Characterization and manipulation of lipid self-assembly to construct stable, portable synthetic lipid bilayers

The overarching goal of this research work is to further our understanding of lipid self-assembly and its organization at an oil-water interface to support the development of synthetic lipid bilayer systems that can be used in biologically relevant fields such as membrane biophysics, protein electrophysiology, development of synthetic biomolecules, drugs, nanoparticles and other applications. Self-assembly kinetics and interfacial properties of lipid monolayers formed at a liquid-air and liquid-liquid interface are characterized using Langmuir-Blodgett trough and pendant drop tensiometer. Insights gained from these studies not only allow us to answer questions related to droplet interface bilayer (DIB; a promising technique to assemble artificial lipid membranes) formation but also enable us to manipulate properties of monolayer in order to improve the potential of droplet interface bilayer by, a) increasing the number of phospholipids that can form DIBs, b) improving the success rate of DIB formation, and c) enhancing the electrical stability of bilayers formed. Owing to its wide range of applicability, novel efforts towards improving the durability and portability of DIB system are presented. In addition, this research work aims at using Nanoscribe direct laser writing — a state-of-the-art 3D printing device, to build 3D micro-scaffolds that can support lipid monolayers and bilayers that are suitable for high resolution optical studies

Evaluation of Population Structure and the Interspecific Relationship of Striped Marlin (Kajikia Audax) and White Marlin (K. Albida) Based on Traditional or Genome-Wide Molecular Markers

The istiophorid billfishes (marlins, spearfishes, and sailfish) are highly migratory pelagic fishes exhibiting broad and continuous spatial distributions in the Atlantic, Pacific, and Indian oceans. These species are targeted by a number of recreational, commercial, artisanal, and subsistence fisheries worldwide, and are also caught as bycatch in pelagic longline fisheries targeting tunas and swordfish. Though stock assessments have not been conducted for all istiophorids, assessments available for some species indicate that many istiophorid stocks are overfished and/or experiencing overfishing. However, the development of stock-specific recovery efforts is often impeded by a lack of information on basic species biology, including stock structure. The species status of some istiophorids is also uncertain, further complicating management efforts as well as strategies to conserve genetic diversity characteristic of distinct evolutionary lineages. In this dissertation, a molecular approach is used to address questions currently contributing uncertainty to the conservation and management of two istiophorid billfishes, white marlin (Kajikia albida) and striped marlin (K. audax). These closely related sister species are distributed in the Atlantic and Indo-Pacific oceans, respectively. Previous assessment of genetic population structure for white marlin based on mitochondrial (mt) DNA and five nuclear microsatellite markers suggested the possibility of population structuring for this species; however, results from the evaluation of mtDNA and 24 microsatellites across a larger number of samples, including a collection of larvae, are consistent with the presence of a single genetic stock (Chapter II). This result highlights the importance of analyses based on large numbers of molecular markers and samples, as well as a biologically informed sampling design, for studies of population structure in highly migratory pelagic species. Compared to the apparent lack of genetic population structure for white marlin, analysis of nearly 4,000 single nucleotide polymorphism (SNP) molecular markers across collections of striped marlin from the Pacific and, for the first time, Indian oceans resolved multiple genetically distinct populations (Chapter III). These populations correspond with striped marlin sampled from the western Indian Ocean, Oceania, North Pacific Ocean, and eastern central Pacific Ocean. Results from individual-based cluster analyses also suggest the presence of a second genetically distinct population in the North Pacific Ocean. Comparisons of replicate sample collections for some regions demonstrate the stability of allele frequencies across multiple generations. Finally, the uncertain species status of striped marlin and white marlin was evaluated using over 12,000 genome-wide SNPs surveyed across large numbers of exemplars per species (white marlin: n = 75, striped marlin: n = 250; Chapter IV). Results from individual-based cluster and maximum likelihood phylogenetic analyses suggest the presence of distinct evolutionary lineages for striped marlin and white marlin. This result is consistent with levels of genetic differentiation between striped marlin and white marlin which are an order of magnitude higher than those calculated between populations of striped marlin. Collectively, results of this dissertation provide practical insights for improving the conservation and management of white marlin and striped marlin, including revised stock structures which should be recognized in assessment and management plans for striped marlin. Future genomic studies should focus on addressing uncertainties regarding rangewide stock structure and species relationships for other istiophorids. Additionally, studies which continue to improve the genomic resources available for istiophorid billfishes and other large pelagic fishes may ultimately facilitate the evaluation of questions previously unexplored for the pelagic marine environment, such as localized adaptation and speciation

