Esta tesis pertenece a la línea de investigación La expresión del proyecto de arquitectura. Análisis y evolución del curso de doctorado de Comunicación Visual en Arquitectura y Diseño de la UPC-ETSAB. La presente investigación se inicia como una exploración en el límite, -uno de los elementos más importantes en la arquitectura que definen el espacio y concretan la realidad-, en un contexto marcado por un pensamiento sistémico -complejo y heterogéneo- y en una realidad en constante renovación. En este escenario, el límite se reconoce como un elemento difuso en la relación de ámbitos o disciplinas. Con una imagen espacial y cognitiva del límite se reconocen los modos de expresión utilizados por los arquitectos para conocer, formular y explorar sus ideas, es decir: aspectos teóricos, de representación y la obra construida. El contexto comprende el trabajo de los últimos veinte años, coincidiendo con la implantación operativa de las tecnologías digitales en los estudios de arquitectura. Los años noventa son los del inicio de jóvenes prácticas que forman la vanguardia avanzada actual y un punto de inflexión en otras existentes, en las que se observa la superación de un período deconstructivo en la posmodernidad. La imagen de límite puede afrontarse desde distintos enfoques, pero su análisis bajo tres puntos de vista permite abordar este estudio con algunas de las posiciones más representativas por la materia tratada y el momento cultural: lo espacial, por la condición creadora de ámbitos de la arquitectura; lo sistémico, por el carácter del pensamiento actual asentado en procesos y relaciones entre objetos; y lo disciplinar, en respuesta al pensamiento complejo y por el interés investigador de las prácticas contemporáneas que promueven la interdisciplinariedad. Bajo estos ángulos, la estructura en capítulos independientes revela distintos modos de enfrentarse a esta cuestión y permite definir similitudes o divergencias entre ellos. La elección de cinco posiciones arquitectónicas diferentes responde a las más representativas caracterizando distintos sistemas y expresiones del límite. De este modo, en el cuerpo central de la tesis se descubre, en diferentes capítulos: un límite físico que busca la abertura sin barreras del espacio por el despliegue de un orden aleatorio y una envolvente que se desdibuja con el contexto, en una Arquitectura de límites difusos personificada por Toyo Ito; un límite perceptivo que diluye las fronteras físicas en una indeterminación de su existencia por la construcción de un sistema escénico-audiovisual y una práctica ambigua entre diferentes disciplinas en la Exploración de un marco visual de Diller Scofidio + Renfro (DS+R); un límite diverso que acota sin delimitar u orienta sin obstáculos una diversidad funcional a través de la articulación de fragmentos y una arquitectura paradójica en El límite como imagen del espacio por MVRDV. Por otro lado, una existencia entre lo real y lo virtual descubre una arquitectura formal, temporal y dinámica que concreta, a través del movimiento, las características del espacio y un límite fluctuante que se redefine por la acción del hombre en Una dualidad del límite: de lo virtual a lo real representada por Asymptote; y por último, la singularidad de una Práctica al límite de la disciplina y de la realidad en Marcos Novak exhiben un límite líquido, que emerge sin fin y una no-arquitectura simbólica. Desde lo espacial, el examen a través de la envolvente evidencia un abanico de conceptos y apariencias que contrastan con la imagen de frontera más o menos estable durante largos periodos de la historia. El límite expresado como una piel, membrana, contorno, margen, umbral, perfil o caparazón determina una interface o una fachada comunicativa a través de la transparencia, una gradación espacial; una articulación entre distintos territorios, o un cerramiento protector como aislamiento frente al exterior. Desde lo sistémico, la comparación de procesos y estrategias manifiesta modelos diferenciados y continuos que minimizan la distancia entre arquitectura, espacio y cuerpo, o construcción y territorio. Por un lado, en los sistemas volumétricos o superficiales, la propia arquitectura habilita atmósferas heterogéneas y reduce las barreras al movimiento o a la visión. Por otro, los sistemas virtuales erigen un límite inmaterial donde el hombre es su motor impulsor y se constata una disolución de jerarquías espaciales y constructivas. El desarrollo de órdenes, encuadres, sistemas dinámicos o procesos emergentes erigen modelos porosos, laberínticos, fluidos y límites informales que suprimen las fronteras lo definen la