RESUMEN

The present review was carried out to describe publications on the use of methylene blue (MB) in pediatrics and neonatology, discussing dose, infusion rate, action characteristics, and possible benefits for a pediatric patient group. The research was performed on the data sources PubMed, BioMed Central, and Embase (updated on Aug 31, 2020) by two independent investigators. The selected articles included human studies that evaluated MB use in pediatric or neonatal patients with vasoplegia due to any cause, regardless of the applied methodology. The MB use and 0 to 18-years-old patients with vasodilatory shock were the adopted criteria. Exclusion criteria were the use of MB in patients without vasoplegia and patients ≥ 18-years-old. The primary endpoint was the increase in mean arterial pressure (MAP). Side effects and dose were also evaluated. Eleven studies were found, of which 10 were case reports, and 1 was a randomized clinical study. Only two of these studies were with neonatal patients (less than 28 days-old), reporting a small number of cases (1 and 6). All studies described the positive action of MB on MAP, allowing the decrease of vasoactive amines in several of them. No severe side effects or death related to the use of the medication were reported. The maximum dose used was 2 mg/kg, but there was no consensus on the infusion rate and drug administration timing. Finally, no theoretical or experimental basis sustains the decision to avoid MB in children claiming it can cause pulmonary hypertension. The same goes for the concern of a possible deleterious effect on inflammatory distress syndrome.

Asunto(s)

Pediatría , Vasoplejía , Adolescente , Niño , Hemodinámica , Humanos , Recién Nacido , Azul de Metileno/efectos adversos , Azul de Metileno/uso terapéutico , Ensayos Clínicos Controlados Aleatorios como Asunto , Vasoplejía/inducido químicamente , Vasoplejía/tratamiento farmacológico

RESUMEN

A bioactive compound isolated from the stem extract of Aristolochia sprucei through High Performance Liquid Chromatography (HPLC) was identified via Nuclear Magnetic Resonance (NMR) as the aristolochic acid (AA). This compound showed an inhibitory effect over the myotoxic activity of Bothrops jararacussu and Bothrops asper venoms, being also effective against the indirect hemolytic activity of B. asper venom. Besides, AA also inhibited the myotoxic activity of BthTX-I and MTX-II with an efficiency greater than 60% against both myotoxins. Docking predictions revealed an interesting mechanism, through which the AA displays an interaction profile consistent with its inhibiting abilities, binding to both active and putative sites of svPLA2. Overall, the present findings indicate that AA may bind to critical regions of myotoxic Asp 49 and Lys49-PLA2s from snake venoms, highlighting the relevance of domains comprising the active and putative sites to inhibit these toxins.

RESUMEN

Rauwolfia vomitoria Afzel. is an antipsychotic plant used by several African communities in the management of psychiatric conditions with good outcomes. Concerns about its dosages on brain activity lead to this investigation of its action on the hippocampal microstructure.Twenty-four adult male Wistar rats of average weight 200 g, were assigned into four groups (n = 6): control; 200, 300 and 400 mg/kg body weight of RVroot bark extract, respectively. The administration was once daily, and orally for seven days. Daily observation of the animals was done till on day eight when they were sacrificed after deep anaesthesia. Each brain was processed for histology and immunohistochemical studies. Animals in the 200, 300 and 400 mg/kg RV groups appeared generally dull and drowsy, and barely fed. Their hippocampal histology showed neuronal atrophy and karyorrhexis, with no difference in cell count, although the pyramidal cell numbers decreased in the 300 and 400 mg/kg RV groups. Neuron-specific enolase decreased in the 400 mg/kg RV group, while neurofilament decreased in all test groups. Glial fibrillary acidic protein expression and density increased in the 200 and 300 mg/kg RV groups, but not the 400 mg/kg RV group, all compared with the control group.The given doses of RV root bark extractin adult Wistar rats showed sedative activities with hippocampal histopathological changes, which may not be reversible, thereby leading to the hippocampal functional deficit.

