Autophagic-like cell death in neutrophils induced by autoantibodies

Human neutrophils undergo autophagic-like cell death following Sialic acid binding immunoglobulin-like lectin-9 (Siglec-9) ligation and concurrent stimulation with certain, but not all, neutrophil survival cytokines. Caspase inhibition by these cytokines is required, but is not sufficient, to trigger this particular form of cell death. Additional mechanisms may involve reactive oxygen species (ROS), and blocking of ROS or prevention of ROS production prevents autophagic-like neutrophil death. Interestingly, human intravenous immunoglobulin (IVIg) preparations contain natural anti-Siglec-9 autoantibodies, which are able to ligate Siglec-9 on neutrophils and induce autophagic-like cell death in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF) and some other survival cytokines. Here, we discuss the pathophysiological and therapeutic implications of these recent findings.

Viable neutrophils release mitochondrial DNA to form neutrophil extracellular traps

Neutrophil extracellular traps (NETs) represent extracellular structures able to bind and kill microorganisms. It is believed that they are generated by neutrophils undergoing cell death, allowing these dying or dead cells to kill microbes. We show that, following priming with granulocyte/macrophage colony-stimulating factor (GM-CSF) and subsequent short-term toll-like receptor 4 (TLR4) or complement factor 5a (C5a) receptor stimulation, viable neutrophils are able to generate NETs. Strikingly, NETs formed by living cells contain mitochondrial, but no nuclear, DNA. Pharmacological or genetic approaches to block reactive oxygen species (ROS) production suggested that NET formation is ROS dependent. Moreover, neutrophil populations stimulated with GM-CSF and C5a showed increased survival compared with resting neutrophils, which did not generate NETs. In conclusion, mitochondrial DNA release by neutrophils and NET formation do not require neutrophil death and do also not limit the lifespan of these cells.

Induction of Bim limits cytokine-mediated prolonged survival of neutrophils

Under inflammatory conditions, neutrophil apoptosis is delayed due to survival-factor exposure, a mechanism that prevents the resolution of inflammation. One important proinflammatory cytokine involved in the regulation of neutrophil survival/activation is granulocyte-macrophage colony-stimulating factor (GM-CSF). Although GM-CSF mediates antiapoptotic effects in neutrophils, it does not prevent apoptosis, and the survival effect is both time dependent and limited. Here, we identified the proapoptotic Bcl-2 family member Bim as an important lifespan limiting molecule in neutrophils, particularly under conditions of survival factor exposure. Strikingly, GM-CSF induced Bim expression in both human and mouse neutrophils that was blocked by pharmacological inhibition of phosphatidylinositol-3 kinase (PI3K). Increased Bim expression was also seen in human immature bone marrow neutrophils as well as in blood neutrophils from septic shock patients; both cell populations are known to be exposed to GM-CSF under in vivo conditions. The functional role of Bim was investigated using Bim-deficient mouse neutrophils in the presence and absence of the survival cytokines interleukin (IL)-3 and GM-CSF. Lack of Bim expression resulted in a much higher efficacy of the survival cytokines to block neutrophil apoptosis. Taken together, these data demonstrate a functional role for Bim in the regulation of neutrophil apoptosis and suggest that GM-CSF and other neutrophil hematopoietins initiate a proapoptotic counterregulation that involves upregulation of Bim.

STAT3 CONTROLS THE NEUTROPHIL MIGRATORY RESPONSE TO CXCR2 AND ITS LIGAND MIP-2 (CXCL2)

Among the first white blood cells to respond to bacterial and fungal infections, neutrophils are produced in the bone marrow, released into circulating blood, and recruited to inflamed tissue. The cytokine granulocyte colony-stimulating factor (G

Cerebrospinal-fluid cytokine and chemokine profile in patients with pneumococcal and meningococcal meningitis

