(s)
Esmalte Dental/efectos de la radiación , Rayos Láser , Esmalte Dental/ultraestructura , Erbio , Humanos , Técnicas In Vitro , Microscopía Electrónica de RastreoRESUMEN
The present study examines the dependence of the ablation threshold on the duration of the applied laser pulses in the dental enamel of human wisdom teeth. To this end, 600 treatments with the Er:YAG laser (lambda=2940 nm) were carried out on a total of 50 extracted teeth. The laser light was coupled into a fluoride glass light guide for this purpose, in order to ensure almost gaussian distribution of the light in a radially symmetrical beam. The beam diameter on the specimen was 610 micro m. The radiant exposure on the tooth surface was varied between 2 and 20 J/cm(2), while the duration of the pulses applied was changed in four steps from 100 micro s to 700 micro s. The irradiated tooth surfaces were examined for visible signs of ablation under a reflected-light microscope. The experiments revealed that, when pulses of shorter duration are used, the limit at which ablation sets in is reduced by up to approx. 3 J/cm(2). This expands the ablation threshold range of Er:YAG laser radiation to between 6 and 10 J/cm(2). In this context, both the pulse duration and the radiant exposure have a statistically significant influence on the ablation threshold (logistic regression, p<0.0001). Although the ablation threshold of the dental enamel can be changed by varying the pulse duration of the Er:YAG laser, no clinical consequences can be expected, as the shift is only slight.
Asunto(s)
Esmalte Dental/efectos de la radiación , Rayos Láser , Erbio , Humanos , Técnicas In Vitro , Dosis de RadiaciónRESUMEN
Myelin-associated glycoprotein (MAG) is expressed in periaxonal membranes of myelinating glia where it is believed to function in glia-axon interactions by binding to a component of the axolemma. Experiments involving Western blot overlay and coimmunoprecipitation demonstrated that MAG binds to a phosphorylated neuronal isoform of microtubule-associated protein 1B (MAP1B) expressed in dorsal root ganglion neurons (DRGNs) and axolemma-enriched fractions from myelinated axons of brain, but not to the isoform of MAP1B expressed by glial cells. The expression of some MAP1B as a neuronal plasma membrane glycoprotein (Tanner, S.L., R. Franzen, H. Jaffe, and R.H. Quarles. 2000. J. Neurochem. 75:553-562.), further documented here by its immunostaining without cell permeabilization, is consistent with it being a binding partner for MAG on the axonal surface. Binding sites for a MAG-Fc chimera on DRGNs colocalized with MAP1B on neuronal varicosities, and MAG and MAP1B also colocalized in the periaxonal region of myelinated axons. In addition, expression of the phosphorylated isoform of MAP1B was increased significantly when DRGNs were cocultured with MAG-transfected COS cells. The interaction of MAG with MAP1B is relevant to the known role of MAG in affecting the cytoskeletal structure and stability of myelinated axons.
Asunto(s)
Proteínas Asociadas a Microtúbulos/metabolismo , Glicoproteína Asociada a Mielina/metabolismo , Neuronas/metabolismo , Animales , Axones/química , Axones/metabolismo , Células COS , Técnicas de Cocultivo , Ganglios Espinales/citología , Proteínas Asociadas a Microtúbulos/análisis , Glicoproteína Asociada a Mielina/análisis , Glicoproteína Asociada a Mielina/genética , Neuroglía/metabolismo , Neuronas/química , Neuronas/ultraestructura , Fosforilación , Unión Proteica/fisiología , Ratas , TransfecciónRESUMEN
The Vilsmeier formylation has been introduced for the solid-phase functionalization of five different 2-carboxyindoles. The aldehyde functionality has been utilized in the preparation of O-benzylhydroxyureas.
Asunto(s)
Técnicas Químicas Combinatorias , Hidroxiurea/síntesis química , Indoles/síntesis química , Urea/análogos & derivados , Urea/síntesis química , Ácidos Carboxílicos/síntesis química , Diseño de Fármacos , Formiatos/química , Tecnología FarmacéuticaRESUMEN
As part of an ongoing lead discovery project we have developed a convenient method for the modification and substitution of indole moieties at the 3-position. Selective bromination of three different 2-carboxyindoles was followed by Suzuki cross-coupling with aryl and heteroaryl boronic acids on a Merrifield resin solid-phase. After column chromatography, yields of the 3- substituted indoles ranged from 42-98%.
