No description available.

This thesis aims to increase our understanding of mechanisms that influence larval dispersal in marine benthic invertebrates, particularly in the absence of strong oceanographic features (e.g. estuarine plumes, upwelling events, or markedly different water masses). Laboratory experiments identified behavioural mechanisms that regulate the vertical distribution of larvae in response to thermal stratification, and field studies in St. George’s Bay, Nova Scotia (NS), Canada, examined the relationship between larval abundance and physical variables (temperature, salinity, fluorescence, etc) and identified mechanisms that regulate larval distributions in situ. In the laboratory, I demonstrated that thermal stratification affects the vertical distribution of larvae by acting as a barrier to migration, or through temperature-dependent vertical swimming velocities. I also developed a random walk based model which highlighted that the key to successfully simulating larval response to temperature was 1) determining the temperature-dependent distribution of vertical swimming velocities and 2) the temporal autocorrelation in these velocities. In the field, the most striking pattern was that the larval distributions for species with similar swimming abilities were significantly correlated to one another at all scales (0.5 to 40 km). This suggests a common mechanism, related to larval swimming ability, which greatly influences the horizontal larval distribution. I found that the spatial scale of variability in larval distributions (~ 3 km) matches that in both the environmental variables and of coherent structures in current velocities (i.e. the tidal excursion). Results from an aggregation-diffusion model suggest that horizontal larval swimming could not be responsible for the observed level of aggregation in the larval horizontal distributions. I suggest that these horizontal patterns are the result of 1) an aggregative process (i.e. larvae swimming against a vertical current and maintaining their vertical position) and 2) a diffusive process which scales the aggregations to the scale of the coherent structures in current velocity (i.e. tidal excursion). In conclusion, this thesis increases our understanding of larval behaviour and its effects on larval dispersal. The results will be particularly useful to those who are interested in mechanisms regulate population connectivity, particularly those using bio-physical models to model dispersal trajectories.

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal

A new Laser Induced Fluorescence Capillary Zone Electrophoresis (LIF-CZE) bioassay to detect and study the catalytic activity of the sulfur assimilating enzyme commonly found in E. coli species; alkanesulfonate monooxygenase (EC 1.14.14.5) is described for the first time. This technique enables the possibility for direct injection onto a capillary for detection without the need for pre-concentration of sample and with minimal sample preparative steps prior to analysis. In this bioassay, a group of Fischer based cyanine dyes and two Oxazine (Nile red) derivatives were designed for further optimization as key Vis-NIR fluorescent substrate. In developing this technique, the test dyes were first assessed for their photophysical properties, based on four criteria; (1) photostable (2) solvatochromism (3) binding affinity towards both the monooxygenase active site and serum albumin and (4) chemical stability in strong electric field strength. Applying key dye characterization procedures including; molar absorptivity determination, quantum yield determination, photostability, solvatochromism and protein interaction studies it was determined that the Fischer indolium cyanine dyes were most suitable for the method development. The data revealed that under the test conditions, reduced flavin, the oxidative monooxygenase catalytically specifically converts the alkylsulfonate substituted cyanine dyes to the corresponding aldehyde. This new bioassay has proven to be quick, portable, sensitive, reliable and the exhibit the possibility of ‘on-the-spot’ detection; advantages not readily realized with other commonly applied techniques such as PCR, SPR, ELISA and GC used to study bacterial sulfur assimilation processes. In addition, recent literature results proposed by other research groups developing similar techniques showed strong reliance on GC analyses. Those assays involve the use of low molecular weight straight chain non-emissive alkanesulfonate substrates. Once enzyme catalysis occurs the aldehyde is formed becomes rather volatile and requires complex and tedious headspace sampling for GC analyses. This feature limits the in vitro applicability and eliminated the possibility in vivo development. Our goal is to further develop, optimize and present this CZE based bioassay as a suitable alternative to the current trends in the field while creating a more robust and sensitive in vitro monooxygenase detection method with the possibilities of in vivo application.