BACKGROUND Bacterial meningitis is characterized by an intense inflammatory reaction contributing to neuronal damage. The aim of this study was to obtain a comparative analysis of cytokines and chemokines in patients with pneumococcal (PM) and meningococcal meningitis (MM) considering that a clear difference between the immune response induced by these pathogens remains unclear. METHODS The cyto/chemokines, IL-1beta, IL-2, IL-6, TNF-alpha, IFN-gamma, IL-10, IL-1Ra, CXCL8/IL-8, CCL2/MCP-1, CLL3/MIP-1alpha, CCL4/MIP-1gamma and G-CSF, were measured in cerebrospinal fluid (CSF) samples from patients with PM and MM. Additionally, a literature review about the expression of cytokines in CSF samples of patients with MB was made. RESULTS Concerning cytokines levels, only IFN-gamma was significantly higher in patients with Streptococcus pneumoniae compared to those with Neisseria meningitidis, regardless of the time when the lumbar puncture (LP) was made. Furthermore, when samples were compared considering the timing of the LP, higher levels of TNF-alpha (P <0.05) were observed in MM patients whose LP was made within 48 h from the initial symptoms of disease. We also observed that the index of release of cyto/chemokines per cell was significantly higher in PM. From the literature review, it was observed that TNF-alpha, IL-1beta and IL-6 are the best studied cytokines, while reports describing the concentration of the cytokine IL-2, IL-1Ra, G-CSF and CCL4/MIP-1beta in CSF samples of patients with bacterial meningitis were not found. CONCLUSION The data obtained in this study and the previously published data show a similar profile of cytokine expression during PM and MM. Nevertheless, the high levels of IFN-gamma and the ability to release high levels of cytokines with a low number of cells are important factors to be considered in the pathogenesis of PM and thereby should be further investigated. Moreover, differences in the early response induced by the pathogens were observed. However, the differences observed are not sufficient to trigger changes in the current therapy of corticosteroids adopted in both the PM and MM.

Prophylaxis of infectious complications with colony-stimulating factors in adult cancer patients undergoing chemotherapy-evidence-based guidelines from the Infectious Diseases Working Party AGIHO of the German Society for Haematology and Medical Oncology (DGHO).

BACKGROUND Current evidence on myelopoietic growth factors is difficult to overview for the practicing haematologist/oncologist. International guidelines are sometimes conflicting, exclude certain patient groups, or cannot directly be applied to the German health system. This guideline by the Infectious Diseases Working Party (AGIHO) of the German Society of Haematology and Medical Oncology (DGHO) gives evidence-based recommendations for the use of G-CSF, pegylated G-CSF, and biosimilars to prevent infectious complications in cancer patients undergoing chemotherapy, including those with haematological malignancies. METHODS We systematically searched and evaluated current evidence. An expert panel discussed the results and recommendations. We then compared our recommendations to current international guidelines. RESULTS We summarised the data from eligible studies in evidence tables, developed recommendations for different entities and risk groups. CONCLUSION Comprehensive literature search and expert panel consensus confirmed many key recommendations given by international guidelines. Evidence for growth factors during acute myeloid leukaemia induction chemotherapy and pegfilgrastim use in haematological malignancies was rated lower compared with other guidelines.

Role of a ribosomal RNA phosphate oxygen during the EF-G–triggered GTP hydrolysis

