<p>Radiation induced bystander effects have given the cancer risk analysis a whole new paradigm. However the actual mechanism involved in producing the effects is still not clear. The basic bystander signal is assumed to be a biological signal. In this study we proposed and tried to quantify the presence of a physical signal in the form of electromagnetic radiation that can trigger a biological response in the bystander cells. In bystander effect studies where the cells are exposed to very low fluence of charged particles there could be several regions that can produce electromagnetic radiation due to the process of atomic/molecular excitations and relaxations. We focused on quantifying the number of ultraviolet photons emitted when charged particles pass through different media that have relevance to radiation biology experiments. The choice of UV photons was made due to the reason that its effects on living cells are very well documented. For this purpose we developed a system which employed the technique of single photon counting to measure the light emitted from samples irradiated under vacuum by a charged particle beam. Photon counting was done using a fast photomultiplier tube (Hamamatsu R7400p) with a peak cathode response at 420 nm wavelength.</p> <p>In the early set of “proof of principle experiments” we tested polystyrene targets for ion beam induced luminescence. Polystyrene is one of the materials that are used as a cell substrate for radiation biology experiments. The luminescence yield from polystyrene was measured in terms of absolute value i.e. number of photons per second per unit solid angle. The output appeared to have a non-linear behavior with the incident Ion fluence: it rose exponentially to an asymptotic value. We irradiated the samples with beam energies varying from 1 MeV to 2.0 MeV and showed saturation at or before an incident fluence rate of 3×10¹³ H⁺/cm²s. The average saturation value for the photon output was found to be 40 × 10⁶ cps. Some measurements were performed using filters to study the emission at specific wavelengths. In the case of filtered light measurements, the photon output was found to saturate at 28×10³, 10×10⁶, and 35×10⁶ cps for wavelengths of 280±5 nm, 320±5 nm and 340±5 nm respectively. Using the IBIL signal evolution characteristics with the ion fluence we determined the ions produce a damage having a cross section of the order of 10^-14 cm² in polystyrene. The average radiant intensity was found to increase at wavelengths of 280 nm and 320 nm when the proton energy was increased. Having found an evidence of a significant production of UV in ion irradiated, biologically relevant, material we extended this study further into the measurements from other relevant materials in radiation biology.</p> <p>Here charged particle irradiation was performed using positively charged protons (H⁺) ranging in energy from 1.2 MeV to 2.2 MeV at a fluence rate of 2.7×10¹⁰ protons mm^-2s^-1.The materials chosen for this study were polypropylene, Mylar, Teflon, and Cellophane as they are all materials commonly used in radiation biology experiments as cell substrates or containers. In addition, we performed measurements of two NIST standard materials derived from living cells: oyster tissue and citrus leaves. These materials were measured as a powder.</p> <p>All the container materials were found to emit UV frequency photons at emission levels that are significant enough to warrant further investigation of the potential biological consequences. In addition, the NIST standard reference materials oyster tissue and citrus leaves also emitted UV when irradiated. This suggested that biological materials may themselves emit UV at significant levels when irradiated with charged particles.</p> <p>We established this further by irradiated cells with β-particles. Cells were plated in Petri-dishes of two different sizes, having different thicknesses of polystyrene (PS) substrate. Exposure of the cell substrates (polystyrene) only resulted in the production of 1035 photons per unit activity in μCi of ⁹⁰Y which was equivalent to an exposure of 840 <em>β</em>-particles/cm² to the substrate. For a collimated electron beam exposure, we observed 158-167 photons per unit μCi (18 β-particles per cm² on the substrate) for different thicknessesof the substrate. Upon irradiating HPV-G cells plated on the PS dishes we determined that the luminescence gradually increased with the increasing exposure of β-particles; reaching up to 250 % of that of the luminescence without any cells for an activity of 180 μCi. For general irradiation conditions we found statistically significant difference in luminescence output for varying cellular densities with cells only and with the application of medium on top of the cells. The colourless medium increased the total luminescence yield while the coloured medium decreased it. When the cells were irradiated using a collimated beam of electrons it was found that the luminescence decreases with the increasing cellular density thus providing an evidence of re-absorption of photons within the surroundings.