Molecular Evolution of Vasopressin and Oxytocin Receptor Genes in Owl Monkeys (aotus Azarai) of Northern Argentina

The arginine vasopressin (AVP) and oxytocin (OT) hormone pathways are involved in a multitude of physiological processes, and their receptor genes (AVPR1A and OXTR) have been implicated in increased partner preference and pair bonding behavior in mammalian lineages. This observation is of considerable importance for understanding social monogamy in primates, which is present in only a small subset of primate taxa, including Azara\u27s owl monkeys (Aotus azarai). Thus, it is the goal of this dissertation to examine the molecular evolution of AVPR1A and OXTR in owl monkeys to better understand how the pro-social behaviors related to those loci may have evolved. However, in order to properly contextualize functional neurogenetic variation related to such sociobehavioral pattterns, it is necessary to first establish the range of molecular variation occurring at non-related genetic loci. To address this issue, we sequenced the entire mitochondrial genome of two species of Aotus (A. azarai and A. nancymaae), and analyzed 39 haplotypes of the mitochondrial COII gene from ten different owl monkey taxa. Next, to understand the recent evolutionary history and genetic structure of our focal owl monkey population, we assessed variation of the mtDNA control region (D-loop) in 118 wild individuals. Furthermore, to establish our knowledge of genetic kinship and individual identity within the wild population, we investigated autosomal variation in the form of 24 short tandem repeat (STR) microsatellite loci. In concert with these analyses, we characterized the molecular features of AVPR1A and OXTR in A. azarai and other platyrrhines through direct sequencing, and demonstrated that there are substantial sequence differences at both loci across primate species. These data provide new clues on the possible basis of pair bonding in New World species, and may help to explain the sporadic appearance of monogamy in this infraorder. Specifically, despite a common molecular origin, we argue that the AVP and OT pathways have evolved in markedly different ways, due in part to their chromosomal locations and their relative proximity to regions of molecular instability. This study reinforces the notion that neurogenetic loci in primates have undergone significant evolutionary changes, and suggests that monogamy has arisen multiple times in the primate order through different molecular mechanisms

Computational Exploration of Flash-Boiling Internal Flow and Near-Nozzle Spray

Gasoline engines operating under the principle of direct injection are susceptible to flash-boiling due to superheated nature of the fuel and the sub-atmospheric in-cylinder pressures during injection. A review of the literature on flash-boiling sprays shows that a majority of the studies have focused on the far-field regions of the spray, with limited attention given to understanding the influences of the injector geometry and the near-nozzle regions of the spray. Modeling the internal nozzle flow and the primary atomization, on which the far-field spray depends, is a challenge. This thesis, therefore, is aimed at understanding the complex flow through a fuel injector nozzle and the nature of the spray in the near-nozzle region, with the help of computer simulations under flash-boiling and non-flash-boiling conditions. In the current study, the simulations were performed using an in-house Eulerian CFD solver called HRMFoam. Improvements to the solver\u27s near-nozzle spray modeling capability are discussed. These improvements include the implementation of a liquid-gas interface-area-density transport equation to model the primary atomization process. The simulations of direct injection of gasoline and gasoline-like sprays were performed on single-hole and multi-hole injectors, for a wide range of operating conditions. Spray characteristics such as the nozzle\u27s coefficient of discharge and the mean droplet diameter in the dense region of the spray were seen to be captured adequately well with the help of a 2D axi-symmetry assumption in the case of single-hole injectors. A novel approach to identify the near-nozzle spray plume boundary in CFD simulations is presented and validated against experimental measurements for a single-hole asymmetric injector. Case studies on single-hole asymmetric injectors revealed a direct correlation between the drill angle of the nozzle and near-nozzle spray plume angle. A hypothesis of the similarity between a stepped-hole two-phase nozzle and a conventional single-phase converging-diverging nozzle is presented. Furthermore, it was observed that flash-boiling jets behave as underexpanded jets, and therefore, are wider. Whereas, non-flash-boiling behave as overexpanded jets, and thus are narrower. Through the case studies on multi-hole injectors, the collapse of the spray or lack thereof was qualitatively and quantitatively characterized. In this process, a resemblance between the experimentally and computationally identified spray collapse mechanism was established. The application of LES modeling to internal and near-nozzle GDI sprays was explored in a pilot study, and the results were qualitatively validated against the experimentally available near-nozzle X-ray radiography measurements. Finally, in another pilot study, an attempt to model the interphase slip velocity is discussed