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Introducción: Rauwolfia vomitoria (RV) Afzel es una planta antipsicótica utilizada por varias comunidades africanas en el tratamiento de enfermedades psiquiátricas con buenos resultados. Las preocupaciones sobre sus efecto sobre la actividad cerebral conducen a esta investigación de su acción sobre la microestructura del hipocampo.Materiales y métodos: Se asignaron veinticuatro ratas Wistar macho adultas de un peso medio de 200 g, en cuatro grupos (n = 6): control; 200, 300 y 400 mg / kg de peso corporal de extracto de corteza de raíz de RV, respectivamente. La administración fue una vez al día y por vía oral durante siete días. Se realizó una observación diaria de los animales hasta el día ocho, cuando fueron sacrificados después de una anestesia profunda. Cada cerebro fue procesado para estudios histológicos e inmunohistoquímicos.Resultados: Los animales en los grupos de RV de 200, 300 y 400 mg / kg parecían generalmente apagados y somnolientos, y apenas alimentados. Su histología hipocampal mostró atrofia neuronal y cariorrexis, sin diferencia en el recuento celular, aunque el número de células piramidales disminuyó en los grupos de RV de 300 y 400 mg / kg. La enolasa específica de neuronas disminuyó en el grupo de RV de 400 mg / kg, mientras que el neurofilamento disminuyó en todos los grupos de prueba. La expresión y densidad de la proteína fibrilar ácida glial aumentó en los grupos de RV de 200 y 300 mg / kg, pero no en el grupo de RV de 400 mg / kg, todos en comparación con el grupo de control.Conclusión: Las dosis administradas de extracto de corteza de raíz de RV en ratas Wistar adultas mostraron actividades sedantes, con cambios histopatológicos del hipocampo, que pueden no ser reversibles, lo que conduce al déficit funcional del hipocampo.

Asunto(s)

Animales , Ratas , Rauwolfia/química , Extractos Vegetales/uso terapéutico , Hipocampo/anatomía & histología , Ratas Wistar

RESUMEN

The nitric oxide/cyclic guanosine monophosphate (NO/cGMP) pathway has a significative influence in hemodynamic changes that occur in transplants. Classically, the ischemia-reperfusion syndrome (IRS) is characterized by hypotension and low vascular resistance, when cGMP and nitric oxide (NO) are increased, contributing to oxidative stress, within an inflammatory context. These mechanisms occur in several types of transplants, such as liver, heart, lungs, kidney, which are a therapeutic choice in several clinical conditions when conventional treatments failed. It is well known the significant relation between graft dysfunction or rejection and ischemia-reperfusion injury that is linked to inflammatory response and NO/cGMP pathway activation. This review aims to study the NO/cGMP pathway in solid organ transplants. Finally, we inquire whether physicians do not underestimate the NO/cGMP pathway.

Asunto(s)

GMP Cíclico , Azul de Metileno , Óxido Nítrico , Trasplantes , GMP Cíclico/metabolismo , Hemodinámica , Humanos , Hipotensión , Hígado/metabolismo , Pulmón/fisiopatología , Óxido Nítrico/metabolismo , Reperfusión , Daño por Reperfusión/fisiopatología , Transducción de Señal , Trasplantes/metabolismo , Resistencia Vascular

RESUMEN

Evidence-based review of the existing literature ultimately recommends stocking of Methylene Blue (MB) as an emergency antidote in the United States. The same is reported around the world in Japan, Greece, Italy and Canada. The observation that MB is always present as the main antidote required in emergency and critical care units calls for a revisit on its effects on the NO/cGMP system to reemphasize its multisystem actions. Therefore, the present review aimed to display the role of MB in emergency units, concerning: 1) Polytrauma and circulatory shock; 2) Neuroprotection, 3) Anaphylaxis and, 4) Overdose and poisoning.

Asunto(s)

Antídotos/uso terapéutico , Cuidados Críticos/métodos , Azul de Metileno/uso terapéutico , Anafilaxia/tratamiento farmacológico , Ensayos Clínicos como Asunto , Sobredosis de Droga/tratamiento farmacológico , Servicio de Urgencia en Hospital , Medicina Basada en la Evidencia , Humanos , Traumatismo Múltiple/tratamiento farmacológico , Enfermedades del Sistema Nervioso/tratamiento farmacológico , Intoxicación/tratamiento farmacológico