ocupación y vinculan el espacio. Desde lo disciplinar, el manejo de forma transversal en esta investigación de técnicas, tecnología y práctica revela nuevos lenguajes, modos de expresión y formalizaciones que determinan una práctica nómada. De este modo, la experiencia interdisciplinar, multidisciplinar o transdisciplinar confirma posiciones ambiguas entre la arquitectura y el arte que implican a los espacios de trabajo y a los lugares de presentación. Las técnicas escénicas, de análisis, animación o simbólicas y las tecnologías audiovisuales, industriales o informáticas destapan una variedad formal y representativa y nuevas oportunidades generativas en la producción que afectan a la interpretación del límite. En este ámbito se verifica una relación directa con las soluciones y los procesos desarrollados. Finalmente en esta tesis se demostrará que el trabajo en el límite, que es lo que hace avanzar a cualquier disciplina, impulsa a la arquitectura de manera esencial. / This research starts as an exploration at the limit, -one of the most important elements in architecture that defines the space and concretizes the reality. , in a context marked by a syslemic thought -complexand heterogeneous- and a realily in constant renewal. At this scenario, the limit is recognized like a diffuse element in the relation of spaces or disciplines. The analysis, under three points of view, and from the beginning of 90 to today- coinciding wilh the operating implantation of the digital technologies in architecture and being the star of actual avant-garde practices and one point of inflection in existing others- allows lackling this study with Ihe most representatives for the matter and the cultural moment treated: the space one, is set for the creative condition of fields of the architecture; the systemic one, for the caracter of a thought established in processes and relations; and the disciplinary one, in response to this thought and for the research work of the contemporary practices that promote the interdisciplinarily. Under these angles, the choice of five architectonic positions allows characterizing several systems and expressions of the limit and defining similarities or divergences among themselves. In this thesis is discovered: a physical limit that looks for the opening without barriers for the display of a fortuitous order and an enveloping that becomes blurred with the context, in an "Arquitectura de limites difusos", byToyo Ito; a perceptive limit that dissolves the physical frontiers in an indetermination of its existence for a scenic-audiovisual system, in the "Exploración de un marco visual", by Diller Scofidio + Renfro; a diverse limit that encloses without delimiting through the joint of functional fragments and the reformutation of models, in "El limite como imagen del espacio", by MVRDV. An existence real-virtual discovers a fluctuating limit and a dynamic temporary system, in "Una dualidad del limite: de lo virtual a lo real", by Asymptote; and a "Práctica al limite de la disciplina y de la realidad" by Marcos Novak exhibits a liquid limit, that emerges endless and a not architecture. From a space view, the exam through an enveloping surface evidences a range of appearances. A skin, membrane, contour, border, doorstep, outline or shell fix an interface or a communicative faryade through transparency; a space gradation, a joint between territories or a protective closing like an isolation to the external. From a systemic view, the strategies manifest models that minimize the distance between architecture, space and body, construclion and territory. The own architecture habilitates heterogeneous atmospheres without barriers or the virtual systems erect immateriallimits whose motor is the man and a dilution hierarchic, of space and constructive are verified. Orders, frameworks, dynamic or emerging processes erect porous, labyrinthical, flowing and informal models thal suppress the frontiers or link the space. From a disciplining view, the use of a cross curricular way of techniques and technology reveals new languages, manners of expression and a nomadic practice. The experience interdisciplinary, multidisciplinary or transdisciplinary explores positions between the architecture and the art. The scenic, analysis or animation techniques and the audiovisual or information technologies emphasize a formal, representative variety and generative opportunities in the production that affect the inlerprelalion of the limit. In Ihis thesis it will be showed thal the work at the limit, that it's what makes move forward any discipline, impels the architecture of essential way.