</p> <p>After establishing the fact that charged particles induce light emission from the materials that have a relevance to the radiation biology experiments. We extended our study further to find out other sources of radiation that could affect the dose distribution in radiation biology experiments. In radiation biology experiments the low doses of radiation are usually delivered usingamicrobeam charged particle accelerator. Microbeams delivers a highly localized and small dose to the biological medium by using a set of collimators that confine the charged particle beam to a very narrow (micron level) region. Since the collimation block a significant proportion of the beam therefore there is a chance of the production of low energy x-rays and secondary electrons. We used Monte Carlo simulations to investigate the production of particle induced x-rays and secondary electrons in the collimation system and its possible effects on the final dose delivery to the biological medium. We found no evidence of the escape of x-rays or secondary electrons from the collimation system for proton energies of up to 3 MeV. The thickness of the collimators was sufficient to reabsorb all the generated low energy x-rays and secondary electrons. However if the proton energy exceeds 3 MeV then a significant proportion of 10 keV and 59 keV (K-α) x-rays can be emitted by the collimator. Further it was established that due to the phase space distribution of particles in various orientations with the beam axis there are significant chances of hitting non-targetted cells in microbeams that employ a collimator to confine the beam.This may happen due to the beam particles travelling obliquely with the beam axis thus passing the collimator edge and hitting the non-targetted cells. Another reason could be the scatter of beam particles inside the collimator.</p> <p>The evidence of the production of UV in materials relevant to the radiation biology experiments suggest that the conclusions and hypotheses derived from some radiation bystander experiments need to be re-thought, as charged particle irradiation leads to some level of UV emission in experimental materials which may be the cause of some “non-targeted” effects.</p> / Doctor of Philosophy (PhD)

Silicon has been the mainstay of the microelectroncs industry for over four decades. There is no material which can match the balance it affords between cost-benefit, mass consumability, process versatility, and nano-scale electron device performance. It is, therefore, the logical (and perhaps inevitable) platform for the development of integrated opto-electronics - a technology that is being aggressively developed to meet the next generation of bandwidth demands that are already beginning to strain interconnect architectures all the way down to the intra-chip level. While silicon-based materials already provide a variety of passive optical functionalities, the success of a genuine silicon-based optoelectronics will depend upon the ability of engineers to overcome those limitations in the optical properties of bulk silicon that occur at critical junctions in device requirements (eg. modulator and laser). Such solutions must not render the device processing incompatible with CMOS, for then the "silicon advantage" is lost. Achieving reliable and efficient electroluminescence in silicon remains the most intractable of these problems to date. Reliability problems in recently developed light emitting devices operating near a wavelength of 1.54 f..Lm, based on the thermally induced formation of silicon nano-clusters in erbium-doped silicon rich silicon oxide thin films, has reinforced the need for a further understanding of the luminescence mechanisms in this material. Indeed, the efficient and stable sensitized photoluminescence from Er3+ ions (near the telecom wavelength), embedded in an oxide matrix, based on a quasi-resonant energy transfer from nanostructured silicon, has the potential to make possible compact waveguide amplifiers and thin film electroluminescence. This thesis represents a study into the luminescence mechanisms in erbium-doped silicon oxide (SiOx, x~2) thin films grown by electron cyclotron