RESUMEN

MLH1 and PMS2 proteins form the MutLα heterodimer, which plays a major role in DNA mismatch repair (MMR) in humans. Mutations in MMR-related proteins are associated with cancer, especially with colon cancer. The N-terminal region of MutLα comprises the N-termini of PMS2 and MLH1 and, similarly, the C-terminal region of MutLα is composed by the C-termini of PMS2 and MLH1, and the two are connected by linker region. The nuclear localization sequences (NLSs) necessary for the nuclear transport of the two proteins are found in this linker region. However, the exact NLS sequences have been controversial, with different sequences reported, particularly for MLH1. The individual components are not imported efficiently, presumably due to their C-termini masking their NLSs. In order to gain insights into the nuclear transport of these proteins, we solved the crystal structures of importin-α bound to peptides corresponding to the supposed NLSs of MLH1 and PMS2 and performed isothermal titration calorimetry to study their binding affinities. Both putative MLH1 and PMS2 NLSs can bind to importin-α as monopartite NLSs, which is in agreement with some previous studies. However, MLH1-NLS has the highest affinity measured by a natural NLS peptide, suggesting a major role of MLH1 protein in nuclear import compared to PMS2. Finally, the role of MLH1 and PMS2 in the nuclear transport of the MutLα heterodimer is discussed.

Asunto(s)

Núcleo Celular/metabolismo , Reparación de la Incompatibilidad de ADN , Endonucleasa PMS2 de Reparación del Emparejamiento Incorrecto/metabolismo , Homólogo 1 de la Proteína MutL/metabolismo , Transporte Activo de Núcleo Celular , Secuencia de Aminoácidos , Animales , Humanos , Carioferinas/metabolismo , Ratones , Endonucleasa PMS2 de Reparación del Emparejamiento Incorrecto/química , Modelos Moleculares , Homólogo 1 de la Proteína MutL/química , Conformación Proteica

RESUMEN

The Neurospora crassa NIT-2 transcription factor belongs to the GATA transcription factor family and plays a fundamental role in the regulation of nitrogen metabolism. Because NIT-2 acts by accessing DNA inside the nucleus, understanding the nuclear import process of NIT-2 is necessary to characterize its function. Thus, in the present study, NIT-2 nuclear transport was investigated using a combination of biochemical, cellular, and biophysical methods. A complemented strain that produced an sfGFP-NIT-2 fusion protein was constructed, and nuclear localization assessments were made under conditions that favored protein translocation to the nucleus. Nuclear translocation was also investigated using HeLa cells, which showed that the putative NIT-2 nuclear localization sequence (NLS; 915TISSKRQRRHSKS927) was recognized by importin-α and that subsequent transport occurred via the classical import pathway. The interaction between the N. crassa importin-α (NcImpα) and the NIT-2 NLS was quantified with calorimetric assays, leading to the observation that the peptide bound to two sites with different affinities, which is typical of a monopartite NLS sequence. The crystal structure of the NcImpα/NIT-2 NLS complex was solved and revealed that the NIT-2 peptide binds to NcImpα with the major NLS-binding site playing a primary role. This result contrasts other recent studies that suggested a major role for the minor NLS-binding site in importin-α from the α2 family, indicating that both sites can be used for different cargo proteins according to specific metabolic requirements.

Asunto(s)

Transporte Activo de Núcleo Celular/fisiología , Proteínas de Unión al ADN/metabolismo , Proteínas Fúngicas/metabolismo , Neurospora crassa/metabolismo , Factores de Transcripción/metabolismo , alfa Carioferinas/metabolismo , Secuencia de Aminoácidos , Sitios de Unión/fisiología , Células Cultivadas , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/genética , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Células HeLa , Humanos , Neurospora crassa/genética , Estructura Secundaria de Proteína , Esporas Fúngicas , Factores de Transcripción/química , Factores de Transcripción/genética , Difracción de Rayos X , alfa Carioferinas/química , alfa Carioferinas/genética