La tesis consta de una parte teórica y de una parte empírica. En la parte teórica se describe el enfoque cognitivo-evolutivo de Kohlberg (capítulo 1), el planteamiento cognitivo-afectivo de Eisenberg (capítulo 2), la teoría de Hoffman y otras sobre la empatía (capítulo 3) y la relación teórica entre el desarrollo del razonamiento moral, el razonamiento moral prosocial y la empatía (capítulo 4). En cada uno de estos temas se ofrecen también resultados de investigación empírica actual.El enfoque cognitivo-evolutivo de Kohlberg versa sobre el desarrollo del razonamiento moral, es decir, el razonamiento sobre dilemas morales en los que las leyes, normas, dictados de las autoridades y obligaciones formales son centrales. El planteamiento cognitivo-afectivo de Eisenberg trata sobre el razonamiento moral prosocial, esto es, el razonamiento sobre dilemas morales en los que las necesidades de una persona están en conflicto con las necesidades de otra en un contexto en el que el papel de las normas, leyes, dictados de las autoridades y obligaciones formales es mínimo. La empatía es la capacidad para ponerse en el lugar del otro junto con sentimientos de preocupación por el otro.Los objetivos de la investigación son a) analizar la influencia de las variables personales sexo, edad y nivel educativo sobre el desarrollo del razonamiento moral, sobre el razonamiento moral prosocial y sobre la empatía de los adolescentes y jóvenes, y b) relacionar teóricamente y examinar las interrelaciones empíricas entre el desarrollo del razonamiento moral, el razonamiento moral prosocial y la empatía. Estos objetivos se operacionalizan en trece hipótesis específicas sometidas a prueba, utilizando para ello una muestra de 556 sujetos de ambos sexos, de edades comprendidas entre los 13 y los 23 años, y cuyos niveles educativos son 3º ESO, 2º BUP, 3º BUP, COU, 1er Ciclo Universitario y 2º Ciclo Universitario. Como instrumentos de evaluación se utilizan el DIT de Rest (1979a), el PROM de Carlo, Eisenberg y Knight (1992), y el IRI de Davis (1980), todos ellos en su versión castellana.A partir de los resultados obtenidos en la investigación se extraen las siguientes conclusiones:* El sexo no produce ningún efecto significativo sobre el razonamiento moral de los adolescentes y jóvenes. Chicos y chicas obtienen similares puntuaciones.* En razonamiento prosocial, el sexo origina diferencias significativas. Los varones puntúan más que las mujeres en razonamiento orientado a la aprobación (aunque sólo los adolescentes), mientras que las mujeres puntúan más que los varones en los razonamientos estereotipado, internalizado y puntuación compuesta PROM.* En empatía también aparecen diferencias significativas debidas al sexo, de forma que varones y mujeres tienen una capacidad cognitiva similar para comprender la situación del otro, pero las mujeres están más predispuestas a una respuesta afectiva ante dicha situación.* La edad ejerce una influencia significativa sobre el razonamiento moral, sobre el razonamiento prosocial y sobre la empatía de los adolescentes y jóvenes.* Respecto al razonamiento moral, se produce con la edad una transición gradual de una moral convencional a una moral derivada de un código racional, de una moral sociónoma a una moral autónoma.* En razonamiento prosocial, los juicios morales llegan a ser menos egocéntricos y más orientados al otro, así como más abstractos, con el avance de la edad.* En empatía, a medida que incrementa la edad de los adolescentes y jóvenes aumentan progresivamente las puntuaciones en toma de perspectiva, fantasía y preocupación empática.* El nivel educativo tiene un efecto significativo sobre el razonamiento moral, sobre el razonamiento prosocial y sobre la empatía de los adolescentes y jóvenes. Tal efecto es similar al producido por la edad.* El razonamiento moral, el razonamiento prosocial y la empatía están interrelacionados empíricamente de forma positiva. Dado que en cada uno de estos tres constructos están implicados procesos cognitivos y afectivos, y que ellos están interrelacionados empíricamente de forma positiva, se concluye, a su vez, una relación positiva entre los procesos cognitivos y afectivos en el desarrollo moral de la persona. / The objectives of present research are a) to analyze the influence of sex, age and formal education variables on adolescents' and youngs' development of moral reasoning, prosocial moral reasoning and empathy, and b) to relate theoretically and to examine the empirical interrelations among development of moral reasoning, prosocial moral reasoning and empathy. A sample of 556 subjects is utilized. As instruments of evaluation are used the DIT, PROM and IRI. The results indicate that sex no has any significant effect on adolescents' and youngs' development of moral reasoning and, however, it affects significantly on prosocial moral reasoning and empathy. Moreover, the age and formal education exert a significant influence on development of moral reasoning, prosocial moral reasoning and empathy. Finally, the development of moral reasoning, prosocial moral reasoning and empathy are empirically positively interrelated. And given that in each one of these three constructs are implicated cognitive and affective processes, can be concluded, in his turn, a positive relation between the cognitive and affective processes on moral reasoning of person.