KRAS is a gene that plays a very important role in the initiation and development of several types of cancer. In particular, 90% of human pancreatic cancers are due to KRAS mutations. KRAS is difficult to target directly and a promising therapeutic path is its indirect inactivation by targeting one of its Synthetic Lethal Partners (SLPs). A gene G is a Synthetic Lethal Partner of KRAS if the simultaneous perturbation of KRAS and G leads to cell death. In the past, efforts to identify KRAS SLPs with high-throughput RNAi screens have been performed. These studies have reported only few top-ranked SLPs. To our knowledge, these screens have never been considered in combination for further examination. This thesis employs integrative analysis of the published screens, utilizing additional, independent data aiming at the detection of more robust therapeutic targets. To this aim, RankSLP, a novel statistical analysis approach was implemented, which for the first time i) consistently integrates existing KRAS-specific RNAi screens, ii) consistently integrates and normalizes the results of various ranking methods, iii) evaluates its findings with the use of external data and iv) explores the effects of random data inclusion. This analysis was able to predict novel SLPs of KRAS and confirm some of the existing ones.

No description available.

Ein Schlüsselbefund der Parkinson-Krankheit auf zellulärer Ebene ist das Auftreten von Protein-Einschlusskörperchen, sogenannten Lewykörperchen. Der Hauptbestandteil dieser Lewykörperchen ist pathologisch aggregiertes, fibrilläres α-Synuklein, ein Protein, welches Einfluss auf präsynaptische Vesikel, Protein- und Enzymfunktionen sowie den Dopaminstoffwechsel und den axonalen Transport hat. Bis heute ungeklärt ist die Ursache der Aggregation des Proteins. Zahlreiche Forschungsaktivitäten werden in diese Richtung unternommen. Die pathologischen Mechanismen, die zur abnormen Aggregation von α-Synuklein führen, bleiben noch weitgehend unbekannt. Ein großer Teil der Literatur unterstützt die Hypothese, dass α-Synuklein bei Punktmutationen oder erhöhter Expression anfällig für Aggregationen ist und damit Neuronen geschädigt werden. Die Einzelheiten dieses sukzessiven Aggregationsprozesses und die Mechanismen, die dabei letztlich den Zelltod verursachen, bleiben unklar. Alterungsprozesse und Umweltfaktoren sind entscheidende Risikofaktoren. Obwohl es immer mehr Hinweise gibt, dass α-Synuklein-Aggregate eine wichtige pathophysiologische Rolle spielen, wird bisher noch unzulänglich verstanden, wie die für die dopaminergen Neurone toxische Wirkung entfaltet wird. Als gesicherter Pathomechanismus der Degeneration der dopaminergen Nervenzellen gilt ein erhöhter oxidativer Stress. Es wird vermutet, dass er zur Aggregation des α-Synukleins beitragen kann. Ziel dieser vorgelegten Arbeit ist es, durch die Verwendung eines geeigneten Zellkulturmodells zur Aufklärung der beschriebenen pathologischen Mechanismen beizutragen. In dieser Studie wurden zwei artifizielle Modellsubstanzen in einer dopaminergen Primärzellkultur eingesetzt, die Pestizide Rotenon und Paraquat, um die pathologischen Verhältnisse in der Substantia nigra zu simulieren. Sie erzeugen oxidativen Stress durch Hemmung der mitochondrialen Atmungskette bzw. Redoxreaktionen mit molekularem Sauerstoff, was zum dopaminergen Zelltod führt. Im Rahmen dieser Studie gelang es, beide Parkinson-Zellkulturmodelle anhand der Lokalisierung, des Aggregationsverhaltens, des Einflusses auf die Mikroglia-Aktivierung sowie des Abbaus von α-Synuklein näher zu charakterisieren. Hierzu wurde α-Synuklein durch Fluoreszenzfärbung, Westernblot und Immunpräzipitation analysiert. Eine kurzzeitige Behandlung mit hoch konzentrierten Toxinen löst eine akute Degeneration dopaminerger Neurone aus, die so nicht der des Idiopathischen Parkinsons entspricht. Beim IPS erfolgt die Degeneration über Jahre hinweg. Um auch den Einfluss der zellulären Alterung auf die α-Synuklein-Aggregation zu zeigen, wurden die in dieser Arbeit verwendeten Zellkulturen über 46 Tage kultiviert und die Pestizid-Konzentration so eingestellt, dass etwa 25-50 % der dopaminergen Neurone absterben. Es konnte gezeigt werden, dass es nach chronischer Behandlung mit dem jeweiligen Pestizid zu den verschiedenen Zeitpunkten Unterschiede in der Lokalisation sowie in der Konformation von α-Synuklein gibt. Die zum Vergleich nur bis zum Tag 11 kultivierten Zellkulturen zeigten nach kurzer Behandlung mit hochkonzentrierter Toxin-Menge eine Ansammlung von α-Synuklein im Soma, aber keine Auswanderung und Lokalisierung in und an den Neuriten, wie es nach chronischer