京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第20440号 / 農博第2225号 / 新制||農||1049(附属図書館) / 学位論文||H29||N5061(農学部図書室) / 京都大学大学院農学研究科応用生命科学専攻 / (主査)教授 植田 充美, 教授 小川 順, 教授 渡邊 隆司 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

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京都大学 / 新制・課程博士 / 博士(農学) / 甲第23237号 / 農博第2444号 / 新制||農||1083(附属図書館) / 学位論文||R3||N5327(農学部図書室) / 京都大学大学院農学研究科応用生命科学専攻 / (主査)教授 植田 充美, 教授 小川 順, 教授 栗原 達夫 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

Contemporary diagnostics rely on expensive, time-consuming, and optically-limited mechanisms that prevent at-home point-of-care molecular diagnostics with the accuracy of laboratory tools and the convenience of affordability. In this Thesis, biosensing was explored with commercial two-dimensional (2D) materials which have been investigated extensively over the last two decades yielding a variety of sensor metrics for detecting biomolecules. 2D materials have intrinsic properties that depend on the quality of material and substrate surface being employed. Here, graphene/SiO2 and monolayer hexagonal boron nitride (hBN) capping layer on graphene/SiO2 field-effect transistors (FETs) were used. Until recently, monolayer hBN has not been commercially available at the wafer-scale and has been observed in the literature to augment the properties of graphene-based devices and better control of processing repeatability. The work in this Thesis combines biochemistry with the wafer-scale production and surface-dependent properties of graphene and monolayer hBN/graphene via a FET fabrication process circumventing the use of photoresist. This was done to avoid photoresist resin that may contaminate the transducer surface and contribute to repeatability issues when studying biochemistry with 2D materials. Briefly, surface engineering of graphene/SiO2 and hBN/graphene/SiO2 was done, and the transfer characteristics were measured as a function of either the concentration of protons, genes, or proteins. Compared to bare 2D materials, the pH sensitivity of the shift in Dirac voltage was enhanced to -99 mV/pH when using 8.6 nm of Al2O3 on hBN/graphene/SiO2 FET. Graphene devices were then engineered for sensing SARS-CoV-2 genome with a signal-to-noise ratio of 3 at 100 aM and a linearized sensitivity of +22 mV/molar decade of SARS-CoV-2 ribonucleic acid and a dynamic range of four orders of magnitude. This was done by conjugating single-stranded deoxyribonucleic acid to sub-percolation threshold gold nanofilms deposited directly on the graphene sensing mesa. Finally, the 2D devices were studied for detecting SARS-CoV-2 spike protein after being functionalized with rabbit immunoglobulin G (IgG) monoclonal antibody (mAb). Additionally, preliminary work was done regarding the partial reduction and fragmentation of anti-SARS-CoV-2 spike protein human mAb IgG in an approach to leverage gold-thiol chemistry for covalently bonding the IgG to the 2D sensing mesa. In summary, the utilization of wide- and zero-bandgap nanomaterials may have profound implications in augmenting molecular diagnosis and treatment of disease through economically decentralizing biosensing. / 2024-01-20T00:00:00Z

This thesis reports the investigation of Bismuth oxybromide (BiOBr) semiconductor material as an efficient photocatalyst for the sunlight harvesting as well as environmental cleanup. I have utilised different synthetic methodologies to obtain BiOBr and its derivatives, such as co-precipitation, ultrasonification, and photo-deposition; and have studied their structural and optical properties by X-ray diffraction and surface analysis techniques. I report the synthesis and characterisation of two new p-n heterojunction systems, AgBr-BiOBr and BiOBr-ZnFe₂O₄, and have performed initial studies on photocatalytic reaction and their catalytic decomposition mechanisms. I have also reported the surface modification method including the deposition of noble metal on BiOBr to investigate the role played by the noble metal and the interactions between semiconductor and metal using various characterisation measurements. Furthermore, a continuous series of BiOBr-BiOI solid solutions were synthesised, characterised and the photocatalytic degradation was performed on the as-obtained semiconductors, to study the band structure properties of the solid solutions.