Thesis--University of Wisconsin--Madison. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 240-248).

Folates (vitamin B9) are essential water soluble vitamins, whose deficiency in humans may contribute to the onset of several diseases, such as anaemia, cancer, cardiovascular diseases, neurological problems as well as defects in embryonic development. Human and other mammals are unable to synthesize ex novo folate obtaining it from exogenous sources, via intestinal absorption. Recently the gut microbiota has been identified as an important source of folates and the selection and use of folate producing microorganisms represents an innovative strategy to increase human folate levels. The aim of this thesis was to gain a fundamental understanding of folate metabolism in Bifidobacterium adolescentis. The work was subdivided in three main phases, also aimed to solve different problems encountered working with Bifidobacterium strains. First, a new identification method (based on PCR-RFLP of hsp60 gene) was specifically developed to identify Bifidobacterium strains. Secondly, Bifidobacterium adolescentis biodiversity was explored in order to recognize representing strains of this species to be screened for their folate production ability. Results showed that this species is characterized by a wide variability and support the idea that a possible new taxonomic re-organization would be required. Finally B. adolescentis folate metabolism was studied using a double approach. A quantitative analysis of folate content was complemented by the examination of expression levels of genes involved in folate related pathways. For the normalization process, required to increase the robustness of the qRT-PCR analysis, an appropriate set of reference genes was tested using two different algorithms. Results demonstrate that B.adolescentis strains may represent an endogenous source of natural folate and they could be used to fortify fermented dairy products. This bio-fortification strategy presents many advantages for the consumer, providing native folate forms more bio-available, and not implicated in the discussed controversy concerning the safety of high intake of synthetic folic acid.

Gut microbial acquisition during the early stage of life is an extremely important event since it affects the health status of the host. In this contest the healthy properties of the genus Bifidobacterium have a central function in newborns. The aim of this thesis was to explore the dynamics of the gut microbial colonization in newborns and to suggest possible strategies to maintain or restore a correct balance of gut bacterial population in infants. The first step of this work was to review the most recent studies on the use of probiotics and prebiotics in infants. Secondly, in order to prevent or treat intestinal disorders that may affect newborns, the capability of selected Bifidobacterium strains to reduce the amount of Enterobacteriaceae and against the infant pathogen Streptococcus agalactiae was evaluated in vitro. Furthermore, the ability of several commercial fibers to stimulate selectively the growth of bifidobacterial strains was checked. Finally, the gut microbial composition in the early stage of life in response to the intrapartum antibiotic prophylaxis (IAP) against group B Streptococcus was studied using q-PCR, DGGE and next generation sequencing. The results globally showed that Bifidobacterium breve B632 strain is the best candidate for the use in a synbiotic product coupled to a mixture of two selected prebiotic fibers (galactooligosaccharides and fructooligosaccharides) for gastrointestinal disorders in infants. Moreover, the early gut microbial composition was affected by IAP treatment with infants showing lower counts of Bifidobacterium spp. and Bacteroides spp. coupled to a decrement of biodiversity of bacteria, compared to control infants. These studies have shown that IAP could affect the early intestinal balance in infants and they have paved the way to the definition of new strategies alternative to antibiotic treatment to control GBS infection in pregnant women.