Rotenon-Behandlung beobachtet werden konnte. Bei den Rotenon-behandelten Zellen ist ein Prozess der Verlagerung und Anhäufung des α-Synuklein aus dem Soma in die Neuriten bereits ab einem früheren Zeitpunkt zu beobachten als in den Kontrollen, welche sich aber mit zunehmendem Alter ähnlich verhalten. Außerdem konnte in den Kulturen, besonders bei den Rotenon-behandelten dopaminergen Neuronen, ein punktförmiges Verteilungsmuster und größere Ansammlungen des Proteins in den Neuriten beobachtet werden, was für eine aggregierte Form des α-Synukleins spricht. Diese Aggregate ließen sich durch die Proteo-Aggreosom-Färbung nachweisen. Auch der Nachweis von am Serin 129 phosphoryliertem α-Synuklein in diesen größeren Ansammlungen gilt als Zeichen für eine aggregierte Form. Die Beobachtung, dass α-Synuklein mit zunehmendem Kulturalter aus dem Soma in die Peripherie austritt, konnte bei der chronischen Paraquat-Behandlung so nicht getätigt werden. Unter Paraquat-Behandlung war keine Herauf-Regulierung der Gesamt-α-Synuklein Expression in der Kultur zu beobachten, wie dies sowohl bei Kontrolle als auch bei Rotenon der Fall ist. Wir vermuten im Paraquat-Modell, dass die sich im Soma ansammelnde Form von α-Synuklein durch vermehrte Einschleusung in den Zellkern zur Toxizität beitragen kann. Auch eine Interaktion von α-Synuklein mit der Zellmembran könnte unter Paraquat-Einfluss zum dopaminergen Zelltod beitragen. Durch Immunpräzipitation und Fluoreszenz-Doppelfärbung von α-Synuklein und Ubiquitin konnten wir den Abbau des fehlgefalteten α-Synukleins in der Zelle durch Ubiquitinierung nachweisen. Unter Rotenon ist ab DIV 14 eine starke Kolokalisation von α-Synuklein und Ubiquitin zu erkennen, die im Verlauf der Kultur nachlässt und nur noch in punktförmigen Aggregaten außerhalb der Zelle zu sehen ist. Unter Paraquat zeigte sich während der gesamten Kultivierung Polyubiquitinierung und Kolokalisation der beiden Proteine in der gesamten Zelle. Die Aggregationsform von α-Synuklein scheint das Proteinabbausystem durch Ubiquitinierung zu beeinflussen, da wir davon ausgehen, dass es sich bei der α-Synuklein-Konformation zu verschiedenen Zeitpunkten in den Paraquat-behandelten dopaminergen Neuronen nicht um die Aggregationsform handelt, die wir unter Rotenon beobachten konnten.

L'objectif principal de cette recherche est d'analyser les effets de l'agrégation sur la prévision de la demande. Cet effet est examiné par l'analyse mathématique et l’étude de simulation. L'analyse est complétée en examinant les résultats sur un ensemble de données réelles. Dans la première partie de cette étude, l'impact de l'agrégation temporelle sur la prévision de la demande a été évalué. En suite, Dans la deuxième partie de cette recherche, l'efficacité des approches BU(Bottom-Up) et TD (Top-Down) est analytiquement évaluée pour prévoir la demande au niveau agrégé et désagrégé. Nous supposons que la série désagrégée suit soit un processus moyenne mobile intégrée d’ordre un, ARIMA (0,1,1), soit un processus autoregressif moyenne mobile d’ordre un, ARIMA (1,0,1) avec leur cas spéciales. / Demand forecasting performance is subject to the uncertainty underlying the time series an organisation is dealing with. There are many approaches that may be used to reduce demand uncertainty and consequently improve the forecasting (and inventory control) performance. An intuitively appealing such approach that is known to be effective is demand aggregation. One approach is to aggregate demand in lower-frequency ‘time buckets’. Such an approach is often referred to, in the academic literature, as temporal aggregation. Another approach discussed in the literature is that associated with cross-sectional aggregation, which involves aggregating different time series to obtain higher level forecasts.This research discusses whether it is appropriate to use the original (not aggregated) data to generate a forecast or one should rather aggregate data first and then generate a forecast. This Ph.D. thesis reveals the conditions under which each approach leads to a superior performance as judged based on forecast accuracy. Throughout this work, it is assumed that the underlying structure of the demand time series follows an AutoRegressive Integrated Moving Average (ARIMA) process.In the first part of our1 research, the effect of temporal aggregation on demand forecasting is analysed. It is assumed that the non-aggregate demand follows an autoregressive moving average process of order one, ARMA(1,1). Additionally, the associated special cases of a first-order autoregressive process, AR(1) and a moving average process of order one, MA(1) are