There is an upsurge of research interest on Alloy 718 additively manufactured (AM) by electron beam powder bed fusion (EB-PBF) technique in aero and land-based gas turbine engines. However, the surface quality of the manufactured components has always been a major challenge. Several factors, including powder particle size, layer thickness, beam parameters, scanning strategies, and inclination angle of the build, govern the surface characteristics. Along with surface roughness resulted from partially melted powder particles, surface defects such as balls, satellites, microcracks as well as up-skin and down-skin surfaces can enhance the vulnerability of the manufactured parts to corrosion. When the surface is unable to withstand the exposed environment adequately, corrosion can be triggered. The surface-induced corrosion failures are increasingly becoming more challenging as the AM components often have complex geometries that render them even more difficult to finish. So, the relatively poor surface finish is the barrier to the full exploitation of the AM industry. In the present study, to achieve the desired surface quality, hence an improved high temperature corrosion performance, shot peening was implemented on Alloy 718 parts manufactured by EB-PBF. The high temperature corrosion behavior of the parts was investigated in an ambient air environment at 650 and 800 °C for up to 336 h. The underlying physical and chemical factors at play of the parts exposed to the corrosive environment were investigated too. The effect of topographical features (e.g., surface roughness) and microstructural characteristics (e.g., grain structure, phases, and defects) on high temperature corrosion behavior were analyzed by 3D surface profilometry, hardness test, optical microscopy (OM), scanning electron microscopy (SEM) equipped with energy disperse spectroscopy (EDS), X-ray diffractometry (XRD) and electron backscatter diffraction (EBSD). The surface roughness and high temperature corrosion rate of the parts was significantly reduced after shot peening.

The interaction at the surfaces of load bearing implant biomaterials with tissues and physiological fluids is an area of crucial importance to all kinds of medical technologies. To achieve the best clinical outcome and restore the function of the diseased tissue, several surface engineering strategies have been discussed by scientific community throughout the world. In the current work, we are focusing on one such technique based on laser surface engineering to achieve the appropriate surface morphology and surface chemistry. Here by using a pulsed and continuous wave laser direct melting techniques we synthesize three dimensional textured surfaces of calcium phosphate (Ca-P) based surface chemistry on Ti-6Al-4V. The influence of each processing type on the micro texture and phase evolution and thereby its associated effect on wettability, in vitro bioactivity, and in vitro biocompatibility are systematically discussed. For samples processed using the pulsed laser, it was realized that with increasing laser scan speed and laser pulse frequency there was a transition from surface textures with sharp circular grooves to surface textures with radial grooves and thereby improved hydrophilicity. For CW laser processing the results demonstrated improved hydrophilicity for the samples processed at 100 μm line spacing as compared to the samples processed at 200 μm line spacing. Owing to the importance of Si for cartilage and hard tissue repair, a preliminary effort for synthesizing Ca-P-SiO2 composite coating on Ti-6Al-4V surface were also conducted. As a future potential technique we also explored the Laser Interference Patterning (LIP) technique to achieve the textured surfaces and developed understanding on their wetting behavior. In the current work, by adjusting the laser processing parameters we were able to synthesize textured coatings with biocompatible phases. The in vitro bioactivity and in vitro vi biocompatibility of the coatings were proved by the precipitation of an apatite like phase following immersion in simulated body fluid (SBF), and increased proliferation and spreading of the MC3T3-E1 like cells. The results and understanding of the current research is encouraging in terms of looking at other bio-ceramic precursor compositions and laser process parameter window for synthesizing better textured biocompatible coatings.