This PhD thesis is focused on cold atmospheric plasma treatments (GP) for microbial inactivation in food applications. In fact GP represents a promising emerging technology alternative to the traditional methods for the decontamination of foods. The objectives of this work were to evaluate: - the effects of GP treatments on microbial inactivation in model systems and in real foods; - the stress response in L. monocytogenes following exposure to different GP treatments. As far as the first aspect, inactivation curves were obtained for some target pathogens, i.e. Listeria monocytogenes and Escherichia coli, by exposing microbial cells to GP generated with two different DBD equipments and processing conditions (exposure time, material of the electrodes). Concerning food applications, the effects of different GP treatments on the inactivation of natural microflora and Listeria monocytogenes, Salmonella Enteritidis and Escherichia coli on the surface of Fuji apples, soya sprouts and black pepper were evaluated. In particular the efficacy of the exposure to gas plasma was assessed immediately after treatments and during storage. Moreover, also possible changes in quality parameters such as colour, pH, Aw, moisture content, oxidation, polyphenol-oxidase activity, antioxidant activity were investigated. Since the lack of knowledge of cell targets of GP may limit its application, the possible mechanism of action of GP was studied against 2 strains of Listeria monocytogenes by evaluating modifications in the fatty acids of the cytoplasmic membrane (through GC/MS analysis) and metabolites detected by SPME-GC/MS and 1H-NMR analyses. Moreover, changes induced by different treatments on the expression of selected genes related to general stress response, virulence or to the metabolism were detected with Reverse Transcription-qPCR. In collaboration with the Scripps Research Institute (La Jolla, CA, USA) also proteomic profiles following gas plasma exposure were analysed through Multidimensional Protein Identification Technology (MudPIT) to evaluate possible changes in metabolic processes.

Much progress has been made in the past decade in elucidating the physiological, pathophysiological and pharmacological role of Hydrogen Sulphide (H2S). Recently a function of H2S virtually in every tissue of the human organism has emerged. However, the H2S-mediated regulation of bone homeostasis has been scarcely investigated. Despite a recent increased interest in the field, many fundamental issues remain indeterminate. The main objective of this study was to increase the basic knowledge on the role of H2S in bone through in vitro and in vivo studies and develop novel therapeutic strategies for bone diseases. Ex vivo experiments revealed that H2S-generating enzymes (Cystathionine-β-synthase, CBS; Cystathionine-γ-lyase, CSE) are expressed in human bone tissues and human bone-derived cells. In vitro experiments evidenced that CBS and CSE expression is a distinctive feature of the transition of mesenchymal stromal cells (h-MSCs) toward mature osteoblast. Furthermore, loss of function experiments on CBS and CSE during osteogenic differentiation of h-MSCs revealed an impaired mineralization ability. In vivo experiments in mice highlighted the role of CBS, CSE and H2S in the maintenance of bone homeostasis and CBS, CSE and H2S were found to be depleted in post-menopausal osteoporosis. Furthermore, our in vitro and in vivo data validated the use of H2S-donors as novel potential candidates for the treatment of bone pathologies. In particular H2S administration prevented and reversed ovariectomy-induced bone loss in mice. Based on these evidences, we firstly developed an H2S-releasing hybrid drug (DM-22) by modifying a clinically relevant anti-resorptive drug in order to improve the therapy of bone loss. DM-22 displayed improved biological properties compared to the parent drug; in particular, it increased the osteogenic differentiation ability of h-MSCs. Secondly, we developed an H2S-releasing scaffold to improve bone regeneration which was permissive for h-MSCs colonization and supported their osteogenic differentiation.

