Die E-Zigarette gewinnt in den letzten Jahren immer mehr an Popularität. Die Frage der Toxizität ist jedoch noch nicht abschließend geklärt, und es besteht weltweite Unsicherheit bei der Verwendung der E-Zigarette. Die vorliegende Arbeit untersucht menschliche Nasenschleimhautzellen nach Dampfexposition mit Propylenglykol, einem Hauptbestandteil der Liquide, auf mögliche akute Entzündungsreaktionen, zytotoxische und genotoxische Wirkungen. Die Nasenschleimhautzellen von 10 Probanden wurden im Air-Liquid-Interface kultiviert und anschließend verschiedenen Konzentrationen von Propylenglykol ausgesetzt. Die Analyse erfolgte unter Verwendung eines Trypanblau-Tests, eines Comet-Assays, eines Mikrokern-Tests und eines IL-6- und IL-8-Sandwich-ELISAs. Der Trypanblau-Test zeigte keine Reduktion der Vitalität. Im Sandwich-ELISA konnte kein Anstieg der IL-6- und IL-8-Konzentrationen festgestellt werden. Im Comet-Assay zeigte das Olive Tail Moment in allen untersuchten Konzentrationen eine Schädigung im Vergleich zur Negativkontrolle. Es zeigte sich auch eine dosisabhängige Schädigung. Ein Unterschied zwischen der Reinsubstanz und der Negativkontrolle konnte im Mikrokern-Test festgestellt werden. Es wurden reparierbare Schäden im Comet-Assay gefunden. Im Mikrokern-Test konnten diese nur in der Reinsubstanzkonzentration bestätigt werden. Die E-Zigarette sollte restriktiv verwendet werden, bis Langzeitstudien vorliegen. Darüber hinaus sollten die Hersteller die Inhaltsstoffe der Flüssigkeiten eindeutig angeben. / The e-cigarette has become increasingly popular in recent years. However, the question of toxicity has not yet been clarified and there is global uncertainty in the use of the e-cigarette. The present work investigates propylene glycol, a major component of the liquids, for possible acute inflammatory reactions, cytotoxic and genotoxic effects on human nasal mucosal cells. The nasal mucosal cells from 10 volunteers were cultivated in the air-liquid-interface and then exposed to different concentrations of propylene glycol. The analysis was carried out using a trypan blue test, comet assay, micronucleus test and IL-6 and IL-8 sandwich-ELISA. The trypan blue test showed no reduction in vitality. No increase in IL-6 and IL-8 concentrations could be detected in the sandwich ELISA. In the comet assay, the Olive Tail Moment showed damage compared to the negative control in all examined concentrations. There was also a dose-dependent damage. A difference between the pure substance and the negative control could be found in the micronucleus test. Repairable damage in the comet assay have been found. In the micronucleus test these could only be confirmed in the pure substance concentration. The e-cigarette should be used restrictively until long-term studies are available. In addition, the manufacturers should clearly declare the ingredients of the liquids.