also considered, and a Single Exponential Smoothing (SES) procedure is used to forecast demand. These demand processes are often encountered in practice and SES is one of the standard estimators used in industry. Theoretical Mean Squared Error expressions are derived for the aggregate and the non-aggregate demand in order to contrast the relevant forecasting performances. The theoretical analysis is validated by an extensive numerical investigation and experimentation with an empirical dataset. The results indicate that performance improvements achieved through the aggregation approach are a function of the aggregation level, the smoothing constant value used for SES and the process parameters.In the second part of our research, the effect of cross-sectional aggregation on demand forecasting is evaluated. More specifically, the relative effectiveness of top-down (TD) and bottom-up (BU) approaches are compared for forecasting the aggregate and sub-aggregate demands. It is assumed that that the sub-aggregate demand follows either a ARMA(1,1) or a non-stationary Integrated Moving Average process of order one, IMA(1,1) and a SES procedure is used to extrapolate future requirements. Such demand processes are often encountered in practice and, as discussed above, SES is one of the standard estimators used in industry (in addition to being the optimal estimator for an IMA(1) process). Theoretical Mean Squared Errors are derived for the BU and TD approach in order to contrast the relevant forecasting performances. The theoretical analysis is supported by an extensive numerical investigation at both the aggregate and sub-aggregate levels in addition to empirically validating our findings on a real dataset from a European superstore. The results show that the superiority of each approach is a function of the series autocorrelation, the cross-correlation between series and the comparison level.Finally, for both parts of the research, valuable insights are offered to practitioners and an agenda for further research in this area is provided.

Conselho Nacional de Desenvolvimento Científico e Tecnológico / This work reports the study of the molecular structure influence of ionic liquid (IL) derived from 1,8-bis(3-methylimidazolium-1-yl)octane in the aggregates formation in a solution ethanol in water (4,75%) and in ethanol (95%). Was varied the anions (Br-, NO3-, BF4-, SCN- e NTf2-) of IL and the aggregation behavior was investigated by various methods, such as differential scanning calorimetry (DSC), conductivity, surface tension, fluorescence and dynamic light scattering (DLS). In the ethanol in water solution (4,75%), the critical aggregation concentration values (cac) (114 to 205 mM), free energy of aggregation (ΔG°a) ( 15 to 18 kJ/mol) and ionization degree (α) (0,37 a 0,44) for the IL with Br-, NO3-, BF4-, SCN- significantly decreased with the increase of anion hydrophobicity. In ethanol, the cac values (165 to 500 mM) and ΔG°a ( 9 to 11 kJ/mol) also decreased with the increase of anion size and hydrophobicity. The free energy adsorption data (ΔG°ads) (-35 to -39 kJ/mol) demonstrated that the ILs, in general, have a good surfactant activity and this property improve with the decrease in the hydrophobic characteristics of anions. In general, in the solvents used in this study, the hydrodynamic radius (Rh) of aggregates ranged between 200 and 500 nm for all the ILs. The critical packing parameter was determined using data from the X-Ray (of IL 4) and surface tension, and showed that aggregates are micelar. The cac data was obtained by the techniques of conductivity, fluorescence and surface tension. Where, was observed that increasing the anion hydrophobicity of dicationic ILs structure favored the formation of aggregates, while less hydrophobic anions improved the surfactant properties of the IL structures. / Este trabalho relata o estudo da influência da estrutura molecular de líquidos iônicos (LI) dicatiônicos, derivados do cátion 1,8-bis(3-metilimidazolil-1-íneo)-octano, na formação de agregados em uma solução etanol em água (4,75%) e em etanol (95%). Variaram-se os ânions (Br-, NO3-, BF4-, SCN- e NTf2-) dos LI e o comportamento de agregação foi investigado através das técnicas de calorimetria exploratória diferencial (DSC), condutividade, tensão superficial, fluorescência e espalhamento de luz dinâmico (DLS). Na solução etanol em água (4,75%), os valores de concentração de agregação crítica (cac) (114 a 205 mM), energia livre de agregação (ΔG°a) ( 15 a 18 kJ/mol) e grau de ionização do contra-íon (α) (0,37 a 0,44) para os LI contendo os ânions Br-, NO3-, BF4-, SCN- diminuíram