Structural Dynamics of Copper Nanomaterials for CO2 Electrocatalysis

Electrons are the currency of the future energy economy. With a host of renewable sources of electrical energy and steadily decreasing cost of generation, our ability to store that electricity via chemical bonds becomes increasingly paramount. Development of materials that allow the transformation of electrical energy to chemical energy, e.g. the electroreduction of CO2 to value-added chemicals and fuels, will push our energy infrastructure to new heights. Despite considerable progress in thermal gas-phase heterogeneous catalysis of CO2, the heterogeneous electrocatalysis of aqueous CO2 under room temperature and neutral pH has remained challenging. The key gating obstacle has been the development of catalysts that effect efficient and selective formation of higher order products such as methane or multicarbons, for which only copper has emerged as a candidate material. To design next-generation electrocatalysts for CO2 conversion to multicarbons (CO2-to-C2+), the relationship between activity/selectivity and catalyst structure must be better understood, to identify the structural motifs that define CO2-to-C2+ active sites. Recent work has consistently revealed that copper nanomaterials undergo considerable structural change under operating conditions. Thus, a central challenge to the understanding of these active surfaces is their dynamic nature under operation in electrochemical conditions, especially as applied to nanoscale electrocatalysts. Hence, this dissertation centers around structural dynamics of copper-based nanocatalysts under CO2 electroreducing conditions.After a brief introduction to the problem statement and fundamental concepts related to heterogeneous electrocatalysis of CO2 to value-added products, I discuss the prospects and existing work around Cu structural evolution in Chapter 1. In Chapter 2, I show how the link between structural evolution and catalytic activity change can be clearly shown on a Cu nanowire catalytic platform, and further show how strategies that mitigate structural change also impact selectivity retention. In Chapter 3, I move to a copper nanoparticle ensemble platform with intriguing activity for multicarbon formation, coupled with a striking structural evolution. I illustrate the dichotomy between the apparent evolution and active structure via conventional ex situ measurements, and the structural and catalytic insights revealed by more comprehensive investigation using preservation strategies. In Chapter 4, I discuss how in situ characterization techniques assist the illumination of such structural evolution under electrocatalytic conditions, and further explore what additional advances are needed to harness these insights. Finally, I close in Chapter 5 with an outlook on materials development for CO2 electrocatalysis as it stands at time of writing. Overall, this dissertation seeks to provide a narrative of the relationship between an electrochemical materials scientist and the concept of structural dynamics. Through the works to be described, this dissertation will take the reader on a journey from emphasizing stabilization and mitigating structural change, towards understanding it for the eventual purpose of leveraging such structural change as another dimension of electrocatalyst design

Study of impact of reduced Sox4 expression levels in homeostasis, cancer and ageing

