RESUMEN
OBJECTIVE: To evaluate the need for postnatal neurosurgical intervention after fetoscopic patch coverage of spina bifida aperta (SBA). METHODS: This was a retrospective analysis of a cohort of 71 fetuses which underwent minimally invasive fetoscopic patch coverage of SBA between 21 + 0 and 29 + 1 weeks of gestation. Postnatal neurosurgical procedures were classified into two types: re-coverage of the SBA within the first 3 months following birth, and shunt placement as treatment of associated hydrocephalus within the first year. RESULTS: Location of the SBA was lumbosacral in 59 cases, lumbar in seven, thoracic in three and sacral in two. In total, 20/71 (28%) patients underwent early postnatal neurosurgical intervention by means of re-coverage of the SBA. This was performed because of cerebrospinal fluid leakage in seven (35%), adhesions with functional deterioration in three (15%), incomplete coverage in five (25%) and skin defect in five (25%) cases. Ventriculoperitoneal shunt placement within 1 year was required in 32 (45%) cases and was preceded by ventriculostomy in two. Three (4%) infants needed Chiari decompression surgery in the first 12 months following birth, because of syringomyelia or gait disturbance. CONCLUSIONS: Fetoscopic patch coverage of SBA may require postnatal re-coverage in some cases. In most cases, conservative wound treatment shows good results, without requiring neurosurgical intervention. The low 1-year-shunt rate is comparable to data of the Management of Myelomeningocele Study and lower compared with published data of patients with postnatal only coverage of SBA.
Asunto(s)
Fetoscopía/efectos adversos , Feto/cirugía , Procedimientos Neuroquirúrgicos/métodos , Espina Bífida Quística/cirugía , Femenino , Fetoscopía/métodos , Edad Gestacional , Humanos , Hidrocefalia/etiología , Hidrocefalia/cirugía , Lactante , Recién Nacido , Región Lumbosacra/embriología , Región Lumbosacra/cirugía , Atención Posnatal/métodos , Embarazo , Reoperación/métodos , Estudios Retrospectivos , Espina Bífida Quística/complicaciones , Espina Bífida Quística/embriología , Derivación VentriculoperitonealRESUMEN
Sphingosine1phosphate (S1P) plays a key role in cell survival, growth, migration, and in angiogenesis. In glioma, it triggers the activity of the S1Preceptor 1 and of the sphingosine kinase 1; thus influencing the survival rate of patients. The aim of the present study was to investigate the antiproliferative effect of the S1P analogue FTY720 (fingolimod) in glioblastoma (GBM) cells. A172, G28, and U87 cells were incubated with micromolar concentrations of FTY720 or temozolomide (TMZ) for 24 to 72 h. Proliferation and half maximal inhibitory concentration (IC50) were determined by using the xCELLigence system. FACS analysis was performed to check the cell cycle distribution of the cells after a 72h incubation with FTY720. This was then compared to TMZincubated and to untreated cells. Gene expression was detected by RTqPCR in A172, G28, U87 and three primary GBMderived cell lines. FTY720 was able to reduce the number of viable cells. The IC50 value was 4.6 µM in A172 cells, 17.3 µM in G28 cells, and 25.2 µM in U87 cells. FTY720 caused a significant arrest of the cell cycle in all cells and stabilized or overexpressed the level of AKT1, MAPK1, PKCE, RAC1, and ROCK1 transcripts. The TP53 transcript level remained stable or was downregulated after treatment with FTY720. FTY720 may be a promising target drug for the treatment of GBM, as it has a strong antiproliferative effect on GBM cells.