significativamente com o aumento da hidrofobicidade do ânion. Em etanol, os dados de cac (165 a 500 mM), ΔG°a ( 9 a 11 kJ/mol) para os LI com os ânions Br-, SCN- e NTf2- também diminuíram com o aumento do volume e hidrofobicidade dos ânions. Dados de energia livre de adsorção (ΔG°ads) (-35 a -39 kJ/mol) demonstraram que os LI dicatiônicos, de forma geral, possuem boa atividade tensoativa e esta propriedade aumenta com a diminuição das características hidrofóbicas dos ânions. Em geral, nos solventes utilizados no estudo, o raio hidrodinâmico (Rh) dos agregados variou entre 200 e 500 nm para todos os LI. Valores de parâmetro de empacotamento crítico (Pc) determinados a partir de dados de raios-X e tensão superficial indicaram que os agregados formados são micelares. Os dados de cac obtidos por condutividade, fluorescência e tensão superficial apresentaram-se de forma concordante. Observou-se que o aumento da hidrofobicidade da estrutura do ânion do LI dicatiônico favoreceu a formação dos agregados, enquanto que ânions menos hidrofóbicos melhoraram as propriedades tensoativas das estruturas dos LI.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / This paper presents the study of influence of molecular structure of ionic liquid-type Gemini imidazolium ìn the formation of aggregates in aqueous solution. The aggregation behavior was investigated through differential scanning calorimetry, conductivity, surface tension, fluorescence, dynamic light scattering techniques and transmission electron microscopy. Among the obtained data, it is possible to highlight the critical aggregation concentration (cac) determination, that is, a concentration range where a variation in physical and chemical properties of the solution emerging from the formation of aggregates can be observed. Differential scanning calorimetry showed that the aggregation can be detected by means of thermal events, characteristics of pure LI. Through the conductivity, it was possible to determine thermodynamic parameters, as the aggregate free energy (ΔG°a), the binding degree of the counterion to the aggregate (α), and the process equilibrium constant (Ka). Surface tension measurements provided data regarding the activity of the surfactant LI studied, while fluorescence, dynamic light scattering and transmission electron microscopy were important to obtain information on the aggregates' size. Critical packing parameter was determined throughout data obtained from X-ray diffraction and surface tension. This parameter allowed the prediction of LI micellar form currently in study. It was observed that the values of cac determined by the methods employed are consistent with each other and it was found that the increase of the carbon chain used as a spacer group promoted the aggregates formation, as well as caused an improvement in the surfactant LI properties. / Este trabalho apresenta o estudo da influência da estrutura molecular dos líquidos iônicos (LI) dicatiônicos derivados do imidazolíneo na formação de agregados em solução aquosa. O comportamento de agregação foi investigado pelas técnicas de calorimetria exploratória diferencial, condutividade, tensão superficial, fluorescência, espalhamento de luz dinâmico e microscopia eletrônica de transmissão. Dentre os dados obtidos, destaca-se a determinação da concentração de agregação crítica, que trata-se de uma faixa de concentração onde pode ser observado a variação nas propriedades físicas e químicas da solução emergente da formação de agregados. Por calorimetria exploratória diferencial observou-se que a agregação pode ser detectada por meio de eventos térmicos característicos dos LI puros. Através da condutividade foi possível determinar parâmetros termodinâmicos como a energia livre de agregação (ΔG°a), grau de ligação do contraíon ao agregado (α) e constante de equilíbrio do processo (Ka). As medidas de tensão superficial forneceram dados a respeito da atividade tensoativa dos LI em estudo enquanto que a fluorescência, espalhamento de luz dinâmico e microscopia eletrônica de transmissão foram importantes para obter informações a sobre o tamanho dos agregado. O parâmetro de empacotamento crítico foi determinado, para isto utilizou-se dados provenientes da difratometria de raios-X e tensão superficial. Este parâmetro possibilitou a previsão da forma micelar para os LI em estudo. Foi observado que os valores de cac determinados pelos métodos empregados são concordantes entre si e verificou-se que o aumento da cadeia carbônica utilizada como grupo espaçador tanto favoreceu a formação dos agregados como também provocou uma melhora nas propriedades tensoativas dos LI.