Asunto(s)
Bromo , Indoles/química , Indoles/síntesis química , Ácidos Carboxílicos/síntesis química , Ácidos Carboxílicos/química , Espectroscopía de Resonancia Magnética , Modelos Moleculares , Conformación Molecular , Estructura Molecular , Espectrometría de Masa por Ionización de Electrospray , Relación Estructura-ActividadRESUMEN
This paper describes two-hybrid interactions amongst barley homeodomain proteins encoded by the Three Amino acid Loop Extension (TALE) superfamily. The class I KNOX protein BKN3 is shown to homodimerise and to associate with proteins encoded by the class I and II Knox genes BKn-1 and BKn-7. Furthermore, JUBEL1 and JUBEL2, two BELL1 homologous proteins, are identified and characterised as interacting partners of BKN3. Differences in the requirements of BKN3 derivatives for interactions with KNOX and JUBEL proteins imply the involvement of overlapping but slightly different domains. This set of results is an example for interactions amongst different classes of plant TALE homeodomain proteins, as previously described for related animal proteins. Apparently identical spatial and temporal expression patterns of BKn-1, BKn-3, BKn-7, JuBel1 and JuBel2, as determined by in situ hybridisation, are compatible with possible interactions of their protein products in planta. Contradictory to the common model, that the transcriptional down-regulation of certain class 1 Knox-genes is the prerequisite for organ differentiation, transcripts of all five genes were, similar to Tkn1 and Tkn2/LeT6 of tomato, detected in incipient and immature leaves as well as in meristematic tissues. A characteristic phenotype is induced by the overexpression of JuBel2 in transgenic tobacco plants.
Asunto(s)
Genes de Plantas , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Hordeum/metabolismo , Proteínas de Plantas , Secuencia de Aminoácidos , Secuencia de Bases , ADN Complementario , Regulación de la Expresión Génica de las Plantas , Proteínas de Homeodominio/química , Hordeum/genética , Datos de Secuencia Molecular , Plantas Modificadas Genéticamente/genética , Plantas Tóxicas , Unión Proteica , Nicotiana/genética , Técnicas del Sistema de Dos HíbridosRESUMEN
The lipid signaling molecule ceramide is formed by the action of acid and neutral sphingomyelinases and degraded by acid and neutral ceramidases. Short-term stimulation of mesangial cells with the pro-inflammatory cytokine interleukin-1beta (IL-1beta) leads to a rapid and transient increase in neutral sphingomyelinase activity (Kaszkin, M., Huwiler, A., Scholz, K., van den Bosch, H., and Pfeilschifter, J. (1998) FEBS Lett. 440, 163-166). In this study, we report on a second delayed peak of activation occurring after hours of IL-1beta treatment. This second phase of activation was first detectable after 2 h of treatment and steadily increased over the next 2 h, reaching maximal values after 4 h. In parallel, a pronounced increase in neutral ceramidase activity was observed, accounting for a constant or even decreased level of ceramide after long-term IL-1beta treatment, despite continuous sphingomyelinase activation. The increase in neutral ceramidase activity was due to expressional up-regulation, as detected by an increase in mRNA levels and enhanced de novo protein synthesis. The increase in neutral ceramidase protein levels and activity could be blocked dose- dependently by the p38 MAPK inhibitor SB 202190, whereas the classical MAPK pathway inhibitor U0126 and the protein kinase C inhibitor Ro 318220 were ineffective. Moreover, cotreatment of cells for 24 h with IL-1beta and SB 202190 led to an increase in ceramide formation. Interestingly, IL-1beta-stimulated neutral ceramidase activation was not reduced in mesangial cells isolated from mice deficient in MAPK-activated protein kinase-2, which is a downstream substrate of p38 MAPK, thus suggesting that the p38 MAPK-mediated induction of neutral ceramidase occurs independently of the MAPK-activated protein kinase-2 pathway. In summary, our results suggest a biphasic regulation of sphingomyelin hydrolysis in cytokine-treated mesangial cells with delayed de novo synthesis of neutral ceramidase counteracting sphingomyelinase activity and apoptosis. Neutral ceramidase may thus represent a novel cytoprotective enzyme for mesangial cells exposed to inflammatory stress conditions.
