La maurocalcine (MCa) est une toxine de 33 acides aminés initialement issue du venin du scorpionScorpio maurus palmatus, et est considérée comme faisant partie de la famille des CPP(Cell Penetrating Peptides) depuis de nombreuses années déjà. La MCa présente donc un intérêtthérapeutique certain dans le domaine de la délivrance intracellulaire de cargos, et lestravaux exposés ici cherchent à caractériser au mieux les propriétés de pénétration de la moléculenative ainsi que celle de certains de ses variants.Après avoir quantifié l’internalisation de plusieurs variants tronqués (linéaires), j’ai pu mettreen évidence le fait que tous ces analogues testés ont une capacité à être internalisés bien plusélevée que celle des CPP de référence (notamment Tat et la pénétratine). Parmi ces variants,l’analogue MCaUF1−9 présente l’avantage d’un temps de rétention relativement élevé au seindes cellules, ainsi que d’une accumulation légèrement accrue en environnement acide (ce quiadvient lors de la formation tumeurs solides). Ce nouveau CPP possède donc un certain potentielthérapeutique mais l’étude de la MCa native, remarquablement stable in vivo, reste plusque jamais d’actualité. / Maurocalcine (MCa) is a 33-mer toxin originally isolated from the venom of the scorpioScorpio maurus palmatus, and has been considered as a cell-penetrating peptide (CPP) for severalyears. MCa presents a therapeutic interest for the intracellular delivery of cargoes, andthis thesis aims to characterise the cell penetration properties of the native molecule as well assome of its variants’.After quantifying several truncated (linear) variants’ internalisation, I have been able tohighlight the fact that all of those analogs possess a higher internalization ability than those ofstandard CPP (especially Tat and penetratin). Among those variants, the analog MCaUF1−9 hasa relatively high rentention time within cells, as well as a slightly increased accumulation whenin an acidic environment (which occurs during solid tumours formation). This new CPP showsa certain therapeutic potential but the study of nativeMCa, remarkably stable in vivo, remainsa priority.

abstract: Nuclear magnetic resonance (NMR) is an important phenomenon involving nuclear magnetic moments in magnetic field, which can provide much information about a wide range of materials, including their chemical composition, chemical environments and nuclear spin interactions. The NMR spectrometer has been extensively developed and used in many areas of research. In this thesis, studies in two different areas using NMR are presented. First, a new kind of nanoparticle, Gd(DTPA) intercalated layered double hydroxide (LDH), has been successfully synthesized in the laboratory of Prof. Dey in SEMTE at ASU. In Chapter II, the NMR relaxation studies of two types of LDH (Mg, Al-LDH and Zn, Al-LDH) are presented and the results show that when they are intercalated with Gd(DTPA) they have a higher relaxivity than current commercial magnetic resonance imaging (MRI) contrast agents, such as DTPA in water solution. So this material may be useful as an MRI contrast agent. Several conditions were examined, such as nanoparticle size, pH and intercalation percentage, to determine the optimal relaxivity of this nanoparticle. Further NMR studies and simulations were conducted to provide an explanation for the high relaxivity. Second, fly ash is a kind of cementitious material, which has been of great interest because, when activated by an alkaline solution, it exhibits the capability for replacing ordinary Portland cement as a concrete binder. However, the reaction of activated fly ash is not fully understood. In chapter III, pore structure and NMR studies of activated fly ash using different activators, including NaOH and KOH (4M and 8M) and Na/K silicate, are presented. The pore structure, degree of order and proportion of different components in the reaction product were obtained, which reveal much about the reaction and makeup of the final product. / Dissertation/Thesis / Ph.D. Physics 2013

The task of this thesis is to study methods for the acquisition perfusní imaging based on dynamic MR imaging with T1 contrast. It describes methods of measurement of T1 relaxation time and the possibility of evaluating the results. It further describes the phantoms and their use. And it is here mentioned for the dynamic acquisition protocol perfusní imaging. There is also described in detail created a program for automatic control of the NMR system. In the experimental measurements are performed on static and dynamic phantom, are also evaluated perfusion parameters from the Flash sequence.