L’importanza del microambiente nella progressione metastatica iniziò a delinearsi sin dalla fine del XIX secolo attraverso la teoria del “seed and soil”, proposta dal chirurgo inglese Stephen Paget (1855-1926), secondo la quale determinati tumori sono in grado di formare metastasi in organi specifici, proprio come un seme trova il terreno giusto per poter attecchire. In termini molecolari, Paget intuì come la cellula tumorale potesse esprimere molecole riconosciute solo in determinati tessuti. Negli stessi anni, il patologo americano James Ewing (1866-1943) elaborò una teoria complementaria, la “anatomical-mechanical hypothesis”, suggerendo l’idea che i sistemi linfatico e vascolare svolgessero un ruolo preponderante nella diffusione passiva delle cellule verso un determinato sito anatomicamente accessibile. Nel 1982, Bissell delineò la teoria moderna secondo cui, alla base dello sviluppo del tumore, sia il microambiente sia le mutazioni genetiche svolgono un ruolo fondamentale. Negli ultimi decenni, infatti, accanto alla definizione di cancro come malattia genetica, si è delineata l’importanza fondamentale delle interrelazioni che intercorrono tra l'epitelio tumorale e il microambiente tissutale nel processo di tumorigenesi. Il tumore è da tempo considerato un tessuto complesso, le cui cellule mutate non agiscono da sole durante la progressione del cancro, ma reclutano le normali cellule circostanti come collaboratori attivi per instaurare un fenotipo neoplastico. Se da un lato l’instabilità genomica dei tumori garantisce loro un vantaggio evolutivo continuo, permettendo a cellule sempre più resistenti e aggressive di sopravvivere alle terapie, dall’altro la loro natura malleabile ha evidenziato la necessità di attuare strategie multiple: molti studi pre-clinici e clinici hanno suggerito l’efficacia di terapie combinate che abbiano come target non solo le cellule tumorali, ma anche le componenti del microambiente e, in particolare, quelle dello stroma. / The relevance of the microenvironment in metastatic progression began to take shape since the late nineteenth century through the theory of the "seed and soil", given by the English surgeon Stephen Paget (1855-1926), according to which certain cancers are able to form metastases in specific organs, just like a seed finds the right soil to take root. In molecular terms, Paget understood as cancer cell could express molecules recognized only in certain tissues. In the same years, the American pathologist James Ewing (1866-1943) developed a complementary theory, "anatomical-mechanical hypothesis ", suggesting that the lymphatic and vascular systems could play a predominant role in the passive diffusion of cells toward a particular site anatomically accessible. In 1982, Bissell delineated the modern theory that, at the basis of tumor growth, both the microenvironment and the genetic mutations play a key role. In recent decades, in fact, next to the definition of cancer as a genetic disease, it has outlined the fundamental importance of the interrelationships that exist between the tumor epithelium and tissue microenvironment in the process of tumorigenesis. The tumor has long been considered a complex tissue, the cells of which do not act alone during the progression of cancer, but they can recruit normal surrounding cells as active collaborators to establish a neoplastic phenotype. While the genomic instability of tumors guarantees them a continuous evolutionary advantage, allowing more and more aggressive and resistant cells to survive the therapies, their malleable nature has highlighted the need to implement multiple strategies: many pre-clinical and clinical studies have suggested the efficacy of combination of therapies targeting not only the tumor cells, but also the components of the microenvironment, in particular those of the stroma.

La PKCε e la PKCδ, chinasi ubiquitariamente distribuite e ad azione pleiotropica, sono implicate del differenziamento, sopravvivenza e proliferazione cellulare. Esse sono coinvolte nel processo differenziativo delle cellule staminali ematopoietiche e in fenomeni patologici associati al compartimento sanguigno. In questa tesi sono presentati i risultati riguardanti lo studio in vitro del ruolo di PKCε e PKCδ nel contesto del differenziamento megacariocitario, in particolare si caratterizza l’espressione e la funzione di queste chinasi nel modello umano e nel modello murino di Megacariocitopoiesi, normale e patologica. Confrontando le cinetiche dei due modelli presi in analisi nello studio è stato possibile osservare come in entrambi PKCε e PKCδ dimostrino avere una chiara e specifica modulazione nel progredire del processo differenziativo. Questi dati, se confrontati, permettono di affermare che PKCε e PKCδ presentano un pattern di espressione opposto e, nel modello umano rispetto a quello murino, reciproco: nell’uomo i livelli di PKCε devono essere down-modulati, mentre nel topo, al contrario, i livelli della