Air bubbles are a promising means of controlling fouling for a range of applications, particularly delaying fouling in marine environments. This work investigates the mechanism by which the collision of an air bubble with a solid removes adsorbed bacteria. A key feature of the work is that the numbers of bacteria were monitored via video microscopy throughout the collision, so we were able to explore the mechanism of bacteria removal. When a bubble collides with a solid, an air–water interface crosses the solid twice, and we were able to distinguish the effects of the first and second air–water interface. The bacterium Pseudomonas aeruginosa was allowed to adhere to smooth polydimethylsiloxane (PDMS) and then a collision with a bubble was investigated for one of three different approach geometries: perpendicular, parallel, and oscillating parallel to the solid surface. Other factors examined were the speed of the bubble, the duration of bacterial adhesion on the solid surface, and the wettability of the solid. Surface wettability was identified as the most significant factor. When the solid dewets, almost all bacteria were removed from hydrophobic surfaces upon the passage of the first air–liquid interface. In contrast, when a thin liquid film remained between the solid and the bubble (a hydrophilic solid), variable amounts of bacteria remained. Although almost all bacteria were initially removed from hydrophobic solids, many bacteria were redeposited on hydrophobic surfaces upon the passage of the second air–liquid interface, especially when the first and second air–liquid interfaces moved in opposite directions. As described previously, a lower velocity of the bubble allows more time for the thin liquid film to drain, and improved removal efficiency on hydrophilic solids. A rougher solid (8 µm diameter hemispherical protrusions) decreased the detachment efficiency because bacteria and liquid were able to shelter in concavities. Air bubbles are capable of removing bacteria over a range of conditions and are a potentially efficient means of combating biofilm growth for practical applications. / Master of Science / A major problem for equipment submerged in seawater is their eventual coverage in marine organisms including bacteria, barnacles, seaweed, and algae. This work investigates how effectively an air bubble removes bacteria adhered to a submerged solid. Adhered bacteria were observed and counted throughout the interaction of a bubble with a solid. When a bubble collides with a solid and is then removed, the bubble edge passes over the solid twice. The edge of the bubble is referred to as an air–liquid interface. The effects on adhered bacteria removal of the first and second passes of the bubble air–liquid interface were observed. Pseudomonas aeruginosa, a bacterial species common to both marine and medical environments, was allowed to adhere to flat solids made up of the polymer polydimethylsiloxane (PDMS) prior to a collision with an air bubble. The air bubble was collided with the solid in three distinct ways: directly from above, across the solid surface in one direction, and across the solid surface in one direction before being pulled back in the other direction. The speed of the bubble, the amount of time bacteria were adhered to the solid prior to bubble collision, and the extent to which the solid could be wet were all also examined for their effects on adhered bacteria removal. The extent to which a solid surface could be dewetted was identified as the most significant factor. For solids that are easily dewetted, almost all adhered bacteria were removed with the passage of the first air–liquid interface. Many bacteria were then redeposited back onto the solid surface upon the passage of the second air–liquid interface, especially when it moved in a direction opposite to the first. In contrast, for solids that are easily wet by water, variable amounts of bacteria remained after the first air–liquid interface swept across its surface. Slower moving air–liquid interfaces were also shown to be more effective at removing adhered bacteria. Solid surfaces with rough patterning made it more difficult to remove bacteria. Air bubbles can be an effective method to combat adhered bacteria and potentially prevent eventual biological growth on different types of underwater applications.

The toxicology of aerosols in occupational settings is often performed through particle collection on a filter followed by reconstitution into cell culture media which can alter the biological effects. Current in vitro exposure systems require additional instruments to control temperature and humidity, making the system bulky and difficult to take to the field. The Portable In Vitro Exposure Cassette (PIVEC) was designed for personal monitoring, characterized using copper nanoparticles, tested with alveolar cells, and set-up for real-time monitoring. Three differently sized copper nanoparticles, 40-800 nm, were dispersed as a dry aerosol and measured gravimetrically and on a number concentration basis to determine the deposition efficiency of the PIVEC. A549 cells, a human alveolar adenocarcinoma epithelial line, were exposed to the aerosols and oxidative stress and cell viability were monitored post-exposure. The deposition efficiency ranged from 0.5% to 18% depending on method of analysis and size of particle. Oxidative stress increased within the first two hours post exposure, however there was no significant difference in cell viability at the four hour time point at deposited doses up to 1.63 mg/cm2. Validation of the PIVEC was done in the laboratory using diesel exhaust. Metal oxide fuel additives are used to reduce emissions; however, additives have been shown to increase emitted nanoparticles. The PIVEC was used to determine the potential cytotoxicity and oxidative activity changes in A549 cells after exposure to either model particles or exhaust generated with or without a commercial, nano-cerium oxide based additive. Acellular experiments suggest a correlation between the deposition and the type of fuel used for the newly designed PIVEC. Cellular results suggest a decrease in cytotoxicity and no statistically significant effect on reactive oxygen species generation with the use of the nano-cerium oxide additive. Rapid monitoring of oxidative stress was performed using an enzyme-based biosensor. The functionalized biosensor uses cytochrome c to measure reactive oxygen species through electrochemical detection during aerosol exposures. When compared to a traditional biological assay, the biosensor response was similar. The PIVEC is a unique device, designed to monitor aerosols using air-liquid interface in vitro techniques including a real-time monitor for oxidative stress.