Asunto(s)
Neoplasias Encefálicas/tratamiento farmacológico , Clorhidrato de Fingolimod/farmacología , Glioblastoma/tratamiento farmacológico , Moduladores de los Receptores de fosfatos y esfingosina 1/farmacología , Quinasas Asociadas a rho/metabolismo , Antineoplásicos Alquilantes/farmacología , Apoptosis/efectos de los fármacos , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Glioblastoma/metabolismo , Glioblastoma/patología , Humanos , Transducción de Señal , Temozolomida/farmacologíaRESUMEN
Platelets provide for primary hemostasis by forming a hemostatic plug at sites of vascular damage. They also provide a surface for the assembly of the coagulation protein complexes that generate thrombin, serve as a nidus for fibrin clots, and secrete factors involved in wound repair. Normal platelet function can be divided into four phases: adhesion, aggregation, secretion, and expression of procoagulant activity. Platelet adhesion initiates plug formation as platelets adhere to the connective tissue at the edges of a wound within seconds after vascular damage. When damage occurs in regions of slow blood flow, platelets adhere to subendothelial collagen, fibronectin, and laminin. However, when damage occurs in regions of rapid flow, platelet adhesion requires the presence of subendothelial von Willebrand factor (vWf) and a specific platelet receptor, the glycoprotein Ib/IX (GPIb/IX) complex. Following initial adhesion, platelets aggregate to complete the formation of a hemostatic plug. Platelet aggregation requires active platelet metabolism, platelet stimulation by agonists such as ADP, thrombin, collagen, or epinephrine; the presence of calcium or magnesium ions and specific plasma proteins such as fibrinogen or vWf; and a platelet receptor, the glycoprotein IIb/IIIa (GPIIb/IIIa) complex. Thus, platelet stimulation results in the generation of intracellular second messengers that transmit the stimulus back to the platelet surface, exposing protein binding sites on GPIIb/IIIa. Fibrinogen (or vWf) then binds to GPIIb/IIIa and crosslinks adjacent platelets to produce platelet aggregates. Platelet stimulation also results in platelet secretion and the elaboration of platelet procoagulant activity. During secretion, substances are released to propagate the aggregation response and to promote wound healing; the expression of procoagulant activity localizes thrombin generation to the site of vascular damage. Disorders of platelet function can be divided into those of congenital and those of acquired origin. Although congenital disorders are uncommon, acquired disorders are encountered frequently in clinical practice. Congenital absence of GPIb/IX and GPIIb/IIIa results in the Bernard-Soulier syndrome (BSS) and Glanzmann thrombasthenia (GT), respectively. Each is an autosomal recessive bleeding disorder in which absence of a protein complex renders the affected platelets incapable of undergoing either vWf-mediated adhesion (BSS) or fibrinogen-mediated aggregation (GT).(ABSTRACT TRUNCATED AT 400 WORDS)
Asunto(s)
Trastornos de las Plaquetas Sanguíneas , Plaquetas/fisiología , Corticoesteroides/uso terapéutico , Antifibrinolíticos/uso terapéutico , Coagulación Sanguínea/fisiología , Transfusión de Componentes Sanguíneos , Trastornos de las Plaquetas Sanguíneas/clasificación , Trastornos de las Plaquetas Sanguíneas/etiología , Trastornos de las Plaquetas Sanguíneas/terapia , Plaquetas/metabolismo , Retracción del Coagulo , Desamino Arginina Vasopresina/uso terapéutico , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos , Estrógenos Conjugados (USP)/uso terapéutico , Humanos , Activación Plaquetaria , Adhesividad Plaquetaria , Agregación Plaquetaria , Transducción de SeñalRESUMEN
The present systematic review examined the effectiveness of bilevel noninvasive positive pressure ventilation (NIPPV) in the management of chronic respiratory failure (CRF) due to severe stable chronic obstructive pulmonary disease (COPD). Randomised controlled trials (RCTs) and non-RCTs (crossover design) of adults with severe stable COPD and CRF receiving bilevel NIPPV via nasal, oronasal or total face mask were identified from electronic databases and manual screening of journals and reference lists. Respiratory function (gas exchange, lung function, ventilatory/breathing pattern, respiratory muscle function and work of breathing) and health-related outcomes (dyspnoea, functional status, exercise tolerance, health-related quality of life (HRQOL), morbidity and mortality) were assessed. In total, 15 studies met the inclusion criteria: six RCTs and nine non-RCTs. RCTs did not find improved gas exchange with bilevel NIPPV, while non-RCTs did. Lung hyperinflation and diaphragmatic work of breathing were reduced in a nonrandomised subset. HRQOL and dyspnoea, the least studied outcomes, showed improvement with bilevel NIPPV. In a subset of individuals on maximal medical treatment regimes for severe stable chronic obstructive pulmonary disease, bilevel noninvasive positive pressure ventilation may have an adjunctive role in the management of chronic respiratory failure through attenuation of compromised respiratory function and improvement in health-related outcomes.