Contrast agents are often crucial in medical imaging for disease diagnosis. Novel contrast agents, such as gold nanoparticles (AuNPs) and lanthanides, are being ex- plored for a variety of clinical applications. Preclinical testing of these contrast agents is necessary before being approved for use in humans, which requires the use of small animal imaging techniques. Small animal imaging demands the detection of these contrast agents in trace amounts at acceptable imaging time and radiation dose. Two such imaging techniques include x-ray fluorescence computed tomography (XFCT) and photon-counting CT (PCCT). XFCT combines the principles of CT with x-ray fluorescence by detecting fluorescent x-rays from contrast agents at various projections to reconstruct contrast agent maps. XFCT can image trace amounts of AuNPs but is limited to small animal imaging due to fluorescent x-ray attenuation and scatter. PCCT uses photon-counting detectors that separate the CT data into energy bins. This enables contrast agent detection by recognizing the energy dependence of x-ray attenuation in different materials, independent of AuNP depth, and can provide anatomical information that XFCT cannot. To achieve the best of both worlds, we modeled and built a table-top x-ray imaging system capable of simultaneous XFCT and PCCT imaging. We used Monte Carlo simulation software for the following work in XFCT imaging of AuNPs. We simulated XFCT induced by x-ray, electron, and proton beams scanning a small animal-sized object (phantom) containing AuNPs with Monte Carlo techniques. XFCT induced by x-rays resulted in the best image quality of AuNPs, however high-energy electron and medium-energy proton XFCT may be feasible for on-board x-ray fluorescence techniques during radiation therapy. We then simulated a scan of a phantom containing AuNPs on a table-top system to optimize the detector arrangement, size, and data acquisition strategy based on the resulting XFCT image quality and available detector equipment. To enable faster XFCT data acquisition, we separately simulated another AuNP phantom and determined the best collimator geometry for Au fluorescent x-ray detection. We also performed experiments on our table-top x-ray imaging system in the lab. Phantoms containing multiples of three lanthanide contrast agents were scanned on our tabletop x-ray imaging system using a photon-counting detector capable of sustaining high x-ray fluxes that enabled PCCT. We used a novel subtraction algorithm for reconstructing separate contrast agent maps; all lanthanides were distinct at low concentrations including gadolinium and holmium that are close in atomic number. Finally, we performed the first simultaneous XFCT and PCCT scan of a phantom and mice containing both gadolinium and gold based on the optimized parameters from our simulations. This dissertation outlines the development of our tabletop x-ray imaging system and the optimization of the complex parameters necessary to obtain XFCT and PCCT images of multiple contrast agents at biologically-relevant concentrations. / Graduate

Ces travaux de thèse s'inscrivent dans le contexte de l'imagerie moléculaire, qui vise à adapter les traitements de pathologies à la variabilité de chaque patient, grâce à l'imagerie de biomarqueurs cellulaires ou moléculaires. En particulier, l'imagerie par résonance magnétique (IRM) couplée a des nanoparticules d’oxyde de fer innovantes pourrait permettre de relever un tel défi.Cette thèse se concentre sur l'étude d'une nouvelle classe d'agents de contraste à base d'oxyde de fer pour l'IRM à haut champ magnétique. En effet, les magnétosomes sont des vésicules d’oxyde de fer produites naturellement par des bactéries appelées bactéries magnétotactiques. De telles bactéries synthétisent ces vésicules magnétiques et les alignent comme l'aiguille d'une boussole, ce qui facilite leur navigation dans les sédiments. Ces bactéries produisent donc des magnétosomes aux propriétés magnétiques exceptionnelles: 50 nm de diamètre, mono-cristallin, mono-domaine magnétique et avec une haute magnétisation à saturation. De plus, une grande variété de souches bactériennes existent dans la nature, et produisent, avec une grande stabilité, des magnétosomes dont la taille, la forme, et le contenu chimique, sont déterminés génétiquement. Enfin, les magnétosomes sont naturellement porteurs d'une membrane bi-lipidique dont le contenu est également déterminé génétiquement. Récemment, le contenu protéique de la membrane des magnétosomes a été mis à jour, ouvrant la voie à la fonctionnalisation de cette dernière par fusion des gènes codant pour des protéines présentes abondamment à la membrane avec ceux codant pour un peptide d’intérêt.Ainsi, l'utilisation de ces micro-organismes pour produire des agents de contraste innovants et fonctionnalisés pour l'imagerie moléculaire par IRM, et les applications qui en découlent, ont été étudiées pendant cette thèse. La production et l'ingénierie des magnétosomes a été réalisée par nos collègues du Laboratoire de Bioénergétique Cellulaire (LBC, CEA Cadarache), et sera présentée et discutée. Des magnétosomes sauvages ont d'abord été caractérisés en tant qu'agents de contraste pour l'IRM. De tel magnétosomes présentent des propriétés contrastantes très intéressantes pour l'IRM, ce qui a été validé à la fois in vitro puis in vivo. L'étude de faisabilité de la production d'un agent de contraste pour l'imagerie moléculaire par IRM en une seule étape, à l'aide des bactéries magnétotactiques, a été réalisée sur un modèle de souris porteur de glioblastome. Sachant par la littérature que les cellules tumorales sur-expriment les intégrines anb3, et que ces dernières peuvent être ciblées par le peptide RGD, il a été choisi de produire des magnétosomes exprimant le peptide RGD à leur membrane. L'affinité de tels magnétosomes pour les cellules tumorales U87 a été vérifiée in vitro, et démontré in vivo par IRM puis cross-validé par histologie. / This work takes place in the context of molecular imaging, which aims at tailoring medical treatments and therapies to the individual context by revealing molecular or cellular phenomenon of medical interest in the less invasive manner. In particular, it can be acheived with MRI molecular imaging using engineered iron-oxide contrast agent.This PhD thesis focuses on the study of a new class of iron-oxide contrast agent for high field MRI. Indeed, magnetosomes are natural iron-oxide vesicles produced by magnetotactic bacteria. These bacteria synthesized such magnetic vesicles and ordered them like a nano-compass in order to facilitate their navigation in sediments. This explains why magnetosomes are awarded with tremendous magnetic properties: around 50 nm, mono-crystalline, single magnetic domain and high saturation magnetization. Furthermore, a wide variety of bacterial strains exist in nature and size and shape of magnetosomes are highly stable within strain and can be very different between strains. Finally, magnetosomes are naturally coated with a bilipidic membrane whose content is genetically determined. Lately, researchers have unravelled magnetosomes membrane protein contents, opening the way to create functionnalized magnetosomes thanks to fusion of the gene coding for a protein of interest with the gene coding for an abundant protein at magnetosomes membrane.A new alternative path using living organisms to tackle the production of engineered high effciency molecular imaging probes have been investigated with magnetotactic bacteria in this PhD. The production and engineering of magnetosomes have been carried out by our partner, the Laboratoire de Bio-energétique Cellulaire (LBC, CEA Cadarache), and will be presented and discussed. We then characterized magnetosomes as contrast agent for high field MRI. We showed they present very promising contrasting properties in vitro, and assessed this observation in vivo by establishing they can be used as effcient blood pool agent after intravenous injection. Afterward, we applied the concept of producing engineered MRI molecular imaging probes in a single step by bacteria, to a mouse model of glioblastoma. Knowing that tumor cells can be actively targeted through anb3 integrins by RGD, we produced RGD functionnalized magnetosomes. We started from showing these RGD magnetosomes have a good affnity for U87 cell in vitro, prior to demonstrate it in vivo on orthotopic U87 mouse model. This in vivo affnity being fnally cross-validated with histology.