resonance plasma enhanced chemical vapour deposition. Importantly, the film growth relies on in-situ erbium doping through the cracking of a volatile organalanthanide Er(tmhd)3 source. Rutherford backscattering spectroscopy has been used to map the film composition space generated from an ECR-PECVD parameter subspace consisting of precursor gas flow rates and the erbium precursor temperature. The response of the film photoluminescence spectra in both visible and infrared bands consistenly reveals three classes of luminescence centres, whose relative ability to emit light is shown in this study to exhibit a considerable degree of variability through the control of the film composition, subsequent thermal anneal temperature, duration, and process ambient. These three classes consist of optically active Er3 + ions, silicon nano-clusters phase separated during thermal annealing, and oxide-based defects (which may additionally include organic chromophores). The latter two of these species show the ability to sensitize the Er3 + luminescence. In fact, sensitization by intrinsically luminescent defects is a rarely studied phenomenon, which seems to be an important phenomenon in the present films owing to a potentially unique Er incorporation complex. To further investigate the ability of the oxide defects in this regard, an optimally luminescent film has been subject to a damaging ion irradiation to induce a photoluminescence quenching. The subsequent recovery of this luminescence with stepwise isochronous annealing has been correlated with Doppler broadening positron annihilation spectroscopy measurements made with a slow positron beam. Irradiation to a sufficiently high fluence has demonstrated a unique ability to de-couple luminescent sensitizers and Er3+ ions, producing enhanced blue and violet emissions. / Thesis / Master of Science (MS)

Quorum sensing is a bacterial signaling system that controls gene expression in a population density-dependent manner. In Gram-negative proteobacteria, the cell density control of luminescence was first observed in the symbiotic marine bacterium Vibrio fischeri and this system is one of the best studied quorum sensing systems. Two-dimensional sodium dodecyl sulfate-polyacrylamide (2D-SDS) gel electrophoresis analysis previously identified several non-Lux proteins in V. fischeri MJ-100 whose expression was dependent on LuxR and 3-oxo-hexanoyl-L-homoserine lactone (3-oxo-C6-HSL). A lacZ reporter was used to show that the promoters for qsrP, acfA, and ribB were directly activated via LuxR-3-oxo-C6-HSL in recombinant Escherichia coli. The sites of transcription initiation were established via primer extension analysis. Based on the position of the lux box-binding site near position â 40, all three promoters appear to have a class II-type promoter structure. Real-time reverse transcription-PCR was used to study the temporal expression of qsrP, acfA, and ribB during the exponential and stationary phases of growth, and electrophoretic mobility shift assays were used to compare the binding affinities of LuxR to the promoters under investigation. In order to fully characterize the LuxR regulon in V. fischeri ES114, microarray analysis was performed in the Greenberg lab (University of Washington) and 18 LuxR-3-oxo-C6-HSL regulated promoters were found including 2 genes (qsrP and acfA) identified previously in MJ-100 in addition to the well-studied lux operon. In collaboration with them, full-length purified LuxR protein was used to show direct interaction between the LuxR protein and 7 genes/operons newly identified out of 13 genes/operons examined. The binding affinity between LuxR proteins and those genes was also measured. Based on the sequence of the lux boxes of the known genes regulated by LuxR and LitR, a position specific weight matrix (PSWM) was created and used to search through the intergenic regions of the V. fischeri ES114 genome. Some potential LuxR and LitR-regulated genes with high score were tested experimently to confirm direct activation. For the LuxR regulon, these possible LuxR-regulated promoters were cloned into a lacZ reporter and tested for their LuxR dependence. Beyond the genes found in microarray, the promoter of the intergenic region VFA0658-0659 was found to be activated by LuxR and 3-oxo-C6-HSL. For the LitR regulon, two LitR-regulated genes found in the microarray were also identified using PSWM and confirmed by real-time PCR to be dependent on LitR for expression. EMSA experiments showed that LitR can specifically bind to the litR boxes of LitR-regulated genes, litR and VF0170 which confirmed that the regulation is direct. / Ph. D.