Elongation factor-catalyzed GTP hydrolysis is a key reaction during the ribosomal elongation cycle. Recent crystal structures of G proteins, such as elongation factor G (EF-G) bound to the ribosome, as well as many biochemical studies, provide evidence that the direct interaction of translational GTPases (trGTPases) with the sarcin-ricin loop (SRL) of ribosomal RNA (rRNA) is pivotal for hydrolysis. However, the precise mechanism remains elusive and is intensively debated. Based on the close proximity of the phosphate oxygen of A2662 of the SRL to the supposedly catalytic histidine of EF-G (His87), we probed this interaction by an atomic mutagenesis approach. We individually replaced either of the two nonbridging phosphate oxygens at A2662 with a methyl group by the introduction of a methylphosphonate instead of the natural phosphate in fully functional, reconstituted bacterial ribosomes. Our major finding was that only one of the two resulting diastereomers, the SP methylphosphonate, was compatible with efficient GTPase activation on EF-G. The same trend was observed for a second trGTPase, namely EF4 (LepA). In addition, we provide evidence that the negative charge of the A2662 phosphate group must be retained for uncompromised activity in GTP hydrolysis. In summary, our data strongly corroborate that the nonbridging proSP phosphate oxygen at the A2662 of the SRL is critically involved in the activation of GTP hydrolysis. A mechanistic scenario is supported in which positioning of the catalytically active, protonated His87 through electrostatic interactions with the A2662 phosphate group and H-bond networks are key features of ribosome-triggered activation of trGTPases.

Cigarette Smoke Exposure Increases Myeloid Cell Production and Lung Bacterial Clearance in Mice

Pneumonia is a leading cause of hospitalization in patients with chronic obstructive pulmonary disease (COPD). Although most COPD patients are smokers, the effects of cigarette smoke exposure on clearance of lung bacterial pathogens and on immune and inflammatory responses are incompletely defined. Here, clearance of Streptococcus pneumoniae and Pseudomonas aeruginosa and associated immune responses were examined in mice exposed to cigarette smoke or following smoking cessation. Mice exposed to cigarette smoke for 6 weeks or 4 months demonstrated decreased lung bacterial burden compared to air-exposed mice when infected 16-24 hours post-exposure. When infection was performed after smoke cessation, bacterial clearance kinetics of mice previously exposed to smoke reversed to comparable levels as those of control mice suggesting that the observed defects were not dependent on adaptive immunological memory to bacterial determinants found in smoke. Comparing cytokine levels and myeloid cell production prior to infection in mice exposed to cigarette smoke relative to mice never exposed or following smoke cessation revealed that reduced bacterial burden was most strongly associated with higher levels of IL-1β and GM-CSF in the lungs and with increased neutrophil reserve and monocyte turnover in the bone marrow. Using serpinb1a-deficient mice with reduced neutrophil numbers and treatment with G-CSF showed that increased neutrophil numbers contribute only in part to the effect of smoke on infection. Our findings indicate that cigarette smoke induces a temporary and reversible increase in clearance of lung pathogens, which correlates with local inflammation and increased myeloid cell output from the bone marrow.

Neurotoxicity of stem cell mobilization chemotherapy with vinorelbine in myeloma patients after bortezomib treatment.

Vinorelbine chemotherapy with G-CSF stimulation is the standard mobilization regimen in Switzerland for multiple myeloma patients. However, with the increasing use of bortezomib during induction treatment, adding the neurotoxic compound vinorelbine for mobilization may aggravate bortezomib-induced polyneuropathy. In this retrospective single-center study, we aimed to explore vinorelbine mediated neuropathy in 106 consecutive bortezomib pretreated myeloma patients. We confirmed that vinorelbine with G-CSF represents a reliable and effective regimen for mobilization of autologous stem cells. However, the single administration of 35 mg/m(2) vinorelbine added significant neurotoxicity. We found that 24 patients (24%) reported vinorelbine mediated neurotoxicity: Aggravation of bortezomib-induced neuropathy was observed in 17 patients (17%), and vinorelbine mobilization induced first occurrence of polyneuropathy in additional 7 patients (7%). We observed that development of polyneuropathy was not associated with differing survival rates. Finally, affected patients reported polyneuropathy associated disease burden as "very high" in 13% and "high" in 50%. Our data indicate that a single administration of vinorelbine to mobilize autologous stem cells is associated with significant additional polyneuropathy in bortezomib pretreated myeloma patients. The efficacy of vinorelbine mobilization should be balanced against its neurotoxic potential.