Asunto(s)
Respiración con Presión Positiva , Enfermedad Pulmonar Obstructiva Crónica/terapia , Estudios Cruzados , Humanos , Evaluación de Resultado en la Atención de Salud , Enfermedad Pulmonar Obstructiva Crónica/fisiopatología , Intercambio Gaseoso Pulmonar , Calidad de Vida , Ensayos Clínicos Controlados Aleatorios como Asunto , Pruebas de Función Respiratoria , Trabajo RespiratorioRESUMEN
We have isolated from an HEL cell cDNA library an alternatively spliced transcript for the platelet membrane glycoprotein IIb (GPIIb) that resulted from the deletion of the 34 amino acids of exon 28 of the GPIIb gene. Confirming an earlier report, we also detected this transcript in platelet mRNA. To determine the consequences of exon 28 deletion on the expression of the GPIIb/IIIa heterodimer, we expressed cDNA for GPIIb-28 in COS-1 cells, either individually or simultaneously with a cDNA for GPIIIa. When recombinant GPIIb-28 was expressed alone, it did not acquire resistance to the enzyme endo-beta-N-acetylglucosaminidase H, was not cleaved into heavy and light chains, and was not transported to the cell surface. However, when recombinant GPIIb-28 was coexpressed with recombinant GPIIIa, GPIIb/IIIa heterodimers were assembled. Nevertheless, these heterodimers failed to complete posttranslational processing and were degraded intracellularly. Exon 28 contains one site for Asn-linked glycosylation. To determine if loss of this glycosylation site was responsible for the effects of exon 28 deletion, we removed the site from the exon 28 of intact GPIIb by oligonucleotide-mediated mutagenesis. However, absence of the carbohydrate appended to exon 28 did not prevent normal GPIIb/IIIa heterodimer expression. Our studies indicate that absence of the amino acids encoded by GPIIb exon 28 sufficiently perturbs the quaternary configuration of the GPIIb/IIIa heterodimer to impair its subsequent intracellular transport and processing. They also indicate that this alternatively spliced form of GPIIb mRNA, although present in megakaryocytes, is unlikely to make a significant contribution to the GPIIb/IIIa complexes expressed on platelets.
Asunto(s)
Deleción Cromosómica , Exones/genética , Expresión Génica , Glicoproteínas de Membrana Plaquetaria/genética , Secuencia de Bases , Sitios de Unión , Línea Celular , Glicosilación , Humanos , Técnicas de Inmunoadsorción , Sustancias Macromoleculares , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Glicoproteínas de Membrana Plaquetaria/metabolismo , Procesamiento Proteico-Postraduccional , Empalme del ARN , ARN Mensajero/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , TransfecciónRESUMEN
The platelet glycoproteins GPIIb and GPIIIa are integral membrane proteins and form calcium-dependent heterodimers in the endoplasmic reticulum (ER). In the absence of heterodimer formation, GPIIb and GPIIIa are retained in the ER and degraded. To produce soluble forms of these proteins, we truncated each at a site just proximal to its transmembrane anchor and expressed the mutants in COS-1 cells. We found that both truncated GPIIIa (GPIIIatr) and GPIIIatr were secreted by the transfected cells. However, GPIIbtr was retained by the cells and was immunoprecipitated as a doublet with a 115,000 molecular weight protein. Incubation of transfected cells with the calcium ionophore A23187 or the calcium chelator 1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl) ester (BAPTA-AM) failed to induce appreciable GPIIbtr secretion, suggesting that formation of intracellular calcium complexes was not a factor in GPIIbtr retention. Further, immunoblotting of immunoprecipitated GPIIbtr and GPIIIatr revealed that the chaperone binding protein (BiP) was associated with each, arguing that BiP alone was not responsible for GPIIbtr retention. These studies indicate that the intracellular retention of GPIIIa involves sequences located in the transmembrane or cytoplasmic domains of the molecule. GPIIb contains an additional retention signal located in the extracellular portion of the molecule whose effect is abrogated by formation of a GPIIb-IIIa heterodimer. This signal may be involved in the fate of nascent GPIIb monomers and the generation of correctly configured GPIIb-IIIa heterodimers.