A study was conducted to investigate the effects of color CRT misconvergence and display luminance on reading performance and perceived image quality. The objectives of this study were (1) to determine the effects of color CRT misconvergence on reading time and errors, (2) to determine the effects of color CRT misconvergence on subjective ratings of image quality, and (3) to examine the interaction between color CRT misconvergence and display luminance upon reading performance and subjective ratings of image quality. Ten participants performed a Tinker Speed of Reading Test on a shadow-mask, color CRT computer workstation. Both search time to find the out-of-context word and errors were recorded. Following the reading task, each participant rated the image quality of the display using a 9-point subjective scale. Reading performance was not affected by CRT misconvergence or character luminance. Increasing misconvergence did result in lower image quality ratings; however, it was found that participants perceived the more luminous displays to have higher image quality. Also, it was determined that image quality ratings were lowest for magenta misconvergence. However, using normalized ratings, it was found that image quality ratings decreased quickest for the yellow characters with increasing misconvergence. These findings indicate that lower levels of misconvergence (1 to 2.5 arcmin) may not affect reading performance adversely. However, since individuals perceive display quality to be poor at these levels of misconvergence, color CRTs should be used only when application absolutely requires them. / Master of Science

The quorum sensing signaling system based on intercellular exchange of N-acyl-homoserine lactones is used by many proteobacteria to regulate the transcription of essential genes in a signal density-dependent manner. It is involved in a number of processes including the development of highly organized bacterial communities, e.g., biofilms, the regulation of expression of virulence factors, production of antibiotics, and bioluminescence. The extensive genetic and biochemical data available on the quorum sensing system in Vibrio fischeri allows the development of a systems biology approach to undertake a spatial and dynamical analysis of the regulation throughout the population. The quorum sensing regulated lux genes are organized in two divergent transcriptional units: luxR and luxICDABEG. The latter contains the genes required for luminescence and the luxI gene necessary for synthesis of an N-acyl-homoserine lactone commonly called autoinducer (AI). The luxR gene codes for a transcriptional regulatory protein that activates the transcription of both operons at a threshold concentration of AI. The positive feedback loop induces a rapid increase of transcription level of the lux genes when a critical population density is reached (reflected by the concentration of AI in the environment). With a combination of molecular biology tools, physiological analysis, and mathematical modeling we identified critical characteristics of the system and expect to assign parameter values in order to achieve a comprehensive understanding of the dynamics. An ordinary differential equation mathematical model is used to investigate the dynamics of the system and derive parameter values. In parallel a novel microfluidic cell culture experimental set-up is used to carefully control environmental parameters as well as to achieve chemostatic conditions for high-density cell populations. An unstable variant of the green fluorescent protein was used as a reporter to follow the time response at a single cell level. Thus spatial organization and noise across the population can be analyzed. Plasmids carrying different genetic constructs were transformed in a recombinant Escherichia coli strain to specifically identify genetic and biochemical elements involved in the regulation of the lux genes under diverse conditions. Then the quantitative data extracted from batch culture and microfluidic assays were used to assign parameter values in the models. The particular question being investigated first is the nature of the regulation to increasing concentration of the signal. The hypothesis tested is that the regulation of the production of the signal by individual cells is biphasic and, therefore, quorum sensing should be robust to global and local variations in cell density. / Master of Science

