Secretory immunoglobulin A: well beyond immune exclusion at mucosal surfaces.

At mucosal surfaces, secretory IgA (SIgA) antibodies serve as the first line of defense against microorganisms through a mechanism called immune exclusion that prevents interaction of neutralized antigens with the epithelium. In addition, SIgA plays a role in the immune balance of the epithelial barrier through selective adhesion to M cells in intestinal Peyer's patches. This mediates the transepithelial retro-transport of the antibody and associated antigens from the intestinal lumen to underlying gut-associated organized lymphoid tissue. In Peyer's patches, SIgA-based immune complexes are internalized by underlying antigen-presenting cells, leaving the antigen with masked epitopes, a form that limits the risk of overwhelming the local immune protection system with danger signals. This translates into the onset of mucosal and systemic responses associated with production of anti-inflammatory cytokines and limited activation of antigen-presenting cells. In the gastrointestinal tract, SIgA exhibits thus properties of a neutralizing agent (immune exclusion) and of an immunopotentiator inducing effector immune responses in a noninflammatory context favorable to preserve local homeostasis.

Digital beamforming and moving target indication for future spaceborne synthetic aperture radar

During the last decade the interest on space-borne Synthetic Aperture Radars (SAR) for remote sensing applications has grown as testified by the number of recent and forthcoming missions as TerraSAR-X, RADARSAT-2, COSMO-kyMed, TanDEM-X and the Spanish SEOSAR/PAZ. In this sense, this thesis proposes to study and analyze the performance of the state-of-the-Art space-borne SAR systems, with modes able to provide Moving Target Indication capabilities (MTI), i.e. moving object detection and estimation. The research will focus on the MTI processing techniques as well as the architecture and/ or configuration of the SAR instrument, setting the limitations of the current systems with MTI capabilities, and proposing efficient solutions for the future missions. Two European projects, to which the Universitat Politècnica de Catalunya provides support, are an excellent framework for the research activities suggested in this thesis. NEWA project proposes a potential European space-borne radar system with MTI capabilities in order to fulfill the upcoming European security policies. This thesis will critically review the state-of-the-Art MTI processing techniques as well as the readiness and maturity level of the developed capabilities. For each one of the techniques a performance analysis will be carried out based on the available technologies, deriving a roadmap and identifying the different technological gaps. In line with this study a simulator tool will be developed in order to validate and evaluate different MTI techniques in the basis of a flexible space-borne radar configuration. The calibration of a SAR system is mandatory for the accurate formation of the SAR images and turns to be critical in the advanced operation modes as MTI. In this sense, the SEOSAR/PAZ project proposes the study and estimation of the radiometric budget. This thesis will also focus on an exhaustive analysis of the radiometric budget considering the current calibration concepts and their possible limitations. In the framework of this project a key point will be the study of the Dual Receive Antenna (DRA) mode, which provides MTI capabilities to the mission. An additional aspect under study is the applicability of the Digital Beamforming on multichannel and/or multistatic radar platforms, which conform potential solutions for the NEWA project with the aim to fully exploit its capability jointly with MTI techniques.