Defective antitumor function of monocyte -derived macrophages from epithelial ovarian cancer patients

An abundance of monocytes and macrophages (MO/MA) in the microenvironment of epithelial ovarian cancer (EOC) suggests possible dual roles for these cells. Certain MO/MA subpopulations may inhibit tumor growth by antibody-dependent cell-mediated cytotoxicity (ADCC), phagocytosis, or stimulation of adaptive immunity. In contrast, other MO/MA subpopulations may support tumor growth by immunosuppressive or pro-angiogenic cytokine production. A better understanding of the phenotype and activity of MO/MA in EOC should lead to greater insight into their role in the immunopathobiology of EOC and hence suggest targets for treatment. We have found differences in the proportions of MO/MA subpopulations in the peripheral blood and ascites of EOC patients compared to normal donors, and differences in MO/MA surface phenotype in the associated tumor environment compared to the systemic circulation. We also demonstrate that, following their activation in vitro, monocyte-derived macrophages (MDM) from the peripheral blood and ascites of EOC patients exhibit antitumor effector activities that are different from the behavior of normal donor cells. The phenotypic characteristics and antitumor activity of CD14+ MO/MA and an isolated subpopulation of CD14brightCD16 −HLA-DR+ MO/MA were compared in samples of normal donor peripheral blood and the peripheral blood and ascites from EOC patients. MDM were cultured with macrophage colony-stimulating factor (M-CSF) and activated with lipopolysaccharide (LPS) or a combination of LPS plus recombinant interferon-gamma. We determined that MO/MA from EOC patients had altered morphology and significantly less ADCC and phagocytic activity than did MO/MA from normal donors. ADCC and phagocytosis are mediated by receptors for the Fe portion of IgG (FcγRs), the expression of which were also found to be deficient on EOC MDM from peripheral blood and ascites. Anti-tumor functions not mediated by the FcγRs, such as macrophage mediated cytotoxicity and cytostasis, were not impaired in EOC MDM compared to normal donor MDM. Our findings also showed that MDM from both EOC patients and normal donors produce M-CSF-stimulated cytokines, including interleukin-8, tumor necrosis factor alpha, and interleukin-6, which have the potential to support ovarian tumor growth and metastasis. These findings may be relevant to the pathogenesis of EOC and to the development of future bioimmunotherapeutic strategies. ^

Impacto clínico de la pancitopenia en pacientes hospitalizados

Objetivo: Determinar etiología, manifestaciones clínicas, morbimortalidad y recursos diagnósticos y terapéuticos utilizados en pacientes internados con pancitopenia. Material y métodos: Estudio protocolizado, descriptivo y observacional de 14 meses. Criterios de inclusión: pacientes internados con pancitopenia definida por hemoglobina (Hb) <12 g/dL; plaquetas <150.000/ mL y leucocitos <3.800/mL. Los datos fueron analizados con Epi Info 6.04. Resultados: Se diagnosticaron 54 casos de pancitopenia. Prevalencia: 22/1.000 egresos. Edad media: 48,72 años (DS±20,64); 29,63% fueron > 65 años y 53,70% hombres. Permanencia media: 17,13 días (DS±13,22) vs 7,25 días (DS±5,4) del Servicio (p<0.0001). Charlson medio: 7,16 (DS±2,96) y APACHEII medio: 12 (DS±5,04). El 83,33% (45/54, IC95%70,71-92,08) de las pancitopenias fueron secundarias a compromiso medular, 22 casos (40,74%; IC95%27,57-54,97) postquimioterapia (15 en neoplasias oncohematológicas y 7 en sólidas), 11 (20,37%; IC95%10,63-33,53) por mieloptisis y 4 casos (7,41%; IC95%2,06-17,89) por megaloblastosis. El 16,66% (9/54; IC95%7,92- 29,29) fue secundaria a hiperesplenismo y el 16,66% asociadas a infecciones (3 casos por SIDA). Se realizó estudio de médula ósea en 19 casos (35,18%). El 96,29% (IC95%87,25-99,55) presentó comórbidas. El síndrome de respuesta inflamatoria sistémica (85,19%), síndrome anémico (77,8%) y púrpura (50%) fueron las manifestaciones clínicas más frecuentes. Presentó sepsis el 81,48% (IC95%68,57-90,75) y el 29,63% (IC95%17,98-46,31) hemorragias. El 81,48% tuvo infecciones; el 50% de origen nosocomial y el 65,91% clínicamente documentadas. El 34,09% (IC95%20,49-49,92) tuvo aislamiento microbiológico, con hemocultivos positivos en 29,55%. El 51,85% (IC95%37,84-65,66) desarrolló neutropenia febril (75% postquimioterapia). El 64,81% recibió hemoderivados y factores estimulantes de colonias (G-CSF) el 46,34% (IC95%32,62-60,39). La mortalidad fue mayor a la media global del Servicio (16,66 vs 8,65%)(p=0.07).- Conclusiones: Las pancitopenias en pacientes hospitalizados se caracterizaron por ser secundarias a compromiso medular, hiperesplenismo e infecciones, asociarse a permanencia prolongada, altos índices de comorbilidad, complicaciones infecciosas, y mayor mortalidad que la media global del Servicio.