Asunto(s)
Amidohidrolasas/metabolismo , Mesangio Glomerular/efectos de los fármacos , Interleucina-1/farmacología , Amidohidrolasas/biosíntesis , Amidohidrolasas/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Células Cultivadas , Ceramidasas , Cartilla de ADN , Activación Enzimática , Mesangio Glomerular/citología , Mesangio Glomerular/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Datos de Secuencia Molecular , Ceramidasa Neutra , ARN Mensajero/genética , Ratas , Esfingomielina Fosfodiesterasa/genética , Regulación hacia Arriba , Proteínas Quinasas p38 Activadas por MitógenosRESUMEN
Recent evidence suggests that the sphingolipid-derived second messenger ceramide and oxidative stress are intimately involved in apoptosis induction. Here we report that exposure of microcapillary glomerular endothelial cells to superoxide-generating substances, including hypoxanthine/xanthine oxidase and the redox cyclers DMNQ and menadione results in a dose-dependent and delayed increase in the lipid signaling molecule ceramide. Long-term incubation of endothelial cells for 2-30 h with either DMNQ or hypoxanthine/xanthine oxidase leads to a continuous increase in ceramide levels. In contrast, short-term stimulation for 1 min up to 1 h had no effect on ceramide formation. The DMNQ-induced delayed ceramide formation is dose-dependently inhibited by reduced glutathione, whereas oxidized glutathione was without effect. Furthermore, N-acetylcysteine completely blocks DMNQ-induced ceramide formation. All superoxide-generating substances were found to dose-dependently trigger endothelial cell apoptosis. In addition, glutathione and N-acetylcysteine also prevented superoxide-induced apoptosis and implied that ceramide represents an important mediator of superoxide-triggered cell responses like apoptosis.
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Ceramidas/biosíntesis , Endotelio Vascular/metabolismo , Glomérulos Renales/metabolismo , Superóxidos/metabolismo , Acetilcisteína/farmacología , Animales , Antioxidantes/farmacología , Apoptosis/efectos de los fármacos , Bovinos , Células Cultivadas , Relación Dosis-Respuesta a Droga , Endotelio Vascular/citología , Endotelio Vascular/efectos de los fármacos , Glutatión/farmacología , Hipoxantina/metabolismo , Hipoxantina/farmacología , Glomérulos Renales/citología , Glomérulos Renales/efectos de los fármacos , Naftoquinonas/farmacología , Superóxidos/farmacología , Vitamina K/farmacología , Xantina Oxidasa/metabolismo , Xantina Oxidasa/farmacologíaRESUMEN
The first goal of this study was to examine the influence that poly(ethylene oxide)-block-poly(D,L-lactide) (PELA) copolymer can have on the wettability, the in vitro controlled delivery capability, and the degradation of poly(D,L-lactide) (PDLLA) foams. These foams were prepared by freeze-drying and contain micropores (10 microm) in addition of macropores (100 microm) organized longitudinally. Weight loss, water absorption, changes in molecular weight, polymolecularity (Mw/Mn) and glass transition temperature (Tg) of PDLLA foams mixed with various amounts of PELA were followed with time. It was found that 10wt% of PELA increased the wettability and the degradation rate of the polymer foams. The release of sulforhodamine (SR) was compared for PDLLA and PDLLA-PELA foams in relation with the foam porosity. An initial burst release was observed only in the case of the 90:10 PDLLA/PELA foam. The ability of the foam of this composition to be integrated and to promote tissue repair and axonal regeneration in the transected rat spinal cord was investigated. After implantation of ca. 20 polymer rods assembled with fibrin-glue, the polymer construct was able to bridge the cord stumps by forming a permissive support for cellular migration, angiogenesis and axonal regrowth.