Biochemical characterization of recombinant human telomerase

Résumé Les télomères sont les structures ADN-protéines des extrémités des chromosomes des eucaryotes. L'ADN télomérique est constitué de courtes séquences répétitives. L'intégrité des télomères est essentielle pour protéger les extrémités des chromosomes contre les systèmes de dégradations et pour les distinguer des cassures de l'ADN double brin. Parce que la machinerie de la réplication de l'ADN n'est pas capable de répliquer l'extrémité des chromosomes, les télomères raccourcissent au fur et à mesure des cycles de réplication. Dès que les télomères atteignent une longueur critique, leur structure protectrice est perdue. Cela induit un signal de dommage de l'ADN et l'arrêt du cycle cellulaire. Pour contrebalancer le raccourcissement des télomères, les cellules qui s'auto régénèrent, dont les cellules de la moelle osseuse, les lymphocytes activés et 80-90% des cellules cancéreuses, expriment la télomérase. C'est une ribonucléoprotéine qui a la capacité de synthétiser des séquences télomériques par transcription inverse d'une courte séquence contenue dans sa propre sous-unité ARN avec laquelle elle est associée. La télomérase humaine est une enzyme processive au niveau de l'addition des nucléotides et aussi des répétitions télomériques. La télomérase de levure et la télomérase humaine sont toutes deux dimériques et il a été montré que la télomérase humaine recombinante contient deux ARN qui coopèrent pour fonctionner ainsi que deux sous-unités catalytiques. Cependant, il n'a pas encore été montré quel est le rôle de la dimérisation dans l'activité de la télomérase. Afin d'élucider ce rôle, nous avons exprimé, reconstitué et purifié la télomérase humaine dimérique recombinante. Et pour étudier l'effet d'ARN mutants sur l'activité de la télomérase, nous avons développé une méthode pour reconstituer et enrichir en hétérodimères de télomérase. Les hétérodimères contiennent une sous-unité ARN sauvage et une sous-unité ARN mutée au niveau de la séquence de la matrice. Sur l'ARN muté nous avons introduit une étiquette aptamer ARN-S1 puis nous avons purifié la télomérase via l'etiquette Si. Nous avons montré que la dimérisation est essentielle pour l'activité de la télomérase. Nos données indiquent que chaque télomérase du dimère allonge leur substrat, l'ADN télomérique, indépendamment l'une de l'autre à chaque cycle d'élongation mais que l'addition itérative de répétitions télomériques nécessite une coopération entre les deux télomérases du dimère. Nous proposons donc un modèle dans lequel les deux télomérases du dimères se lient et allongent deux substrats télomères et que pendant l'élongation processive les deux enzymes subissent un changement de conformation de manière coordonnée, ce changement va permettre le repositionnement des substrats pour d'autres cycles d'additions de répétitions télomériques. Dyskeratosis congenita est une maladie mortelle due majoritairement au disfonctionnement de la moelle osseuse. Dans la forme autosomale de la maladie, l'ARN de la télomérase contient des mutations. En utilisant notre système de reconstitution, nous avons montré que ces ARN mutés, qui ont perdu leur activité enzymatique dans le cas d'un homodimère de mutants, sont dominant négatifs quand ils sont présents dans les hétérodimères sauvage/mutant. Cet effet trans-dominant négatif pourrait contribuer à la progression de la maladie. Abstract Telomeres are protein-DNA structures at the ends of linear eukaryotic chromosomes. The telomeric DNA consists of tandemly repeated sequences. Telomeric integrity is essential to protect chromosomal ends from nucleolytic degradation and to prevent their recognition as DNA double strand breaks. Due to the inability of the conventional DNA replication machinery to replicate terminal DNA stretches, telomeres shorten with continuous rounds of DNA replication. As soon as telomeres reach a critical length, their protective structure is lost and the deprotected telomeres will induce a DNA damage response leading to cell cycle arrest. To counteract telomere shortening, self-renewing cells, including bone marrow cells, activated lymphocytes and 80-90% of cancer cells express the cellular reverse transcriptase telomerase, which has the capacity to synthesize telomeric repeats by reverse transcription of a short template sequence encoded by its stably associated RNA subunit. Human telomerase is a processive enzyme for nucleotide as well as repeat addition. Both yeast and human telomerase are dimeric enzymes and recombinant human telomerase has been shown to contain two functionally cooperating RNAs and most probably also two protein subunits. However, it has remained unclear how dimerization may contribute to telomerase activity. To study the role of dimerization, we expressed, reconstituted and purified recombinant human telomerase. We also developed a new method to reconstitute and enrich for telomerase heterodimers containing wild-type (wt) and mutant telomerase RNA subunits. To this end we introduced an S1-RNA-aptamer tag into telomerase RNA and purified telomerase reconstituted with a mixture of untagged and tagged RNA via the S1-tag. Using this experimental system, we introduced template mutations in the tagged RNA subunit and examined the effect of mutant RNAs on wt telomerase activity in wt/mutant heterodimers. We obtained evidence that dimerization is essential for telomerase activity. Our data indicate that the two subunits elongate telomere substrates independently of each other during single rounds of elongation, but that iterative addition of telomeric repeats requires cooperation between the two subunits. We suggest a model, in which dimeric telomerases bind and elongate two telomere substrates and that the two subunits undergo coordinated conformational changes during processive elongation that enable repositioning the substrates for subsequent rounds of repeat addition. Dyskeratosis congenita is a multisystemic disease with bone marrow failure as the major cause of death. The autosomal form of this disease was found to harbor mutations in the telomerase RNA. Using our reconstitution system, we tested whether mutant dyskeratosis telomerase RNAs behaved in a dominant negative manner. We observed that dyskeratosis telomerase RNA mutants, which lacked enzymatic activity were dominant negative, when present in wt/ mutant heterodimers. The transdominant negative effect of these mutants may contribute to disease progression.