RESUMEN

Plant aquaporins are water channels implicated in various physiological processes, including growth, development and adaptation to stress. In this study, the Tonoplast Intrinsic Protein (TIP) gene subfamily of Eucalyptus, an economically important woody species, was investigated and characterized. A genome-wide survey of the Eucalyptus grandis genome revealed the presence of eleven putative TIP genes (referred as EgTIP), which were individually assigned by phylogeny to each of the classical TIP1-5 groups. Homology modeling confirmed the presence of the two highly conserved NPA (Asn-Pro-Ala) motifs in the identified EgTIPs. Residue variations in the corresponding selectivity filters, that might reflect differences in EgTIP substrate specificity, were observed. All EgTIP genes, except EgTIP5.1, were transcribed and the majority of them showed organ/tissue-enriched expression. Inspection of the EgTIP promoters revealed the presence of common cis-regulatory elements implicated in abiotic stress and hormone responses pointing to an involvement of the identified genes in abiotic stress responses. In line with these observations, additional gene expression profiling demonstrated increased expression under polyethylene glycol-imposed osmotic stress. Overall, the results obtained suggest that these novel EgTIPs might be functionally implicated in eucalyptus adaptation to stress.

RESUMEN

AIM: To set up and test the feasibility of a handmade apparatus adapted for exhaled breath condensate (EBC) collection in medium-sized animals. MATERIALS AND METHODS: The apparatus was produced using an 18-mm thick u-shaped borosilicate glass. The u-shaped tube body is 25 cm in diameter, and the horizontal portions are 12 cm in diameter. The base consists of a tube joint 14/20 or 14 mm thick by 20 cm in diameter, and has a length of 5 cm. This has a hole that is plugged for condensate flow to a 1.5 mL polypropylene microtube that stores the condensate during collection. Was placed inside a styrofoam box and immersed in crushed ice and salt to ensure cooling. The temperature was monitored and maintained throughout the collection at -10°C. One of the outputs of the u-shaped tube was connected to the expiratory limb of the ventilator. RESULTS: An experimental model of ALI, induced by oleic acid (OA) was adopted to determine the concentration of biomarkers of oxidative stress: malondialdehyde (MDA), glutathione (GSH), and nitrite/nitrate (NOx). The proposed model allows measurement of NOx, MDA, and GSH. However, the NOx and MDA levels in the EBC were not significant. It was only possible to observe an upward trend, which suggests a temporal evolution of the presence of these markers in the EBC. CONCLUSION: The EBC collection method adapted is effective to generate sufficient content that allows to determine the levels of different biomarkers, such as NOx, MDA, and GSH, that are involved in oxidative and inflammatory stress processes during respiratory diseases.

Asunto(s)

Lesión Pulmonar Aguda/metabolismo , Modelos Animales de Enfermedad , Animales , Biomarcadores/análisis , Pruebas Respiratorias/instrumentación , Estudios de Factibilidad , Ácido Oléico , Conejos

RESUMEN

Neurospora crassa is a filamentous fungus that has been extensively studied as a model organism for eukaryotic biology, providing fundamental insights into cellular processes such as cell signaling, growth and differentiation. To advance in the study of this multicellular organism, an understanding of the specific mechanisms for protein transport into the cell nucleus is essential. Importin-α (Imp-α) is the receptor for cargo proteins that contain specific nuclear localization signals (NLSs) that play a key role in the classical nuclear import pathway. Structures of Imp-α from different organisms (yeast, rice, mouse, and human) have been determined, revealing that this receptor possesses a conserved structural scaffold. However, recent studies have demonstrated that the Impα mechanism of action may vary significantly for different organisms or for different isoforms from the same organism. Therefore, structural, functional, and biophysical characterization of different Impα proteins is necessary to understand the selectivity of nuclear transport. Here, we determined the first crystal structure of an Impα from a filamentous fungus which is also the highest resolution Impα structure already solved to date (1.75 Å). In addition, we performed calorimetric analysis to determine the affinity and thermodynamic parameters of the interaction between Imp-α and the classical SV40 NLS peptide. The comparison of these data with previous studies on Impα proteins led us to demonstrate that N. crassa Imp-α possess specific features that are distinct from mammalian Imp-α but exhibit important similarities to rice Imp-α, particularly at the minor NLS binding site.