Les systèmes d’actionnement à distance pour la manipulation d’objets de taille micrométrique ont connu un développement sans précédent ces dernières années dans les laboratoires de recherche. Ils permettent de contrôler à distance le déplacement et l’orientation d’objets en appliquant des champs de force à distance. Ils sont donc spécialement adaptés pour agir dans des milieux confinés pour lesquels les problèmes d’accessibilité empêchent l’utilisation de manipulateurs robotisés classiques. De plus la faible inertie de ces systèmes permet d’atteindre des cadences de manipulation importantes. Parmi les effets physiques exploitables pour actionner ces systèmes une attention particulière est portée sur les effets magnétiques, en raison des niveaux de forces élevés atteignables. L’état de l’art montre un nombre de travaux importants dans le domaine de l’actionnement magnétique en environnement liquide. Cependant les vitesses atteignables sont limitées par les frottements visqueux. Quelques études font état de l’utilisation de l’actionnement magnétique en milieu ambiant, mais les forces d’adhésion entre le substrat et la particule rend le système peu répétable. Cette thèse propose une approche originale alliant faibles frottements visqueux et grande répétabilité en considérant un milieu de travail peu étudié : l’interface air/liquide. Cette thèse s’intéresse plus particulièrement à la modélisation, la caractérisation, l’analyse des performances et la commande d’un système d’actionnement magnétique à l’interface air/liquide pour des applications à l’échelle micrométrique. / In recent years research laboratories have shown an increasing interest for non-contact actuation systems at micrometer scale. These systems control both the displacement and the orientation of the objects using remote force fields. They are of major interest in confined spaces in which traditional approaches based on robotic grippers are not suitable. In addition high manipulation throughputs can be reached due to the low inertia of these systems. Several physical principles can be considered as the actuation source. Among them a special attention is given to magnetic forces due to the high forces that can be applied to the objects. A large amount of work has been carried out on magnetic actuation systems for manipulation in liquid environments. However velocities are limited by viscous effects. A few studies are reported on magnetic systems in ambient environments. However repeatability is decreased by adhesion forces between the substrate and the objects. This work proposes an original approach with a good trade off between low viscous effects and high repeatability based on the use of a specific environment: the air/liquid interface. This thesis presents the modelling, the characterization, the performance analysis and the control of a magnetic actuation system at the air/liquid interface for applications at the micrometer scale.

Although there are over 1,600 Engineered Nanomaterials (ENMs)-containing consumer products available, our understanding of ENM safety is still limited. Airborne ENMs can readily enter the human body through inhalation potentially leading to many adverse health effects such as cardiovascular and pulmonary diseases. The conventional in vitro submerged cell culture method was developed decades ago and has been widely used as a fast screening method to elucidate cellular toxicity upon exposure to hazardous materials; however, it has many limitations compared with the in vivo models. Our group has previously utilized and validated an integrated low flow system capable of generating and depositing airborne nanoparticles (NPs) directly onto cells at an air-liquid interface (ALI) condition, and our results confirmed that this exposure system produced reproducible toxicological data for ENMs including gold (Au), 16% silver coated onto silica (16% Ag-SiO2), and copper oxide (CuO). To further improve this ALI method for an even closer representation of the in vivo model, a co-culture model containing three cell lines (A549, THP-1 differentiated macrophages, and EA.hy 926) was established and validated for testing ENMs toxicity. The co-culture model was exposed to 16% Ag-SiO2 and CuO NPs under the same protocol (4 h ALI exposure with a concentration of 3.5 mg/m3) as monoculture (A549 only) for comparison. Toxicity was assessed by measuring cell viability, reactive oxygen species (ROS) production, lactate dehydrogenase (LDH) release, and interleukin (IL) 8 level. Results showed that 16% Ag-SiO2 NPs induced higher ROS generation, and CuO NPs produced a significant level of proinflammatory response compared with monoculture. In addition, the co-culture model exhibited a similar response with the primary human bronchial epithelia cell line (HBEC) in terms of ROS and IL-8 responses after CuO NPs exposure, suggesting a more advanced refinement of the conventional model for in vitro inhalation study.