An anonymous reputation mechanism for cloud computing networks using volunteer resources

One of the major problems when using non-dedicated volunteer resources in adistributed network is the high volatility of these hosts since they can go offlineor become unavailable at any time without control. Furthermore, the use ofvolunteer resources implies some security issues due to the fact that they aregenerally anonymous entities which we know nothing about. So, how to trustin someone we do not know?.Over the last years an important number of reputation-based trust solutionshave been designed to evaluate the participants' behavior in a system.However, most of these solutions are addressed to P2P and ad-hoc mobilenetworks that may not fit well with other kinds of distributed systems thatcould take advantage of volunteer resources as recent cloud computinginfrastructures.In this paper we propose a first approach to design an anonymous reputationmechanism for CoDeS [1], a middleware for building fogs where deployingservices using volunteer resources. The participants are reputation clients(RC), a reputation authority (RA) and a certification authority (CA). Users needa valid public key certificate from the CA to register to the RA and obtain thedata needed to participate into the system, as now an opaque identifier thatwe call here pseudonym and an initial reputation value that users provide toother users when interacting together. The mechanism prevents not only themanipulation of the provided reputation values but also any disclosure of theusers' identities to any other users or authorities so the anonymity isguaranteed.

Novel micelle carriers for cyclosporin A topical ocular delivery: in vivo cornea penetration, ocular distribution and efficacy studies.

Cornea transplantation is one of the most performed graft procedures worldwide with an impressive success rate of 90%. However, for "high-risk" patients with particular ocular diseases in addition to the required surgery, the success rate is drastically reduced to 50%. In these cases, cyclosporin A (CsA) is frequently used to prevent the cornea rejection by a systemic treatment with possible systemic side effects for the patients. To overcome these problems, it is a challenge to prepare well-tolerated topical CsA formulations. Normally high amounts of oils or surfactants are needed for the solubilization of the very hydrophobic CsA. Furthermore, it is in general difficult to obtain ocular therapeutic drug levels with topical instillations due to the corneal barriers that efficiently protect the intraocular structures from foreign substances thus also from drugs. The aim of this study was to investigate in vivo the effects of a novel CsA topical aqueous formulation. This formulation was based on nanosized polymeric micelles as drug carriers. An established rat model for the prevention of cornea graft rejection after a keratoplasty procedure was used. After instillation of the novel formulation with fluorescent labeled micelles, confocal analysis of flat-mounted corneas clearly showed that the nanosized carriers were able to penetrate into all corneal layers. The efficacy of a 0.5% CsA micelle formulation was tested and compared to a physiological saline solution and to a systemic administration of CsA. In our studies, the topical CsA treatment was carried out for 14 days, and the three parameters (a) cornea transparency, (b) edema, and (c) neovascularization were evaluated by clinical observation and scoring. Compared to the control group, the treated group showed a significant higher cornea transparency and significant lower edema after 7 and 13 days of the surgery. At the end point of the study, the neovascularization was reduced by 50% in the CsA-micelle treated animals. The success rate of cornea graft transplantation was 73% in treated animals against 25% for the control group. This result was as good as observed for a systemic CsA treatment in the same animal model. This new formulation has the same efficacy like a systemic treatment but without the serious CsA systemic side effects. Ocular drug levels of transplanted and healthy rat eyes were dosed by UPLC/MS and showed a high CsA value in the cornea (11710 ± 7530 ng(CsA)/g(tissue) and 6470 ± 1730 ng(CsA)/g(tissue), respectively). In conclusion, the applied formulation has the capacity to overcome the ocular surface barriers, the micelles formed a drug reservoir in the cornea from, where a sustained release of CsA can take place. This novel formulation for topical application of CsA is clearly an effective and well-tolerated alternative to the systemic treatment for the prevention of corneal graft rejection.