Pliocene and Pleistocene eastern equatorial Pacific Mg/Ca subsurface temperature

During the early Pliocene warm period (~4.6-4.2 Ma) in the Eastern Equatorial Pacific upwelling region, sea surface temperatures were warm in comparison to modern conditions. Warm upwelling regions have global effects on the heat budget and atmospheric circulation, and are argued to have contributed to Pliocene warmth. Though warm upwelling regions could be explained by weak winds and/or a deep thermocline, the temporal and spatial evolution of the equatorial thermocline is poorly understood. Here we reconstruct temporal and spatial changes in subsurface temperature to monitor thermocline depth and show the thermocline was deeper during the early Pliocene warm period than it is today. We measured subsurface temperature records from Eastern Equatorial Pacific ODP transect Sites 848, 849, and 853 using Mg/Ca records from Globorotalia tumida, which has a depth habitat of ~50-100 m. In the early Pliocene, subsurface temperatures were ~4-5°C warmer than modern temperatures, indicating the thermocline was relatively deep. Subsurface temperatures steeply cooled ~2-3°C from 4.8 to 4.0 Ma and continued to cool an additional 2-3°C from 4.0 Ma to present. Compared to records from other regions, the data suggests the pronounced subsurface cooling between 4.8 and 4.0 Ma was a regional signal related to restriction of the Isthmus of Panama, while continued cooling from 4.0 Ma to present was likely related to global processes that changed global thermocline structure. Additionally, the spatial evolution of the equatorial thermocline along a N-S transect across ODP Sites 853, 849 and 848 suggests an intensification of the southeast trades from the Pliocene to present. Large-scale atmospheric and oceanographic circulation processes link high and low latitude climate through their influence on equatorial thermocline source water regions and consequently the equatorial thermocline. Through these low latitude/high latitude linkages, changes in the equatorial thermocline and thermocline source water played an important role in the transition from the warm Pliocene to the cold Pleistocene.