X-ray Luminescent Computed Tomography (XLCT) is a new imaging modality which is under extensive trials at present. The modality works by selective excitation of X-ray sensitive nanophosphors and detecting the optical signal thus generated. This system can be used towards recreating high quality tomographic slices even with low X-ray dose. There have been many studies which have reported successful validation of the underlying philosophy. However, there is still lack of information about optimal settings or combination of imaging parameters, which could yield best outputs. Research groups participating in this area have reported results on basis of dose, signal to noise ratio or resolution only. In this thesis, the candidate has evaluated XLCT taking into consideration noise and resolution in terms of composite indices. Simulations have been performed for various beam widths and noise & resolution metrics deduced. This information has been used in evaluating quality of images on basis of CT Figure of Merit & a modified Wang-Bovik Image Quality index. Simulations indicate the presence of an optimal setting which can be set prior to extensive scans. The conducted study, although focusing on a particular implementation, hopes to establish a paradigm in finding best settings for any XLCT system. Scanning with an optimal setting preconfigured can help in vastly reducing the cost and risks involved with this imaging modality. / Master of Science

京都大学 / 新制・課程博士 / 博士(人間・環境学) / 甲第23975号 / 人博第1027号 / 新制||人||242(附属図書館) / 2022||人博||1027(吉田南総合図書館) / 京都大学大学院人間・環境学研究科相関環境学専攻 / (主査)教授 田部 勢津久, 教授 吉田 寿雄, 教授 中村 敏浩, 教授 田中 勝久 / 学位規則第4条第1項該当 / Doctor of Human and Environmental Studies / Kyoto University / DFAM

This study investigates the photoluminescence (PL), cathodoluminescence (CL), radioluminescence (RL) and ionoluminescence (IL) of natural and synthetic minerals. The natural minerals (fluorapatite, leucophanite, meliphanite and zircon) are mostly from Ilímaussaq Alkaline Complex in South Greenland, Langesundsfjord in Norway and from different localities within Scotland. Synthetic fluorapatite (manufactured as part of the present study) and zircon doped with rare earth elements (REE) were used to compare single and multidoped materials. This study has shown that many of the generally accepted applications of luminescence are not as straightforward as often suggested by the current literature. For example, the study demonstrates how site distribution of REE, based on luminescence, is greatly affected by the dopant level and structural changes, and that different conclusions can be drawn on the same sample depending on method applied. Furthermore, it is clearly demonstrated that using luminescence as a tool for quantitative trace element determination is not going to be a standard technique in the near future if ever. The two main findings supporting this conclusion are the non-linear intensity decrease between different REE activators in the same sample and a large variation between activators at the concentration at which self-quenching starts. In contrast to the general perception that luminescence related to REE is mostly independent of the host, this study has shown a strong interaction between host and REE activators. This conclusion is supported by the change in the activator’s coordination polyhedron observed with single-crystal and powder X-ray diffraction combined with full chemical characterisation. When combining the weak interaction between some REE with the strong host interaction this study has shown the potential for designing new types of colour tuneable and “white light” LEDs based on natural minerals. This study also reveals that zircon doped with Gd³⁺ and Eu³⁺ can potentially have quantum-cutting properties.

Ce travail porte sur l’élaboration et la caractérisation physico-chimique de nanoparticules hybrides à base d’apatites phosphocalciques biomimétiques proches du minéral osseux, en vue d’applications dans le domaine du diagnostic de cancers voire de thérapeutique. Dans cette étude, une formulation colloïdale a été développée en milieu aqueux à partir de sels aisément manipulables et en présence d’un dérivé phospholipidique (2-aminoéthylphosphate, AEP) jouant le rôle d’agent dispersant et permettant de contrôler la taille moyenne des nanoparticules (dans la gamme 30-100 nm). L’effet de paramètres expérimentaux majeurs (pH, concentrations, température) a été déterminé. La complémentarité des données analytiques (analyses chimiques, spectroscopie FTIR, diffraction des rayons X, diffusion de la lumière, MET, mesures de potentiel zêta) nous a permis de proposer un modèle descriptif des nanoparticules colloïdales mettant en jeu la présence de complexes entre Ca2+ et AEP- en surface de nanocristaux d’apatite. La possibilité de conférer des propriétés de luminescence a été démontrée, par substitution d’ions Ca2+ par des ions europium Eu3+, et une durée de vie de luminescence de l’ordre de la milliseconde permet d’envisager l’étude de matériel biologique. Plus ponctuellement, l’adsorption additionnelle d’acide folique a été étudiée, avec pour objectif final le ciblage de cellules cancéreuses. Divers aspects liés à une potentielle utilisation dans le domaine biomédical ont également été abordés, tels que la purification de telles suspensions par dialyse, la possibilité d’une remise en suspension après lyophilisation, l’évaluation de leur cytotoxicité, l’étude de leur potentiel proinflammatoire par interaction avec des macrophages humains, et une étude préliminaire de l’internalisation de ces nanoparticules par des cellules cancéreuses. Ce travail a permis de développer une « preuve