Designing business models of cloud platforms

Cloud computing and its three facets (Software as a Service (SaaS), Platform as a Service (PaaS), and Infrastructure as a Service (IaaS)) are terms that denote new developments in the software industry. In particular, PaaS solutions, also referred to as cloud platforms, are changing the way software is being produced, distributed, consumed, and priced. Software vendors have started considering cloud platforms as a strategic option but are battling to redefine their offerings to embrace PaaS. In contrast to SaaS and IaaS, PaaS allows for value co-creation with partners to develop complementary components and applications. It thus requires multisided business models that bring together two or more distinct customer segments. Understanding how to design PaaS business models to establish a flourishing ecosystem is crucial for software vendors. This doctoral thesis aims to address this issue in three interrelated research parts. First, based on case study research, the thesis provides a deeper understanding of current PaaS business models and their evolution. Second, it analyses and simulates consumers' preferences regarding PaaS business models, using a conjoint approach to find out what determines the choice of cloud platforms. Finally, building on the previous research outcomes, the third part introduces a design theory for the emerging class of PaaS business models, which is grounded on an extensive action design research study with a large European software vendor. Understanding PaaS business models from a market as well as a consumer perspective will, together with the design theory, inform and guide decision makers in their business model innovation plans. It also closes gaps in the research related to PaaS business model design and more generally related to platform business models.

Dual-channel electrospray microchip.

A dual-channel electrospray microchip has been developed for electrospray ionization mass spectrometry (ESI-MS) where aqueous samples are mixed at the Taylor cone with an organic buffer. Due to the fast and effective mixing in the Taylor cone, the aqueous sample can be well ionized with a high ion intensity. The influence of geometric parameters such as the distance between the two microchannels at their junction at the tip of the emitter has been investigated together with chemical parameters such as the organic buffer composition.

Sumak Kawsay, or the concept of living well, and perceptions about natural, urban and rural areas: the case of Cuenca (Ecuador)

This research explores critical environmental concepts from the standpoint of political ecology. It contrasts the Ecuadorian Kichua indigenous concept of Sumak Kawsay (i.e., living well), with discourses about natural, urban and rural areas. Sumak Kawsay was included in Ecuador’s 2008 constitution as an alternative to the neoclassical idea of development. This work reveals interesting insights from a variety of social and political viewpoints from both rural and urban areas. The research is based on the medium sized city of Cuenca, which is located in the southern Ecuadorian Andes. We used an adaptation of the Q Method to investigate stakeholders’ perceptions. Four remarkably well-defined viewpoints were revealed. Although these views contrast strongly with Sumak Kawsay, it is possible to transform this paradigm in the foundation of Cuenca’s public policies and institutional structure.

Decreased well-being after job loss : testing omitted causes

Job loss is widely known to lead to a substantial decrease in workers' subjective well-being. Functionalist theories explain this fact by arguing that the fundamental needs that work fulfills are absent during unemployment. Recent evidence from longitudinal studies however contradicts this approach, showing that workers who find a new job do not fully regain their former level of well-being upon reemployment. Therefore other mechanisms must be at work. We suggest that changes in social or economic domains of workers' lives - triggered by job displacement - lead to the observed changes in well-being. Drawing on a unique data set from a survey of workers displaced by plant closure in Switzerland after the financial crisis of 2008, our analysis confirms the previous result that finding a job after displacement does not completely restore workers' pre-displacement level of well-being. The factors that best explain this outcome are changes in social domains, notably changes in workers' job - related social status and their relationships to friends. This result provides valuable insights about the long lasting scars job displacement leaves on workers' lives.