Contribution to active multi-beam reconfigurable antennas for L and S bands

La presente tesis doctoral con título "Contribution to Active Multi-Beam Reconfigurable Antennas for L and S Bands" ha sido desarrollada por el investigador ingeniero de telecomunicación estudiante de doctorado Javier García-Gasco Trujillo en el Grupo de Radiación del Departamento de Señales, Sistemas y Radiocomunicaciones de la ETSI de Telecomunicación de la Universidad Politécnica de Madrid bajo la dirección de los doctores Manuel Sierra Pérez y José Manuel Fernández González. Durante décadas, el desarrollo de antenas de apuntamiento electrónico ha estado limitado al área militar. Su alto coste y su gran complejidad eran los mayores obstáculos que frenaban la introducción de esta tecnología en aplicaciones comerciales de gran escala. La reciente aparición de componentes de estado sólido prácticos, fiables, y de bajo coste ha roto la barrera del coste y ha reducido la complejidad, haciendo que las antenas reconfigurables de apuntamiento electrónico sean una opción viable en un futuro cercano. De esta manera, las antenas phased array podrían llegar a ser la joya de la corona que permitan alcanzar los futuros retos presentes en los sistemas de comunicaciones tanto civiles como militares. Así pues, ahora es el momento de investigar en el desarrollo de antenas de apuntamiento electrónico de bajo coste, donde los nuevos componentes de estado sólido comerciales forman el núcleo duro de la arquitectura. De esta forma, el estudio e implementación de estos arrays de antenas activas de apuntamiento electrónico capaces de controlar la fase y amplitud de las distintas señales implicadas es uno de los grandes retos de nuestro tiempo. Esta tesis se enfrenta a este desafío, proponiendo novedosas redes de apuntamiento electrónico e innovadores módulos de transmisión/recepción (T/R) utilizando componentes de estado sólido de bajo coste, que podrán integrar asequibles antenas activas reconfigurables multihaz en bandas L y S. En la primera parte de la tesis se realiza una descripción del estado del arte de las antenas phased array, incluyendo su base teórica y sus ventajas competitivas. Debido a que las contribuciones obtenidas en la presente tesis han sido realizadas dentro de distintos proyectos de investigación, donde se han manejada antenas de simple/doble polarización circular y simple/doble banda de trabajo, se describen detenidamente los dos proyectos más relevantes de la investigación: el radar de basura espacial de la Agencia Espacial Europea (ESA), Space Situational Awareness (SSA); y la estación base de seguimiento y control de satélites de órbita baja, GEOdesic Dome Array (GEODA). Sin lugar a dudas, los dispositivos desfasadores son uno de los componentes clave en el diseño de antenas phased arrays. Recientemente se ha observado una gran variación en el precio final de estos dispositivos, llegando en ocasiones a límites inasequibles. Así pues, se han propuesto distintas técnicas de conformación de haz alternativas a la utilización de componentes desfasadores comerciales: el desfasador de líneas conmutadas, la red de haz conmutado, y una novedosa red desfasadora divisora/combinadora de potencia. Para mostrar un uso práctico de las mismas, se ha propuesto el uso de las tres alternativas para el caso práctico del subarray de cinco elementos de la celda GEODA-SARAS. Tras dicho estudio se obtiene que la novedosa red desfasadora divisora/combinadora de potencia propuesta es la que mejor relación comportamiento/coste presenta. Para verificar su correcto funcionamiento se construye y mide los dos bloques principales de los que está compuesta la red total, comprobando que en efecto la red responde según lo esperado. La estructura más simple que permite realizar un barrido plano es el array triangular de tres elementos. Se ha realizado el diseño de una nueva red multihaz que es capaz de proporcionar tres haces ortogonales en un ángulo de elevación _0 y un haz adicional en la dirección broadside utilizando el mencionado array triangular de tres elementos como antena. En primer lugar se realizar una breve introducción al estado del arte de las redes clásicas multihaz. Así mismo se comentan innovadores diseños de redes multihaz sin pérdidas. El estudio da paso a las redes disipativas, de tal forma que se analiza su base matemática y se muestran distintas aplicaciones en arrays triangulares de tres elementos. Finalmente, la novedosa red básica propuesta se presenta, mostrando simulaciones y medidas de la misma para el caso prácticoo de GEODA. También se ha diseñado, construido y medido una red compuesta por dos redes básicas complementarias capaz de proporcionar seis haces cuasi-ortogonales en una dirección _0 con dos haces superpuestos en broadside. La red propuesta queda totalmente validada con la fabricación y medida de estos con prototipos. Las cadenas de RF de los módulos T/R de la nueva antena GEODA-SARAS no son algo trivial. Con el fin de mostrar el desarrollo de una cadena compleja con una gran densidad de componentes de estado sólido, se presenta una descripción detallada de los distintos componentes que integran las cadenas de RF tanto en transmisión como en recepción de la nueva antena GEODA-SARAS. Tras presentar las especificaciones de la antena GEODA-SARA y su diagrama de bloques esquemático se describen los dos bloques principales de las cadenas de RF: la celda de cinco elementos, y el módulo de conversión de panel. De la misma manera también se presentará el módulo de calibración integrado dentro de los dos bloques principales. Para comprobar que el funcionamiento esperado de la placa es el adecuado, se realizará un