Asunto(s)

Neurospora crassa/metabolismo , Señales de Localización Nuclear , alfa Carioferinas/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Modelos Moleculares , Neurospora crassa/genética , Señales de Localización Nuclear/química , Señales de Localización Nuclear/genética , Unión Proteica , Conformación Proteica , alfa Carioferinas/química , alfa Carioferinas/genética

RESUMEN

Importin-α recognizes cargo proteins that contain classical nuclear localization sequences (NLS) and, in complex with importin-ß, is able to translocate nuclear proteins through the nuclear pore complex. The filamentous fungus Neurospora crassa is a well studied organism that has been widely used as a model organism for fundamental aspects of eukaryotic biology, and is important for understanding the specific mechanisms of protein transport to the cell nucleus. In this work, the crystallization and preliminary X-ray diffraction analysis of importin-α from N. crassa (IMPα-Nc) complexed with a classical NLS peptide (SV40 NLS) are reported. IMPα-Nc-SV40 NLS crystals diffracted X-rays to 2.0 Šresolution and the structure was solved by molecular-replacement techniques, leading to a monomeric structure. The observation of the electron-density map indicated the presence of SV40 NLSs interacting at both the minor and major NLS-binding sites of the protein.

Asunto(s)

Cristalización/métodos , Cristalografía por Rayos X/métodos , Neurospora crassa/metabolismo , Oligopéptidos/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , alfa Carioferinas/química , alfa Carioferinas/metabolismo , Núcleo Celular/metabolismo , Unión Proteica , Proteínas Recombinantes/genética , alfa Carioferinas/genética

RESUMEN

The influenza virus has been a challenge to science due to its ability to withstand new environmental conditions. Taking into account the development of virus sequence databases, computational approaches can be helpful to understand virus behavior over time. Furthermore, they can suggest new directions to deal with influenza. This work presents triplet entropy analysis as a potential phylodynamic tool to quantify nucleotide organization of viral sequences. The application of this measure to segments of hemagglutinin (HA) and neuraminidase (NA) of H1N1 and H3N2 virus subtypes has shown some variability effects along timeline, inferring about virus evolution. Sequences were divided by year and compared for virus subtype (H1N1 and H3N2). The nonparametric Mann-Whitney test was used for comparison between groups. Results show that differentiation in entropy precedes differentiation in GC content for both groups. Considering the HA fragment, both triplet entropy as well as GC concentration show intersection in 2009, year of the recent pandemic. Some conclusions about possible flu evolutionary lines were drawn.

Asunto(s)

Glicoproteínas Hemaglutininas del Virus de la Influenza/genética , Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H3N2 del Virus de la Influenza A/genética , Neuraminidasa/genética , Composición de Base , Evolución Molecular , Humanos , Modelos Genéticos , Filogenia , Análisis de Secuencia de ADN , Estadísticas no Paramétricas , Termodinámica

RESUMEN

Neurospora crassa has been widely used as a model organism and contributed to the development of biochemistry and molecular biology by allowing the identification of many metabolic pathways and mechanisms responsible for gene regulation. Nuclear proteins are synthesized in the cytoplasm and need to be translocated to the nucleus to exert their functions which the importin-α receptor has a key role for the classical nuclear import pathway. In an attempt to get structural information of the nuclear transport process in N. crassa, we present herein the cloning, expression, purification and structural studies with N-terminally truncated IMPα from N. crassa (IMPα-Nc). Circular dichroism analysis revealed that the IMPα-Nc obtained is correctly folded and presents a high structural conservation compared to other importins-α. Dynamic light scattering, analytical size-exclusion chromatography experiments and molecular dynamics simulations indicated that the IMPα-Nc unbound to any ligand may present low stability in solution. The IMPα-Nc theoretical model displayed high similarity of its inner concave surface, which binds the cargo proteins containing the nuclear localization sequences, among IMPα from different species. However, the presence of non-conserved amino acids relatively close to the NLS binding region may influence the binding specificity of IMPα-Nc to cargo proteins.