Human practices have resulted in great damage to the environment. Carbon-depletion, water and air pollution, as well as global warming are examples of the environmental footprints caused by several industries and their related applications. Detergency (cleaning) is widely practiced operation in household, industry and institutional sectors and thus consumes significant amounts of water, energy and chemicals and, therefore, contributes appreciably to the environmental destruction. This process is still not fully understood, on a molecular level, and not acceptably optimised. Therefore, this study is a contribution toward a better fundamental understanding and optimisation of protein stain removal from interfaces, which may ultimately result in the development of environmentally friendly and sustainable cleaning products and technologies. In this research, the cleaning of rubisco, a grassy protein stain, from different surfaces (hydrophobic, hydrophilic, and dyed) using different cleaning agent formulations was investigated. These studies encompassed experimental work and fundamental analysis in terms of mathematical modelling. The results revealed a consistent correlation between enzyme adsorption kinetics and stain cleanability. Higher adsorption and desorption absolute rates ( a k and d k ) resulted in higher enzyme mobility and thus higher stain cleanability regardless of the enzyme adsorbed amount. Surface chemistry underlying the stain has indirectly influenced stain cleanability through the alteration of enzyme adsorption kinetics. Such fundamental findings may aid in screening enzyme candidates for detergent formulations and may also assist in designing easily cleanable surfaces. Another fundamental finding is the cooperative cleaning mechanism of surfactant and enzyme of rubisco stain from different interfaces. The break down of intermolecular physical bonds between interfacial rubisco molecules by surfactants and the break down of the intramolecular covalent bonds by enzyme resulted in a higher protein displacement from interfaces. The overall protein removal by the two different actions of surfactant and enzyme showed a superiority of a biosurfactant-enzyme formulation. This finding may have significant implications on developing sustainable detergents that have superior cleaning performance and no or minimal environmental hazard. Overall, the findings reported in this Ph.D. thesis may form a basis for further comprehensive scientific research, which may ultimately provide detergent market with more efficient and optimum cleaning products and technologies.

Cystic Fibrosis (CF) is a degenerative disorder that is often associated with chronic lung disease. CF is caused by mutations in the gene encoding the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) chloride channel, which lead to defective chloride and sodium ion movement across epithelia. Subsequent dehydration of the airway surface liquid (ASL) on airway epithelia, is associated with poor mucociliary clearance and chronic lung infection. The monogenic nature of CF, along with the accessibility of the lung, makes the disease amenable to gene replacement therapy. Gene therapy clinical trials have focused on replacing the mutated CFTR with a functional copy, which has led to improved chloride transport, but has shown no significant effects on sodium transport. An alternative strategy for CF gene therapy therefore, could be to reduce the expression of the epithelial sodium channel (ENaC) in the lung, using RNA interference (RNAi), combined with CFTR delivery. Developing a dual-function gene transfer vector could potentially restore chloride and sodium levels in the ASL and help alleviate CF lung disease. The aim of this thesis was to develop a recombinant lentivirus delivery system capable of simultaneously delivering CFTR expression and knocking down ENaC expression in the airways. A modular HIV vector genome plasmid was developed to allow simple insertion of various promoter elements, transgenes and knockdown sequences, for subsequent virus production. Insertion of the CFTR transgene and a short-hairpin RNA (shRNA) sequence targeting the ENaC alpha subunit (ENaCα) resulted in significant knockdown of human ENaCα and simultaneous expression of CFTR in A549 (human lung carcinoma) cell culture. Replacement of the ENaCα shRNA with an shRNA targeting the transcription factor BACH1 resulted in target gene knockdown and concomitant HMOX1 up-regulation, confirming specific knockdown effects, and demonstrating that the dual-function rLV vector could mediate target gene knockdown irrespective of the target. Attempts were made to knock down BACH1 in primary cultures of human bronchial epithelial cells grown at the air-liquid interface (ALI), but improved transduction efficiencies from the apical surface will be required to generate successful knockdown in this experimental model. These studies provide proof-of-principle for the utility of this versatile dual-function prototype virus. The dual function vector not only has the potential for treatment of CF lung disease, but could be readily altered to target other lung diseases where combinations of prolonged target gene knockdown and gene expression/up-regulation could collectively provide an appropriate therapy. In conclusion, the focus on the rational design of gene transfer vectors for specific therapeutic effects will aid the development and translation of gene therapy approaches to human studies.