The bone marrow (BM) niche provides hematopoietic stem (HSC) and progenitor cells with many exogenous cues that tightly regulate homeostasis. These cues orchestrate cellular decisions, which are difficult to dissect and analyze in vivo. This thesis introduces a novel in vitro platform that permits systematic studies of BM-relevant factors that regulate homeostasis. Specifically, the role of 3D patterned adhesion ligands and soluble cytokines were studied in a combinatorial fashion. Analysis of human HSC differentiation and proliferation at both population and single cell level showed synergistic and antagonistic effects of adhesion- and cytokine-related signals. Those effects were dependent on the cytokine concentration and the distribution and number of adhesion ligands. The aim of this thesis was to model the in vivo bone marrow with its porous 3D structure and different sized niche compartments using a microcavity culture carrier. The developed culture system presented extracellular matrix (ECM) adhesion ligands to the HSCs in various defined dimensions ranging from single- to multi-cell capacity. The 3D open well geometry of the microcavity carriers also allowed HSCs to freely explore different scenarios including homing, migration, adhesion, or suspension. Furthermore, the developed setup offered straightforward accessibility to analytical methods like cytometry and quantitative microscopy. Single cell analysis of adherent HSCs showed decreased DNA synthesis and higher levels of stem cell marker expression within single cell microcavities under low cytokine conditions . This effect was reflected in a decline of proliferation and differentiation with decreasing microcavity size. When the cytokine concentration was increased2 beyond physiological levels the inhibitory effect on proliferation and differentiation due to single-cell-microcavity adherence was diminished. This result highlighted the fine balance between adhesion related and soluble cues regulating HSC fate. Within small microcavities more adhesion related receptors were engaged due to the 3D character of the culture carrier compared to multi-cell wells or conventional 2D cell culture plates. This study demonstrated that adhesion-related signal activation leads to reduced proliferation and differentiation. This geometry-based effect could be reversed by increased cytokine supplementation in the culture media. For plane substrates, HSCs attachment to fibronectin or heparin initiated early cell cycle entry compared to non-adherent cells during the initial 24h. Cytokine supplemented media favored integrin activation that induced fast adhesion, ultimately leading to early cell cycle activation. However, after prolonged cell culture the system balanced itself with a lower cycling rate of adherent versus non-adherent HSCs. Furthermore, HSCs within the 3-dimensionality of the microcavities cycled less than 2D adherent cells. These findings additionally supported the above stated idea of limited HSC proliferation as a consequence of more adhesion-related signals overwriting cytokine driven expansion. To complement the various in vitro studies, an in vivo repopulation study was performed. Cultured HSCs derived from single cell microcavities outperformed freshly isolated HSCs in a competitive repopulation assay, indicating that carefully engineered substrates are capable of preserving stem cell potential. Overall the reported findings provide a promising in vitro culture strategy that allows the stem cell field to gain a better understanding of the impact of distinct exogenous signals on human HSCs, which discloses new concepts for the wide scientific community working towards tissue engineering and regenerative medicine. / Die Homöostase der Hämatopoietischen Stamm- und Vorläuferzellen (HSC) in der Knochenmark Nische wird von einer Vielzahl exogener Faktoren gezielt reguliert. Diese Faktoren orchestrieren intrazelluläre Vorgänge, deren in vivo Analyse kompliziert ist. Die vorliegende These widmet sich einem neuen biotechnologischen Ansatz, der systematische Studien von Knochenmark-relevanten Faktoren ermöglicht. Im Speziellen wurde die Rolle 3D-präsentierter Zell Adhäsionsliganden in Kombination mit verschiedenen Konzentrationen löslicher Zytokine untersucht. Die Auswertung der Proliferation und Differenzierung von humanen HSC auf Einzelzell- und Populationsebene offenbarte die synergistischen und antagonistischen Effekte von Adhäsions- und Zytokinsignalen in ihrer Abhängigkeit von der Verteilung und der Anzahl von Adhäsionsliganden sowie der Zytokinkonzentration. Um die poröse Struktur des Knochenmarks in vivo-ähnlich darzustellen, wurde eine Zellkultur Plattform mit Mikrokavitäten verschiedenster Dimensionen von Multi- bis Einzelzellgröße entwickelt und mit Molekülen der extrazellulären Matrix beschichtet. Die Vorteile dieser Plattform liegen in der offenen 3D-Geometrie dieses mikrokavitäten Kultursystems, die den Zellen ermöglichte verschiedene Wachstumsbedingungen bezüglich Homing, Migration, Adhäsion oder Suspension frei zu erkunden. Das leicht zugängliche Setup eignete sich zudem hervorragend für die zytometrische Analyse der Zellen oder die quantitative Mikroskopie. Die Einzelzellanalyse adhärenter HSC ergab eine Reduktion von DNA Synthese und eine höhere Expression von Stammzelloberflächenfaktoren innerhalb der Einzelzell-Mikrokavitäten bei niedrigen Zytokinkonzentrationen . Dieser Effekt spiegelte sich auch auf Populationsebene in verminderter Proliferation und Differenzierung mit abnehmender Größe der Mikrokavitäten wider. Wurde die Zytokinkonzentration jedoch weit über physiologische Bedingungen erhöht, verminderte sich der Effekt (reduzierte DNA Synthese und höhere Stammzellfaktorexpression) beschrieben für die Einzelzellmikrokavitäten. Dieses Ergebnis verdeutlicht die empfindliche intrazelluläre Balance, vermittelt durch Adhäsionsignale und löslichen Faktoren, die das Verhalten von HSCs regulieren. Aufgrund des 3D-Charakters des Zellkulturträgers wurden innerhalb kleiner Mikrokavitäten mehr Adhäsionsrezeptoren ringsum die Zelle aktiviert. Dieser Vorteil gegenüber den Multizellkavitäten oder der herkömmlichen 2D–Zellkultur ermöglichte eine hohe Anzahl adhäsionsvermittelter Signale mit entsprechend höherer Proliferations-inhibitorischer Wirkung. Je höher die Konzentration der Zytokine war, desto stärker erfolgte die Stimulation der Proliferation und Differenzierung. Auf 2D Substraten, initiierte Adhäsion zu Fibronektin und Heparin innerhalb der ersten 24h einen frühen Zell-Zyklus-Start im Gegensatz zu nicht adhärenten Zellen. Die Zytokine im Zellmedium förderten die Integrin Aktivierung, was zu einer schnellen Zelladhäsion führte. Die Adhäsionsrezeptoren wiederum kooperieren mit Zytokinrezeptoren im Zellinneren und begünstigten damit einen zeitigeren Zell-Zyklus- Start. Allerdings stellte sich danach ein Gleichgewicht im Kultursystem ein, wobei weniger adhärente Zellen als nicht-adhärente Zellen den Zellzyklus durchliefen. Des Weiteren war die Zellzyklusrate innerhalb von 3D Mikrokavitäten niedriger verglichen mit herkömmlichen 2D Substraten. Diese Ergebnisse bestätigen ferner obenstehende These, dass Zytokin-induzierte Zellexpansion durch erhöhte Zelladhäsions-vermittelte Signale überschrieben wird. Um die in vitro Studien zu komplettieren wurde ein in vivo Repopulationsversuch durchgeführt. HSC kultiviert auf Einzel-Zell-Mikrokavitäten übertrafen frisch isolierte Konkurrenz-Zellen in einem kompetitiven Repopulationsversuch. Dieses erste Ergebnis zeigt, dass sich der Zellgröße entsprechende Biomaterialien für die erfolgreiche Stammzell-Kultur eignen. Die Ergebnisse dieser Arbeit bieten eine vielversprechende in vitro Zellkulturstrategie, die ein besseres Verständnis der Einflüsse von exogenen Signalen auf HSC erlaubt und damit eine Grundlage für neue Erkenntnisse in Richtung erfolgreicheres Tissue Engineering und klinische Anwendungen im Bereich der regenerativen Medizin bildet.