análisis que tratará entre otros datos: la potencia máxima en la entrada del transmisor (comprobando la saturación de la cadena), señal de recepción mínima y máxima (verificando el rango de sensibilidad requerido), y el factor G/T (cumpliendo la especificación necesaria). Así mismo se mostrará un breve estudio del efecto de la cuantificación de la fase en el conformado de haz de RF. Los estudios muestran que la composición de las cadenas de RF permite el cumplimiento de las especificaciones necesarias. Finalmente la tesis muestra las conclusiones globales del trabajo realizado y las líneas futuras a seguir para continuar con esta línea de investigación. ABSTRACT This PhD thesis named "Contribution to Active Multi-Beam Reconfigurable Antennas for L and S Bands", has been written by the Electrical Engineer MSc. researcher Javier García-Gasco Trujillo in the Grupo de Radiación of the Departamento de Señales, Sistemas y Radiocomunicaciones from the ETSI de Telecomunicación of the Universidad Politécnica de Madrid. For decades, the implementation of electronically steerable phased array antennas was confined to the military area. Their high cost and complexity were the major obstacles to introduce this technology in large scale commercial applications. The recent emergence of new practical, low-cost, and highly reliable solid state devices; breaks the barrier of cost and reduces the complexity, making active phased arrays a viable future option. Thus, phased array antennas could be the crown jewel that allow to meet the future challenges in military and civilian communication systems. Now is time to deploy low-cost phased array antennas, where newly commercial components form the core of the architecture. Therefore, the study and implementation of these novel low-cost and highly efficient solid state phased array blocks capable of controlling signal phase/amplitude accurately is one of the great challenges of our time. This thesis faces this challenge, proposing innovative electronic beam steering networks and transmitter/ receiver (T/R) modules using affordable solid state components, which could integrate fair reconfigurable phased array antennas working in L and S bands. In the first part of the thesis, a description of the state of art of phased array antennas, including their fundamentals and their competitive advantages, is presented. Since thesis contributions have been carried out for different research projects, where antennas with single/double circular polarization and single/double working frequency bands have been examined, frameworks of the two more important projects are detailed: the Space Situational Awareness (SSA) programme from the European Space Agency (ESA), and the GEOdesic Dome Array (GEODA) project from ISDEFE-INSA and the ESA. Undoubtedly, phase shifter devices are one of the key components of phased array antennas. Recent years have witnessed wide fluctuations in commercial phase shifter prices, which sometimes led to unaffordable limit. Several RF steering technique alternatives to the commercial phase shifters are proposed, summarized, and compared: the switched line phase shifter, the switched-beam network, and the novel phase shifter power splitter/combiner network. In order to show a practical use of the three different techniques, the five element GEODA-SARAS subarray is proposed as a real case of study. Finally, a practical study of a newly phase shifter power splitter/combiner network for a subarray of five radiating elements with triangular distribution is shown. Measurements of the two different phase shifter power splitter/combiner prototypes integrating the whole network are also depicted, demonstrating their proper performance. A triangular cell of three radiating elements is the simplest way to obtain a planar scanner. A new multibeam network configuration that provides three orthogonal beams in a desired _0 elevation angle and an extra one in the broadside steering direction for a triangular array of three radiating elements is introduced. Firstly, a short introduction to the state of art of classical multi-beam networks is presented. Lossless network analysis, including original lossless network designs, are also commented. General dissipative network theory as well as applications for array antennas of three radiating elements are depicted. The proposed final basic multi-beam network are simulated, built and measured to the GEODA cell practical case. A combined network that provides six orthogonal beams in a desired _0 elevation angle and a double seventh one in the broadside direction by using two complementary proposed basic networks will be shown. Measurements of the whole system will be also depicted, verifying the expected behavior. GEODA-SARAS T/R module RF chains are not a trivial design. A thorough description of all the components compounding GEODA-SARAS T/R module RF chains is presented. After presenting the general specifications of the GEODA-SARAS antenna and its block diagrams; two main blocks of the RF chains, the five element cell and the panel conversion module, are depicted and analyzed. Calibration module integrated within the two main blocks are also depicted. Signal flow throw the system analyzing critical situations such as maximum transmitted power (testing the chain unsaturation), minimum and maximum receiving signal (verifying sensitivity range), maximum receiver interference signals (assuring a proper reception), and G/T factor (fulfilling the technical specification) are evaluated. Phase quantization error effects are also listed. Finally, the manuscript contains the conclusions drawn of the present research and the future work.