de concept » permettant d’envisager l’utilisation future de tels nano-systèmes colloïdaux dans le domaine biomédical, et en particulier en oncologie. / This work deals with the synthesis and physico-chemical characterization of hybrid nanoparticles based on biomimetic calcium phosphate apatites close to bone mineral, in view of applications in the field of cancer diagnosis, or therapeutics. In this study, a colloidal formulation has been developed in aqueous medium, from easily-handled salts and in the presence of a phospholipid moiety (2-aminoethylphosphate, AEP) acting as dispersing agent and allowing the control of the mean nanoparticle size (in the range 30-100 nm). The effect of major experimental parameters (pH, concentrations, temperature) has been determined. Complementary analytical data (chemical analyses, FTIR spectroscopy, XRD, dynamic light scattering, TEM, zeta potential measurements) enabled us to propose a descriptive model for the colloidal nanoparticles, involving the presence of complexes between Ca2+ and AEP- on the surface of apatite nanocrystals. The possibility to confer luminescence properties was demonstrated by way of ionic substitutions of some Ca2+ ions by europium Eu3+ ions, allowed us to envision the study of biological material. The additional adsorption of folic acid was also addressed, with the final aim to target cancer cell. Other aspects linked to a potential future use of these nano-systems in the biomedical field were also examined, such as the purification of these suspensions by dialysis, the possibility to resuspend the nanoparticles after freeze-drying, the evaluation of their cytotoxicity, the study of the pro-inflammatory potential by following interactions with human macrophages, and a preliminary study of their internalization by cancer cells. This work enabled us to develop a « proof of concept » allowing one to envision the future use of such colloidal nano-systems in the biomedical field, and in particular in oncology.

Durant mon Doctorat de Sciences Chimiques, effectué en cotutelle entre l'Université de Bologne et l'Université de Aix-Marseille, j'ai pris en charge la synthèse et la caractérisation d'une série de composés aromatiques soufrés qui bénéficient de propriétés photophysiques intéressantes. Ce travail de thèse a été concentré sur deux classes de composés : la première s'apparente à des astérisques moléculaires contenant un coeur benzénique persoufré, fonctionnalisé avec des unités périphériques aromatiques. Dans chaque cas, des groupes stériquement encombrants ont été greffés sur les unités aromatiques situées à la périphérie des molécules. Ces molecules offrent la capacité de fonctionner comme luminophore AIE (Emission Induite par l'Agrégation) à l'état solide ou dans des milieux à haute rigidité. La deuxième famille de molécule est basée sur la même géométrie en astérisque. Cependant, chaque ligand de cette série utilise des unités terpyridyles comme unité périphérique, capables d'interagir avec des métaux de transition comme le fer(III) et le zinc. Suite à la complexation du zinc les propriétés photophysiques peuvent être modulées en fonction de la rigidité du système. Enfin, on a synthetisé un troisième type de système qui a été obtenu par le remplacement du coeur benzénique des astérisques par une fonction pyrene polysoufrée, en conservant les mêmes unités périphériques. Ce composé a montré des propriétés photophysiques intéressantes mais aussi des capacités de coordinations vers les ions métalliques (Fe(III), Zn(II) et le Nd(III)), ce qui a permis la génération d'émission dans le domaine du proche infrarouge. / The aim of this thesis was the synthesis and photophysical characterization of some new polysulfurated aromatic compounds: this class of molecules can offer intriguing properties, potentially useful for the construction of new materials for optoelectronic devices. Two main families of compounds have been synthesized: the first is represented by a series of small molecular asterisks, with peripheral aromatic units, showing luminescence in solid phase or in highly rigid conditions. All compounds with peripheral substituents display an AIE behavior (Aggregation Induced Emission) with radiative deactivation of the triplet states. Taking inspiration from these smaller asterisks, a larger molecule with the same geometry has been designed, decorated with terpyridyl moieties as the outermost units: this compound shows great affinity for the coordination of several transition metal ions, changing luminescence properties after the interaction with zinc ions. With the same intentions, a tetrasulfurated pyrene-core molecule with terpyridyl external units has been synthesized and isolated: this ligand exhibits good coordination capabilities towards transition metal ions, giving rise to luminescent nanoaggregates upon addition of zinc(II), characterized by DLS and AFM microscopy. In addition a NIR emission is recorded after coordination of neodymium(III), showing evidence of an intramolecular energy transfer process.