Asunto(s)

Núcleo Celular/metabolismo , Señales de Localización Nuclear/metabolismo , alfa Carioferinas/metabolismo , Secuencia de Aminoácidos , Cromatografía en Gel , Dicroismo Circular , Clonación Molecular , Ligandos , Modelos Moleculares , Simulación de Dinámica Molecular , Neurospora crassa/metabolismo , Estabilidad Proteica , Alineación de Secuencia , alfa Carioferinas/química , alfa Carioferinas/genética , alfa Carioferinas/aislamiento & purificación , beta Carioferinas/metabolismo

RESUMEN

Flap endonuclease 1 (FEN1) is a member of the nuclease family and is structurally conserved from bacteriophages to humans. This protein is involved in multiple DNA-processing pathways, including Okazaki fragment maturation, stalled replication-fork rescue, telomere maintenance, long-patch base-excision repair and apoptotic DNA fragmentation. FEN1 has three functional motifs that are responsible for its nuclease, PCNA-interaction and nuclear localization activities, respectively. It has been shown that the C-terminal nuclear localization sequence (NLS) facilitates nuclear localization of the enzyme during the S phase of the cell cycle and in response to DNA damage. To determine the structural basis of the recognition of FEN1 by the nuclear import receptor importin α, the crystal structure of the complex of importin α with a peptide corresponding to the FEN1 NLS was solved. Structural studies confirmed the binding of the FEN1 NLS as a classical bipartite NLS; however, in contrast to the previously proposed (354)KRKX(8)KKK(367) sequence, it is the (354)KRX(10)KKAK(369) sequence that binds to importin α. This result explains the incomplete inhibition of localization that was observed on mutating residues (365)KKK(367). Acidic and polar residues in the X(10) linker region close to the basic clusters play an important role in binding to importin α. These results suggest that the basic residues in the N-terminal basic cluster of bipartite NLSs may play roles that are more critical than those of the many basic residues in the C-terminal basic cluster.

Asunto(s)

Endonucleasas de ADN Solapado/metabolismo , Señales de Localización Nuclear/metabolismo , alfa Carioferinas/metabolismo , Transporte Activo de Núcleo Celular , Secuencia de Aminoácidos , Animales , Sitios de Unión , Cristalografía por Rayos X , Endonucleasas de ADN Solapado/química , Humanos , Ratones , Modelos Moleculares , Datos de Secuencia Molecular , Señales de Localización Nuclear/química , Unión Proteica , Conformación Proteica , alfa Carioferinas/química

RESUMEN

Ku70 and Ku80 form a heterodimeric complex involved in multiple nuclear processes. This complex plays a key role in DNA repair due to its ability to bind DNA double-strand breaks and facilitate repair by the nonhomologous end-joining pathway. Ku70 and Ku80 have been proposed to contain bipartite and monopartite nuclear localization sequences (NLSs), respectively, that allow them to be translocated to the nucleus independently of each other via the classical importin-α (Impα)/importin-ß-mediated nuclear import pathway. To determine the structural basis of the recognition of Ku70 and Ku80 proteins by Impα, we solved the crystal structures of the complexes of Impα with the peptides corresponding to the Ku70 and Ku80 NLSs. Our structural studies confirm the binding of the Ku80 NLS as a classical monopartite NLS but reveal an unexpected binding mode for Ku70 NLS with only one basic cluster bound to the receptor. Both Ku70 and Ku80 therefore contain monopartite NLSs, and sequences outside the basic cluster make favorable interactions with Impα, suggesting that this may be a general feature in monopartite NLSs. We show that the Ku70 NLS has a higher affinity for Impα than the Ku80 NLS, consistent with more extensive interactions in its N-terminal region. The prospect of nuclear import of Ku70 and Ku80 independently of each other provides a powerful regulatory mechanism for the function of the Ku70/Ku80 heterodimer and independent functions of the two proteins.

Asunto(s)

Antígenos Nucleares/metabolismo , Núcleo Celular/metabolismo , Proteínas de Unión al ADN/metabolismo , alfa Carioferinas/metabolismo , Secuencia de Aminoácidos , Animales , Antígenos Nucleares/química , Transporte Biológico , Proteínas de Unión al ADN/química , Humanos , Autoantígeno Ku , Modelos Moleculares , Datos de Secuencia Molecular , alfa Carioferinas/química