Les nanoparticules (NPs) représentent un danger potentiel pour la santé des travailleurs et du grand public, notamment en cas d’exposition par voie respiratoire. Si une NP est évaluée in vivo comme toxique chez l’animal, cela peut inciter à prendre des mesures pour réduire l’exposition de l’Homme à celle-ci, avant qu’il y ait des conséquences sanitaires graves. Les études in vivo sont donc d’une importance capitale afin de diminuer les potentiels risques sanitaires des NPs pour l’Homme. Néanmoins, dans un contexte de réduction du nombre d’animaux utilisés et compte tenu du nombre important de NPs existantes et de leur grande diversité physico-chimique, la toxicologie a besoin de modèles alternatifs, comme le in vitro, permettant de prédire de manière fiable les potentiels effets pulmonaires chez l’Homme. Des progrès importants ont été faits pour développer des modèles in vitro pulmonaires plus physiologiques et des méthodes d’exposition permettant de simuler l’inhalation de NPs in vitro. Cependant, des incertitudes existent quant à la capacité de ces nouveaux modèles in vitro à prédire les réponses observées in vivo dans les poumons après exposition à des NPs. Dans ce contexte, l’objectif de ce travail a été d’évaluer la capacité de plusieurs méthodes in vitro, de complexité différente, à prédire les effets toxiques observés in vivo chez le rat après exposition aiguë (24h) pulmonaire à des NPs métalliques faiblement solubles de TiO2 et de CeO2. Dans un premier temps, des expérimentations in vitro ont été effectuées afin d’évaluer si exposer des cellules alvéolaires à l’interface air-liquide (ALI) à des aérosols de NPs de TiO2 et de CeO2, générait des résultats différents par rapport à des expositions classiques à des suspensions en submergé. Dans un second temps, des expérimentations in vivo par aspiration intratrachéale ont été réalisées afin de comparer les réponses toxiques pulmonaires in vitro avec celles obtenues in vivo. Pour comparer les réponses pulmonaires in vivo et in vitro, des référentiels de dose similaires, notamment la masse par unité de surface ou par macrophage, ont été utilisés. Après 24h d’exposition, des réponses biologiques significatives (inflammation principalement) ont été observées in vitro à des doses inférieures à l’ALI par rapport au submergé. Nous avons par ailleurs souligné la nécessité de prendre en compte les doses réellement déposées sur les cellules ainsi que le débit de dose pour effectuer les comparaisons entre les deux méthodes d’exposition in vitro utilisées. Nous avons ensuite comparé les résultats in vitro avec ceux obtenus in vivo. Nous avons constaté que la méthode ALI générait des résultats plus prédictifs du in vivo, en termes de niveau d’activation des réponses toxiques à 24h. Finalement, nous avons établi un classement des quatre NPs utilisées dans notre étude et celles-ci ont été classées similairement in vivo et in vitro et quelle que soit la méthode utilisée in vitro. Nous avons par ailleurs montré l’importance de considérer la surface active des NPs pour établir ce classement. En conclusion, notre approche nous a permis de mieux évaluer le fossé existant entre le in vivo et le in vitro. Nos résultats soulignent l’intérêt d’utiliser des méthodes in vitro plus réalistes et plus proches de la physiologie humaine dans le but de modéliser les potentiels effets indésirables des NPs pour l’Homme. Cela ouvre des perspectives quant à l’utilisation et au développement de méthodologies in vitro de plus en plus représentatives des conditions d’exposition in vivo. / Nanoparticles (NPs) represent a potential danger for workers and public, especially after inhalation. When a NP is shown toxic for the lungs in vivo in animals, this can incite regulators to implement measures to reduce human exposure risks. The in vivo studies are thus of utmost importance in reducing the potential health risks for humans. However, in a context of a diminution in the number of animals used in experimentations and considering the high number of NPs used and their physicochemical diversity, there is an urgent need for alternative methods, like the in vitro, which could be used to predict the potential health effects of NPs in human. Many progresses have been made recently to develop more physiological cell models and exposure methods simulating the inhalation of NPs in vitro. Nevertheless, uncertainties remain regarding the capacity of these new in vitro methods to predict