Asunto(s)
Mutagénesis Sitio-Dirigida , Glicoproteínas de Membrana Plaquetaria/metabolismo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Calcimicina/farmacología , Línea Celular , Membrana Celular/metabolismo , ADN/genética , Ácido Egtácico/análogos & derivados , Ácido Egtácico/farmacología , Retículo Endoplásmico/metabolismo , Cinética , Sustancias Macromoleculares , Datos de Secuencia Molecular , Plásmidos , Glicoproteínas de Membrana Plaquetaria/genética , Glicoproteínas de Membrana Plaquetaria/aislamiento & purificación , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , TransfecciónRESUMEN
The precursor of platelet membrane glycoprotein IIb (GPIIb) undergoes endoproteolytic cleavage into heavy and light chains post-translation. Endoproteolysis occurs within a 17-amino acid stretch of the precursor that contains 4 arginine residues, 3 in dibasic sequences [Lys-Arg (855-856) and Arg-Arg (858-859)] and a single arginine at 871. To determine the site of GPIIb cleavage and its role in the function of the glycoprotein IIb/IIIa heterodimer, we mutated arginine 856, the di-arginine sequence 858-859, and arginine 871 and coexpressed the mutants with glycoprotein IIIa (GPIIIa) in COS-1 cells. Each GPIIb mutant formed recombinant GPIIb-IIIa heterodimers, but mutants lacking arginine at 856 or 858-859 failed to undergo cleavage. Nevertheless, heterodimers containing the uncleaved GPIIb were expressed on the cell surface. Because endoproteolysis most often occurs after arginines in dibasic sequences, we next expressed GPIIb mutants containing lysine at 856 or aspartic acid at 855 with GPIIIa. Both mutants were cleaved and surface-expressed, indicating that the dibasic sequence at 858-859, but not at 855-856, is required for GPIIb cleavage. Lastly, we tested the function of GPIIb-IIIa containing uncleaved GPIIb by measuring adhesion of transfected cells to immobilized fibrinogen. We found no difference in the adhesion of cells expressing either wild-type or mutant GPIIb, indicating GPIIb-IIIa heterodimers containing uncleaved GPIIb maintain their ability to interact with fibrinogen.
Asunto(s)
Glicoproteínas de Membrana Plaquetaria/metabolismo , Secuencia de Aminoácidos , Animales , Línea Celular , Escherichia coli/genética , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Glicoproteínas de Membrana Plaquetaria/genética , Procesamiento Proteico-Postraduccional , TransfecciónRESUMEN
The integrin alpha IIb beta 3, a calcium-dependent heterodimer, plays a critical role in platelet aggregation. The alpha IIb subunit of the heterodimer contains four highly conserved putative calcium-binding domains in its extracellular portion. During studies of the molecular basis of Glanzmann thrombasthenia in a child of mixed Caucasian background whose platelets expressed little alpha IIb beta 3 on their surface, we found the patient heterozygous for a two amino acid deletion in the fourth alpha IIb calcium-binding domain. When this alpha IIb mutant was expressed in COS-1 cells, we found that the deletion did not interfere with the assembly of alpha IIb beta 3 heterodimers, but altered their conformation such that they were neither recognized by the heterodimer-specific antibody A2A9 nor able to undergo further intracellular processing or transport to the cell surface. These results suggest that the calcium-binding domains in alpha IIb play an important role maintaining the overall conformation of alpha IIb beta 3. To confirm this suggestion, we deleted each of the four 12 amino acid calcium-binding domains in alpha IIb by in vitro mutagenesis and expressed the mutants along with beta 3 in COS-1 cells. Each construct formed a heterodimer with beta 3, but none of the heterodimers interacted with A2A9 or underwent further intracellular processing. These data indicate that the calcium-binding domains in alpha IIb are not involved in alpha IIb beta 3 heterodimer formation, but their presence is required for the intracellular transport of alpha IIb beta 3 to the cell surface.
Asunto(s)
Calcio/metabolismo , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/genética , Conformación Proteica , Eliminación de Secuencia , Trombastenia/genética , Adulto , Alelos , Secuencia de Aminoácidos , Animales , Antígenos de Plaqueta Humana/química , Antígenos de Plaqueta Humana/genética , Antígenos de Plaqueta Humana/inmunología , Secuencia de Bases , Sitios de Unión , Transporte Biológico , Plaquetas/metabolismo , Línea Celular Transformada , Chlorocebus aethiops , Clonación Molecular , Epítopos/química , Epítopos/inmunología , Femenino , Citometría de Flujo , Humanos , Recién Nacido , Masculino , Datos de Secuencia Molecular , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/química , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/inmunología , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , TransfecciónRESUMEN
A phase II trial of Tomudex (raltitrexed, ZD 1694), a new thymidylate synthase inhibitor, was performed in patients with recurrent or metastatic squamous cell carcinoma of the head and neck. This trial demonstrated that Tomudex was well tolerated in this patient population. Nausea and vomiting were minimal, and hematologic toxicities were relatively infrequent. Only one patient was withdrawn from the study due to toxicity (grade 4 diarrhea). One patient exsanguinated from a rent in the carotid artery in an area of tumor involvement, and was categorized as a grade 5 toxicity. Thus 25/27 patients were able to complete at least 2 cycles of treatment. Tomudex demonstrated a 3.7% response rate (95% CI 0.1-19%), with a median survival of 6 months in this highly resistant disease population. Tomudex is not considered active enough as monotherapy for further evaluation in this disease population.