RESUMEN

OBJECTIVE: There are subsets of patients with ischemic cardiomyopathy for whom the optimal treatment strategies are not clear. The objective of this study was to delineate the relationship between clinical outcomes and surgical procedure in patients who were treated either with a coronary artery bypass graft or with a coronary artery bypass graft and additional ventricular restoration. METHODS: The study population comprised 137 consecutive patients with anterior myocardial infarction. All patients had an ejection fraction <50% and left ventricle end-systolic volume index >80 ml/m(2). The patients were divided into a viable and a non-viable group according to anterior myocardium viability, which was determined by a thallium-201 test. The viable group underwent a revascularization and was randomized into two groups for additional ventricular reconstruction. Group 1a comprised 35 patients with viable anterior wall who underwent surgical revascularization. Group 1b comprised 39 patients with viable anterior wall who underwent surgical revascularization and ventricular restoration. Group 2 comprised 69 patients with non-viable anterior wall who underwent revascularization and ventricular reconstruction. The preoperative and postoperative ejection fractions, end-systolic volume, mitral regurgitation, mortality, and heart failure symptoms were compared among groups. RESULTS: Complete 2-year follow-up was achieved in 127 (92.7%) patients. Ejection fraction improved in all groups compared with preoperative values and it was greater in group 1b than in group 1a (p<0.001) at 2 years. There were no postoperative deaths in group 1a, one in group 1b, and two in group 2. After 2 years, group 1b was significantly smaller than group 1a (p<0.01) in relation to mitral regurgitation of grades 1 to 2+. End-systolic volume was significantly smaller in group 1b than in group 1a (p<0.001), it was smaller in group 1a than in group 2 (p<0.001), and it was smaller in group 1b than in group 2 (p<0.001). Heart failure class (NYHA) was reduced in all groups and events were significantly smaller in patients with end-systolic volume lesser than 120 ml/m(2) (p<0.05). CONCLUSION: We have demonstrated that the short-term and mid-term outcomes of coronary artery surgery alone in patients with a large left ventricle are inferior to coronary artery surgery plus ventricular restoration.

Asunto(s)

Puente de Arteria Coronaria/métodos , Isquemia Miocárdica/patología , Isquemia Miocárdica/cirugía , Anciano , Ecocardiografía , Femenino , Estudios de Seguimiento , Ventrículos Cardíacos/patología , Ventrículos Cardíacos/fisiopatología , Ventrículos Cardíacos/cirugía , Humanos , Masculino , Persona de Mediana Edad , Insuficiencia de la Válvula Mitral/etiología , Infarto del Miocardio/patología , Infarto del Miocardio/fisiopatología , Isquemia Miocárdica/fisiopatología , Complicaciones Posoperatorias , Estudios Prospectivos , Volumen Sistólico/fisiología , Tasa de Supervivencia , Resultado del Tratamiento

RESUMEN

The crystal structure of an acidic phospholipase A(2) isolated from Bothrops jararacussu venom (BthA-I) chemically modified with p-bromophenacyl bromide (BPB) has been determined at 1.85 Angstroms resolution. The catalytic, platelet-aggregation inhibition, anticoagulant and hypotensive activities of BthA-I are abolished by ligand binding. Electron-density maps permitted unambiguous identification of inhibitor covalently bound to His48 in the substrate-binding cleft. The BthA-I-BPB complex contains three structural regions that are modified after inhibitor binding: the Ca(2+)-binding loop, beta-wing and C-terminal regions. Comparison of BthA-I-BPB with two other BPB-inhibited PLA(2) structures suggests that in the absence of Na(+) ions at the Ca(2+)-binding loop, this loop and other regions of the PLA(2)s undergo structural changes. The BthA-I-BPB structure reveals a novel oligomeric conformation. This conformation is more energetically and conformationally stable than the native structure and the abolition of pharmacological activities by the ligand may be related to the oligomeric structural changes. A residue of the ;pancreatic' loop (Lys69), which is usually attributed as providing the anticoagulant effect, is in the dimeric interface of BthA-I-BPB, leading to a new hypothesis regarding the abolition of this activity by BPB.

Asunto(s)

Acetofenonas/química , Fosfolipasas A/antagonistas & inhibidores , Fosfolipasas A/química , Animales , Anticoagulantes/química , Antihipertensivos/química , Sitios de Unión/efectos de los fármacos , Bothrops , Cristalización , Cristalografía por Rayos X , Ligandos , Modelos Moleculares , Fosfolipasas A/farmacología , Inhibidores de Agregación Plaquetaria/química , Conformación Proteica , Estructura Cuaternaria de Proteína/efectos de los fármacos , Estructura Terciaria de Proteína/efectos de los fármacos , Venenos de Víboras/enzimología

RESUMEN

BnSP-7 and BnSP-6, two Lys49-phospholipase A2 isolated from Bothrops neuwiedi pauloensis snake venom, were co-crystallized with alpha-tocopherol and X-ray diffraction data were collected for both complexes (2.2 and 2.6 A). A new "alternative" quaternary conformation for these two complexes compared with all other dimeric Lys49-PLA2 has been observed.