chinasi risultano up-modulati durante lo stesso processo. Analogamente, le CD34+ in differenziazione presentano una costante e maggiore espressione di PKCδ durante la maturazione MK, mentre nel modello murino tale proteina risulta down-modulata nella fase più tardiva di formazione della piastrina. Le chinasi mostrano in oltre di agire, nei due modelli, attraverso pathways distinti e cioè RhoA nel topo e Bcl-xL nell’uomo. È stato inoltre verificato che l’aberrante differenziamento MK osservato nella mielofibrosi primaria (PMF), è associato a difetti di espressione di PKCε e di Bcl-xL e che una forzata down-modulazione di PKCε porta ad un ripristino di un normale livello di espressione di Bcl-xL così come della popolazione di megacariociti formanti propiastrine. I dati ottenuti indicano quindi che PKCε e PKCδ svolgono un ruolo importante nel corretto differenziamento MK e che PKCε potrebbe essere un potenziale nuovo target terapeutico nelle PMF. / Protein kinases C (PKC) are known to be ubiquitously distributed and to have pleiotropic effects. Isoforms epsilon (PKCε) and delta (PKCδ) are involved in the regulation of cell growth, survival and differentiation; in particular, they have been also investigated for their role in the hematopoiesis and in aberrant processes of differentiation along the erythroid and megakaryocytic lineages. In this PhD thesis, the results of an in vitro study about the role of these two kinases in models of megakaryocytic (MK) differentiation, both normal and pathological, are presented. The observations about PKCε and PKCδ kinetics show how these proteins have a specific modulation during the MK differentiation that results in an opposite pattern of expression and, in the murine model if compared with the human model, also a reciprocal one. In particular, in human megakaryocytopoiesis, PKCε results down-modulated, whereas in mouse its levels increase. Instead, PKCδ shows a high and steady expression in maturing CD34+ MK committed, but it is strongly down-modulated during the latest phases of platelet maturation in the murine model. The study also elucidates the different pathways PKCε and PKCδ work through, being an inhibitory action of PKCε on RhoA during proplatelets (ppt) formation in the mouse model while, in the human MK differentiation, platelets production is regulated by PKCδ through Bcl-xL. In this dissertation it is also demonstrated how in an aberrant megakaryocytopoiesis, as in the pathologic model of primary myeloproliferative neoplasm (PMF), PKCε is strongly deregulated and it results in an altered Bcl-xL expression. A forced down-modulation of this kinase restores a normal MK differentiation and ppt maturation. Therefore, the data presented show that PKCε and PKCδ play a key role in proper megakaryocyte maturation and that PKCε could be a potential new therapeutic target for PMF.

Bifidobacterium spp. are known as probiotic strains and recently new features emphasize their importance for human health, as EPSs and folate production. The relationship between Bifidobacterium spp. and their hosts is unknown, but probably links to peculiarities in the bifidobacterial cell-wall structures or to bifidobacterial ability to metabolize substrates from the host diet. Recently, a richness and diversity of bifidobacteria was observed in Callithrix jacchus and Saguinus midas, introducing the existence of a storehouse in primate guts. Several techniques were developed to deepen the microbial diversity, mainly based on the PCR. The RFLP-PCR of 16S rRNA gene represents a fast tool to distinguish human or animal origin bifidobacteria, useful in “Microbial Source Tracking” and probiotic selection. The project aim was the exploration of the bifidobacterial occurrence and diversity in evolutionary primate hosts to improve the knowledge about bifidobacteria distribution in non-human primates, and to identify bifidobacteria with new probiotic features (EPSs and folate production). 17 subjects from Strepsirrhini, Eulemur macaco, Eulemur rubriventer, Hapalemur alaotrensis and Lemur catta, and from Simiiformes, the New World Monkeys Callithrix jaccus, Pithecia pithecia, Saguinus oedipus and Saguinus imperator, and the Old World Monkeys, Chlorocebo aethiops and Macaca Sylvanus, were studied. Strains tested for probiotics traits, acid and bile tolerance, revealed B. aesculapii, B. myosotis and B. spp. MRM_8.19 strains as the most resistance. The folate production on strains from ring-tailed lemur and common marmoset revealed autotrophy only in strains from common marmoset. The distribution of microbial communities in non-human primates from 8 babies of common marmosets, golden faced saki and Barbary macaques and 11 adults of ring-tail lemurs, black lemurs, red-bellied lemur, Alaotran bamboo lemur, Barbary macaques, grivet, cotton top-tamarin and emperor tamarin, was carried out using ARDRA and rep-PCR. Results revealed a richness in both abundance and diversity of Bifidobacterium in primates.