the biological responses observed in vivo into the lungs after exposure to NPs. In this context, the aim of our study was to assess the ability of several in vitro methods, differing in complexity, to predict the adverse responses observed in vivo in rat lungs after acute exposure (24h) to several metallic and poorly soluble TiO2 and CeO2 NPs. For this, in vitro experimentations were first performed to assess if exposing alveolar cells in monoculture or in co-culture at the ALI interface to aerosols of NPs, generated different results compared to classic exposure in submerged conditions to suspensions. In a second step, rats were exposed by intratracheal aspiration of NP suspensions to compare the biological responses in vitro to those obtained in vivo. To compare the pulmonary responses in vivo and in vitro, similar dose metrics were selected, including the mass per surface unit or per macrophage. After 24h of exposure, significant biological responses (mostly inflammation) were observed at lower doses at the ALI compared to in submerged conditions. Moreover, we highlighted the necessity to take into account the deposited dose on the cells and the timing of the dose delivery in order to compare the two exposure methods used in vitro. When we compared the responses in vitro to those observed in vivo, we noticed that the ALI methods generated more predictive results than the submerged one, in term of biological activation levels after 24h of exposure. Finally, a ranking of the four NPs used in our study was provided and the NPs were ranked similarly both in vivo and in vitro and whatever the exposure method used in vitro. We also showed the importance of the surface area when ranking the poorly soluble NPs. In conclusion, the gap existing between the in vivo and the in vitro has been evaluated in our study. Our results highlighted the relevance of using more realistic in vitro exposure methods to model the potential adverse effects of NPs for human. This brings perspectives about using and developing in vitro methods mimicking more closely the in vivo exposure conditions.

The interfacial structuring of three non-halogenated ionic liquids was examined at the air-liquid interface for both dry and humid condition by analysis of X-ray reflectometry data using a slab model approach. The aim was to investigate the effect of humidity on the air-liquid interface. Pure ionic liquids as well as solutions of 5% w/w and 20% w/w in propylene carbonate were examined. Three different cations were used, a phosphonium cation ([P6,6,6,14]+) and two dialkylimidazolium cations ([C6C1Im]+ and [C10C1Im]+). The anion was bis(mandelato)borate ([BMB]-), which is a non-commercial anion that has been shown to exhibit excellent tribological properties. The results reveal that the presence of ambient water has an impact on the interfacial structure and the layering of all pure ionic liquids, as well as most cases of ionic liquid solutions. Exposure to humidity had the largest response in the case of all pure and diluted solutions of [C6C1Im][BMB], for which a more pronounced layering was observed. The [P6,6,6,14][BMB] solutions did not exhibit any significant changes when exposed to ambient water, with the 20% w/w solution proving to be the most stable. / Gränsskiktsstrukturen hos tre olika icke-halogenerade jonvätskor undersöktes vid luft-vätskegränsskiktet för både torra och fuktiga förhållanden genom analys av data från en röntgen reflektometer med en “slab model approach”. Målet var att undersöka effekten av luftfuktighet på luft-vätskagränsskiktet. Rena jonvätskor såväl som lösningar av 5 och 20 vikt-% utspädda i propylenkarbonat undersöktes. Tre olika katjoner användes, en fosforkatjon ([P6,6,6,14]+) samt två dialkylimidazoliumkatjoner ([C6C1Im]+ och [C10C1Im]+). Anjonen var bis(madelato)borat ([BMB]-), vilket är en icke-kommersiell anjon som har visat bra tribologiska egenskaper. Resultatet visar att närvaron av omgivande vatten har en påverkan på gränsskiktsstrukturen samt skiktning hos alla rena jonvätskor och de flesta utspädda lösningar. Exponering för fuktighet hade den största inverkan på alla rena och utspädda lösningar av [C6C1Im][BMB], för vilka en mer distinkt skiktning kunde observeras. [P6,6,6,14][BMB] lösningar uppvisade inga signifikanta ändringar vid exponering till fuktig luft, där den lösning med 20 vikt-% visade sig vara mest stabil.

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