Asunto(s)
Materiales Biocompatibles , Factor 1 de Crecimiento de Fibroblastos/administración & dosificación , Regeneración Nerviosa/fisiología , Poliésteres , Polietilenglicoles , Traumatismos de la Médula Espinal/fisiopatología , Traumatismos de la Médula Espinal/terapia , Animales , Técnicas In Vitro , Ensayo de Materiales , Microscopía Electrónica de Rastreo , Ratas , Ratas Wistar , Rodaminas/administración & dosificación , Rodaminas/farmacocinética , Traumatismos de la Médula Espinal/patologíaRESUMEN
The ability of DRG-derived neurons to survive and attach onto macroporous polylactide (PLA) foams was assessed in vitro. The foams were fabricated using a thermally induced polymer-solvent phase separation. Two types of pore structures, namely oriented or interconnected pores, can be produced, depending on the mechanism of phase separation, which in turn can be predicted by the thermodynamics of the polymer-solvent pair. Coating of the porous foams with polyvinylalcohol (PVA) considerably improved the wettability of the foams and allowed for cell culture. The in vitro biocompatibility of the PVA-coated supports was demonstrated by measuring cell viability and neuritogenesis. Microscopic observations of the cells seeded onto the polymer foams showed that the interconnected pore networks were more favorable to cell attachment than the anisotropic ones. The capacity of highly oriented foams to support in vivo peripheral nerve regeneration was studied in rats. A sciatic nerve gap of 5-mm length was bridged with a polymer implant showing macrotubes of 100 microm diameter. At 4 weeks postoperatively, the polymer implant was still present. It was well integrated and had restored an anatomic continuity. An abundant cell migration was observed at the outer surface of the polymer implant, but not within the macrotubes. This dense cellular microenvironment was found to be favorable for axogenesis.
Asunto(s)
Materiales Biocompatibles/química , Regeneración Nerviosa , Poliésteres/química , Prótesis e Implantes , Nervio Ciático/fisiología , Animales , Biodegradación Ambiental , Adhesión Celular , Materiales Biocompatibles Revestidos/química , Ganglios Espinales/citología , Ensayo de Materiales , Microscopía Confocal , Microscopía Electrónica de Rastreo , Neuritas/ultraestructura , Neuronas Aferentes/citología , Alcohol Polivinílico/química , Porosidad , Ratas , Ratas Wistar , Nervio Ciático/lesiones , Propiedades de SuperficieRESUMEN
We determined the mutation spectra in Salmonella of four chlorinated butenoic acid analogues (BA-1 through BA-4) of the drinking water mutagen 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX) and compared the results with those generated previously by us for MX and a related compound, MCF. We then considered relationships between the properties of mutagenic potency and mutational specificity for these six chlorinated butenoic acid analogues. In TA98, the three most potent mutagens, BA-3, BA-4, MX, and the organic extract, all induced large percentages of complex frameshifts (33-67%), which distinguish these agents from any other class of compound studied previously. In TA100, which has only GC sites for mutation recovery, >71% of the mutations induced by all of the agents were GC-->TA transversions. The availability of both GC and TA sites for mutation in TA104 resulted in greater distinctions in mutational specificity than in TA100. MX targeted GC sites almost exclusively (98%); the structurally similar BA-4 and BA-2 produced mutations at similar frequencies at both GC and AT sites; and the structurally similar BA-3 and BA-1 induced most mutations at AT sites (69%). Thus, large variations in structural properties influencing relative mutagenic potency appeared to be distinct from the more localized similar structural features influencing mutagenic specificity in TA104. Among a set of physicochemical properties examined for the six butenoic acids, a significant correlation was found between pK(a) and mutagenic potency in TA100, even when the unionized fraction of the activity dose was considered. In addition, a correlation in CLOGP for BA-1 to BA-4 suggested a role for bioavailability in determining mutagenic potency. These results illustrate the potential value of structural analyses for exploring the relationship between chemical structure and mutational mechanisms. To our knowledge, this is the first study in which such analyses have been applied to structural analogues for which both mutagenic potency and mutation spectra date were available.