This thesis focuses on the application of nanocrystalline transition metal oxide TiO2, WO3 and NiO thin films in new “green” building technologies. Specifically, their physicochemical properties in photocatalytic, self-cleaning and gas sensing applications are studied. There is an intimate connection between comfort issues, health, with connections to energy efficiency, leading to a need for intelligent building materials and green architecture. The importance of good indoor environment is augmented by the fact that modern man in developed countries spends some 90 % of his time inside buildings and vehicles. Poor air quality may lead to discomfort of the person inhabiting a building and in ultimately cause adverse health effects. Thin films of nanocrystalline TiO2 were prepared using reactive DC magnetron sputtering. Crystalline mesoporous films of WO3 and NiO were prepared using advanced gas deposition technique (AGD). The crystal structure, morphology, optical and chemical properties of the films were characterized by using grazing incidence X-ray diffraction (GIXRD), scanning electron microscopy (SEM), UV/Vis spectroscopy and X-ray photoelectron spectroscopy (XPS), respectively. The photocatalytic properties and adsorption of both organic and inorganic molecules on pure and functionalized films were probed by in situ Fourier transform infrared spectroscopy (FTIR). The gas sensing properties of sensors based on TiO2, WO3 and NiO were investigated by conductivity measurements and noise spectroscopy. It was found for the first time that NiO based thin film sensors can be used to detect H2S and NO2 at low temperatures – down to room temperature. Hybrid WO3 sensors functionalized with multiwalled carbon nanotubes (MWCNTs) were used to detect NO2, CO and NH3 gases. These hybrid gas sensors show improved recovery properties compared to unmodified WO3 sensors. TiO2 based gas sensors were able to detect low concentrations of H2S by noise spectroscopy provided that the sensors were irradiated by UV light. Furthermore we show that sulphur is photo-fixated in crystalline TiO2 films upon simultaneous SO2 gas exposure and UV irradiation. Studies of the kinetics and identity of the photo-fixated sulphur complexes show that these are formed by photo-induced reactions between oxygen and SO2 at oxygen surface vacancy sites in TiO2. The sulphur modified TiO2 films show interesting self-cleaning properties compared to the pure films. / Felaktigt tryckt som Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology 739