Decentralization in Croatia: problems and possible solutions

El 2001 Croàcia va posar en marxa la iniciativa destinada a afluixar un alt grau de centralització mitjançant l'ampliació dels mandats de les unitats locals i el canvi de les fonts de finançament de les funcions públiques. No obstant això, els passos inicials en el procés de descentralització no ha estat seguida per altres mesures de descentralització fiscal, i en conseqüència, el seu nivell s'ha mantingut pràcticament sense canvis. El treball es proposa elaborar els principals problemes i obstacles en relació amb l'aplicació efectiva del procés de descentralització i donar tres grups de propostes per a la descentralització: (i) la divisió dels poders obligatoris entre les diferents unitats locals, (ii) el canvi en el finançament i (Iii) modificar la divisió territorial del país.

Study of protein haptenation by amoxicillin through the use of a biotinylated antibiotic.

Allergic reactions towards β-lactam antibiotics pose an important clinical problem. The ability of small molecules, such as a β-lactams, to bind covalently to proteins, in a process known as haptenation, is considered necessary for induction of a specific immunological response. Identification of the proteins modified by β-lactams and elucidation of the relevance of this process in allergic reactions requires sensitive tools. Here we describe the preparation and characterization of a biotinylated amoxicillin analog (AX-B) as a tool for the study of protein haptenation by amoxicillin (AX). AX-B, obtained by the inclusion of a biotin moiety at the lateral chain of AX, showed a chemical reactivity identical to AX. Covalent modification of proteins by AX-B was reduced by excess AX and vice versa, suggesting competition for binding to the same targets. From an immunological point of view, AX and AX-B behaved similarly in RAST inhibition studies with sera of patients with non-selective allergy towards β-lactams, whereas, as expected, competition by AX-B was poorer with sera of AX-selective patients, which recognize AX lateral chain. Use of AX-B followed by biotin detection allowed the observation of human serum albumin (HSA) modification by concentrations 100-fold lower that when using AX followed by immunological detection. Incubation of human serum with AX-B led to the haptenation of all of the previously identified major AX targets. In addition, some new targets could be detected. Interestingly, AX-B allowed the detection of intracellular protein adducts, which showed a cell type-specific pattern. This opens the possibility of following the formation and fate of AX-B adducts in cells. Thus, AX-B may constitute a valuable tool for the identification of AX targets with high sensitivity as well as for the elucidation of the mechanisms involved in allergy towards β-lactams.

Fish oil after abdominal aorta aneurysm surgery

OBJECTIVE: Fish oil (FO) may attenuate the inflammatory response after major surgery such as abdominal aortic aneurysm (AAA) surgery. We aimed at evaluating the clinical impact and safety aspects of a FO containing parenteral nutrition (PN) after AAA surgery. METHODS: Intervention consisted in 4 days of either standard (STD: Lipofundin medium-chain triglyceride (MCT): long-chain triglyceride (LCT)50%-MCT50%) or FO containing PN (FO: Lipoplus: LCT40%-MCT50%-FO10%). Energy target were set at 1.3 times the preoperative resting energy expenditure by indirect calorimetry. Blood sampling on days 0, 2, 3 and 4. Glucose turnover by the (2)H(2)-glucose method. Muscle microdialysis. Clinical data: maximal daily T degrees, intensive care unit (ICU) and hospital stay. RESULTS: Both solutions were clinically well tolerated, without any differences in laboratory safety parameters, inflammatory, metabolic data, or in organ failures. Plasma tocopherol increased similarly; with FO, docosahexaenoic and eicosapentaenoic acid increased significantly by day 4 versus baseline or STD. To increased postoperatively, with a trend to lower values in FO group (P=0.09). After FO, a trend toward shorter ICU stay (1.6+/-0.4 versus 2.3+/-0.4), and hospital stay (9.9+/-2.4 versus 11.3+/-2.7 days: P=0.19) was observed. CONCLUSIONS: Both lipid emulsions were well tolerated. FO-PN enhanced the plasma n-3 polyunsaturated fatty acid content, and was associated with trends to lower body temperature and shorter length of stay.