Asunto(s)

Cristalización/métodos , Fosfolipasas A/química , Difracción de Rayos X/métodos , alfa-Tocoferol/química , Animales , Sitios de Unión , Bothrops/clasificación , Venenos de Crotálidos/química , Cisteína/química , Cisteína/metabolismo , Fosfolipasas A2 Grupo II , Fosfolipasas A2 Grupo IV , Lisina/química , Lisina/metabolismo , Fosfolipasas A2 , Proteínas de Reptiles , Venenos de Serpiente

RESUMEN

An acidic phospholipase A(2) (PLA(2)) isolated from Bothrops jararacussu snake venom was crystallized with two inhibitors: alpha-tocopherol (vitamin E) and p-bromophenacyl bromide (BPB). The crystals diffracted at 1.45- and 1.85-A resolution, respectively, for the complexes with alpha-tocopherol and p-bromophenacyl bromide. The crystals are not isomorphous with those of the native protein, suggesting the inhibitors binding was successful and changes in the quaternary structure may have occurred.

Asunto(s)

Acetofenonas/metabolismo , Bothrops , Venenos de Crotálidos/enzimología , Fosfolipasas A/química , Fosfolipasas A/metabolismo , Vitamina E/metabolismo , Animales , Antioxidantes/metabolismo , Sitios de Unión , Venenos de Crotálidos/química , Cristalización , Inhibidores Enzimáticos/metabolismo , Difracción de Rayos X

RESUMEN

As plaquetas podem estar envolvidas na fisiopatologia da asma liberando mediadores inflamatórios e/ou espasmogênicos, bem como interagindo com outras células inflamatórias. O objetivo deste trabalho foi investigar a agregaçao plaquetária de crianças com asma crônica grave. Foram selecionadas 16 crianças entre 7 e 15 anos de idade, tratadas com brocodilatadores inalatórios e beclometasona (no máximo 750 mug/dia). Doadores de sangue foram selecionados como controles-sadios. Plasma em plaquetas (PRP) e plasma pobre em plaquetas (PPP) foram preparados de acordo com procedimento padrao. A agregaçao de plaquetas foi estudada em agregômetro Payton de dois canais, calibrados com PRP (0 por cento) e PPP (100 por cento) segundo a transmitância de luz. Os estímulos utilizados para induzir a agregaçao plaquetária foram adrenalina (1-30 muM) ou adenosina bifosfato (ADP,1-30 muM). Os resultados foram expressos como percentagem da transmitância máxima. As medianas da agregaçao plaquetária dos pacientes foram respectivamente (adrenalina/ADP 1,3,10 e 30 muM): 0/0, 30/48/41/52, 45/58. As medianas da agregaçao plaquetária dos controles-sadios foram respectivamente (adrenalina/ADP 1,3,10 e 30 muM): 0/0, 12/44, 29/65, 54/74. A análise estatística demontra que a agregaçao de plaquetas dos pacientes foi significativamente menor que a de controles-sadios quando o estímulo foi ADP nas concentraçoes 10 e 30 muM. Em contrapartida, frente aos estímulos adrenalina (1-30 muM) ou mesmo ADP em concentraçoes menores (1-3 muM), a agregaçao de plaquetas dos pacientes foi similar a de controles-sadios. Concluímos que plaquetas de crianças com asma crônica grave sao hiporresponsivas quando o estímulo é ADP em altas concentraçoes. Os mecanismos bioquímicos e molecular envolvidos nesse fenômeno encontram-se sob investigaçao.

Asunto(s)

Humanos , Masculino , Femenino , Niño , Adolescente , Agregación Plaquetaria/fisiología , Asma/fisiopatología , Adenosina Difosfato/efectos adversos , Asma/sangre , Enfermedad Crónica , Epinefrina/efectos adversos