Tesis doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 12-05-2014Sox4 belongs to the SoxC class of transcription factors (encompassing Sox4, Sox11 and Sox12) within the Sry-related HMG box-containing superfamily of proteins. Sox4 expression peaks during embryonic development in neural crest and mesenchyme, where it regulates the differentiation and survival of mesenchymal and neural progenitors. Sox4 expression has also been found in thymus, spleen, developing pancreas and hair follicles. During embryogenesis, Sox4 has been implicated in a plethora of processes including the maturation of B and T lymphocytes, the modulation of myeloid differentiation, pancreatic islet development, osteoblast development and proper cardiac cushion formation. Sox4-/- mice display embryonic lethality at E14.5, mainly due to profound cardiac malformations, concomitant with defective haematopoiesis and pancreatic islet formation defects. In adults, the Sox4 is restricted to few tissues such as haematopoietic compartment, female reproductive system, intestinal crypts, pancreatic islets and activated hair follicles. These constrictions suggest a tight regulation to ensure proper tissue homeostasis; indeed, deregulated Sox4 expression is frequently associated to malignant cellular transformation. Sox4 can act as a potent oncogene that promotes the survival of transformed cells and favours anchorage-independent cell growth and motility. High expression of Sox4 is associated with leukemogenesis, inhibition of apoptosis and increased cell viability. Moreover, increased Sox4 expression is often linked to the acquisition of an invasive phenotype by promoting the initiation of the EMT transcriptional program, and therefore is often associated to metastatic colonization of distant tissues. To study the role of Sox4 in the adult organism, we first generated mice with reduced whole-body Sox4 expression. These mice display a plethora of age-related degenerative disorders and reduced spontaneous cancer incidence, indicating a role for this protein in maintaining adult tissue homeostasis and in tumour growth. To specifically address a role for Sox4 in adult stem cells, we conditionally deleted Sox4 (Sox4cKO) in stratified epithelia. Sox4cKO mice show increased skin stem cell quiescence and DNA damage accumulation, accompanied by resistance to chemical carcinogenesis. These phenotypes correlate with downregulation of cell cycle, DNA repair and skin stem cell differentiation genes. Altogether, our findings highlight the importance of Sox4 in adult tissue homeostasis and cancerLa expresión de Sox4 es máxima durante el desarrollo embrionario en la cresta neural y el mesénquima, donde regula la diferenciación y desarrollo de precursores neuronales y mesenquimales. Así mismo, también se ha detectado expresión de Sox4 en otros tejidos como timo, bazo, páncreas y folículos pilosos. Durante la embriogénesis, Sox4 está implicado en un vasto panel de procesos que incluyen maduración de linfocitos B y T, modulación de la diferenciación mieloide, desarrollo de islotes pancreáticos y osteoblastos, y adecuada formación del canal auriculoventricular. Ratones deficientes en la expresión de Sox4 presentan letalidad embrionaria debido a profundas malformaciones cardíacas, que cursan con defectos en hematopoyesis y en la formación de islotes pancreáticos. En adultos la expresión de Sox4 se restringe a unos pocos tejidos como la médula ósea, sistema reproductivo femenino, criptas intestinales, islotes pancreáticos y folículos pilosos activados. Dichas constricciones son sugestivas de una fuerte regulación que asegure una adecuada homeostasis tisular; de hecho, la desregulación de la expresión de Sox4 en adultos se encuentra asociada a transformación celular. Sox4 puede actuar como un potente oncogén que promueve la supervivencia de células transformadas y favorece el crecimiento independiente de anclaje a substrato. La expresión elevada de Sox4 está estrechamente asociada a leukemogenesis, inhibición de la apoptosis y aumento de la viabilidad celular. Además, el aumento de la expresión de Sox4 se asocia frecuentemente a la adquisición de un fenotipo invasivo dado a que es capaz de promover el programa transcripcional asociado a la EMT, y por lo tanto a menudo se asocial a la colonización metastática de tejidos distantes. Para estudiar Sox4 en adultos, hemos generado ratones con niveles reducidos de Sox4 en todo el organismo. Dichos ratones muestran un panel de enfermedades asociadas al envejecimiento y resistencia frente al cáncer espontáneo, indicando un papel para Sox4 en el mantenimiento de la homeostasis tisular y en tumorogénesis. Para estudiar en detalle el papel de Sox4 en células madre adultas, hemos delecionado Sox4 de forma condicional en epitelio estratificado (ratones Sox4cKO). Estos ratones muestran mayor quiescencia en las células madre de la piel junto a una mayor acumulación de daño en el DNA y resistencia a carcinogénesis química. Dichos fenotipos correlacionan con una regulación negativa de genes implicados en ciclo celular, reparación de daño en el DNA y diferenciación de células madre de la piel. Estos resultados resaltan la importancia de Sox4 en la homeostasis tisular adulta y cáncer

Development of the control system of the ALICE Transition Radiation Detector and of a test environment for quality-assurance of its front-end electronics

Im Rahmen dieser Arbeit wurde das Detektor-Kontroll-System (DCS) für den Übergangsstrahlungsdetektor (TRD) des ALICE Experiments am Large Hadron Collider entwickelt. Das TRD Kontrollsystem ist vollständig implementiert als eine detektororientierte Hierarchie von Objekten, welche sich wie End-Zustandsautomaten verhalten. Es kontrolliert und überwacht über 65 tausend front-end Elektronik (FEE) Einheiten, einige hundert low-voltage und eintausend high-voltage Kanäle, sowie weitere Subsysteme wie Kühlung und Gasversorgung. Die Inbetriebnahme des TRD Kontrollsystems fand während mehrerer Datennahmen mit ALICE unter Verwendung von Ereignissen aus der kosmischen Strahlung statt. In einem weiteren Teil dieser Arbeit wurde ein Test-setup zur Qualitätssicherung der Massenproduktion von über viertausend FEE Readout-boards mit insgesamt 1.2 Millionen elektronischen Auslesekanälen des TRD entwickelt. Die Hardware- und Softwarekomponenten werden im Detail beschrieben. Zusätzlich wurde vorher eine Reihe von Leistungsuntersuchungen durchgeführt, welche die Strahlungstoleranz des TRAP-chips überprüft, der den Haupt\-bestandteil der TRD-FEE darstellt

CHARACTERIZATION OF THE MULTIFUNCTIONAL PROPERTIES OF ZINC OXIDE BASED MATERIALS BY SCANNING PROBE MICROSCOPY TECHNIQUES

Ph.DDOCTOR OF PHILOSOPH