Dissociation of cytokine-induced phosphorylation of Bad and activation of PKB/akt: Involvement of MEK upstream of Bad phosphorylation

The phosphatidylinositol 3-kinase (PI3K)-signaling pathway has emerged as an important component of cytokine-mediated survival of hemopoietic cells. Recently, the protein kinase PKB/akt (referred to here as PKB) has been identified as a downstream target of PI3K necessary for survival. PKB has also been implicated in the phosphorylation of Bad, potentially linking the survival effects of cytokines with the Bcl-2 family. We have shown that granulocyte/macrophage colony-stimulating factor (GM-CSF) maintains survival in the absence of PI3K activity, and we now show that when PKB activation is also completely blocked, GM-CSF is still able to stimulate phosphorylation of Bad. Interleukin 3 (IL-3), on the other hand, requires PI3K for survival, and blocking PI3K partially inhibited Bad phosphorylation. IL-4, unique among the cytokines in that it lacks the ability to activate the p21ras–mitogen-activated protein kinase (MAPK) cascade, was found to activate PKB and promote cell survival, but it did not stimulate Bad phosphorylation. Finally, although our data suggest that the MAPK pathway is not required for inhibition of apoptosis, we provide evidence that phosphorylation of Bad may be occurring via a MAPK/ERK kinase (MEK)-dependent pathway. Together, these results demonstrate that although PI3K may contribute to phosphorylation of Bad in some instances, there is at least one other PI3K-independent pathway involved, possibly via activation of MEK. Our data also suggest that although phosphorylation of Bad may be one means by which cytokines can inhibit apoptosis, it may be neither sufficient nor necessary for the survival effect.

Constitutive and impaired signaling of leptin receptors containing the Gln → Pro extracellular domain fatty mutation

Leptin (OB), an adipocyte-secreted circulating hormone, and its receptor (OB-R) are key components of an endocrine loop that regulates mammalian body weight. In this report we have analyzed signal transduction activities of OB-R containing the fatty mutation [OB-R(fa)], a single amino acid substitution at position 269 (Gln → Pro) in the OB-R extracellular domain that results in the obese phenotype of the fatty rat. We find that this mutant receptor exhibits both ligand-independent transcriptional activation via interleukin 6 and hematopoietin receptor response elements and ligand-independent activation of signal transducer and activator of transcription (STAT) proteins 1 and 3. However, OB-R(fa) is unable to constitutively activate STAT5B and is highly impaired for ligand induced activation of STAT5B compared with OB-R(wt). Introduction of the fatty mutation into a OB-R/G-CSF-R chimera generates a receptor with constitutive character that is similar but distinct from that of OB-R(fa). Constitutive mutant OB-R(fa) receptor signaling is repressed by coexpression of OB-R(wt). The implications of an extracellular domain amino acid substitution generating a cytokine receptor with a partially constitutive phenotype are discussed both in terms of the mechanism of OB-R triggering and the biology of the fatty rat.