The use of ultrasound contrast agents increases the information available for reconstruction during ultrasound imaging. Previously studied microbubbles, consisting of a gas core, are subject to limitations such as a short lifetime and a large size. Droplets with a liquid perfluorocarbon core that is stabilized by cellulose nanofibers eliminate these drawbacks, but require further investigation. By studying the frequency-dependent attenuation profile of the cellulose nanofiber coated perfluorocarbon droplets within an ultrasound field, information about the droplets as oscillators can be retrieved, enabling characterization of their physical properties. In this study, the frequency-dependent attenuation profile was experimentally acquired and compared between two concentrations, using flat transducers covering the frequency range of 1-15 MHz. The data collected in the time domain was processed and transformed into the frequency domain and the attenuation was calculated across the entire frequency range. Among the frequencies studied, the attenuation increases with frequency and covers the range of approximately 0.25-8.3 dB/cm and 0.01-3.3 dB/cm at the concentrations of 50 million droplets/ml and 10 million droplets/ml respectively. The attenuation reaches a minimum at 3 MHz within the highest concentration, compared to 4.43 MHz within the lowest. The increase of the attenuation with frequency is explained by the droplets not exhibiting large oscillations within the range covered. It is probable that the droplets do exhibit oscillations, due to a viscosity lower than that of water, but a resonance frequency is not found within the spectrum studied. This could be explained by a shell elasticity or a small droplet radius placing the resonance frequency outside of the spectrum studied, or high levels of damping broadening the resonance peak. Localizing the resonance frequency would enable characterization of these physical properties of the cellulose nanofiber shell as well as the perfluorocarbon liquid core of the droplets. The increase of the attenuation with frequency demonstrates that the droplets do not produce a maximized amount of scattering at a specific frequency within the range studied, which is observed among other oscillating particles implemented as ultrasound contrast agents. The attenuation is, however, larger than that of blood across all frequencies except for those among which the attenuation reaches its minimum. Potential errors that are affecting the results include droplet vaporization, the formation of flocs after the mechanical agitation has ceased, the experimental setup, the settings on the pulse generator, the sensitivity of the transducers and the processing code.

Today, microbubbles are one of the most commonly used ultrasound contrast agents, since their high compressibility results in a strongly scattered signal. Despite this advantage, microbubbles experience limitations by the decreased stability and large diameter. The cellulose nanofiber (CNF) stabilized perfluoropentane (PFC5) droplets have the possibility of eliminating these drawbacks. In order to examine the droplet behavior and scattering ability when exposed to ultrasound, the acoustic response of the droplets is studied and compared with that of microbubbles (MBs). Therefore, this thesis aims to design an experimental set-up and a processing method to determine the phase velocity, bulk modulus and compressibility of the CNF-coated PFC5 droplets. The experimental study of the acoustic characterization uses pulse-echo spectroscopy with an aluminum reflector and seven flat transducers covering the frequency range 0.7 to 14.1 MHz. By using fast Fourier transform, while accounting for the 2πn ambiguity, the phase velocity profiles are obtained. The dispersions within this frequency spectrum are 1391-1487 m/s and 1387-1488 m/s for the concentrations 10 ∙ 106 and 50 ∙ 106 droplets/ml, respectively. These profiles display an increasing phase velocity with frequency and a slight increase in dispersion with concentration. These results agree with theory and studies examining the phase velocity of MBs. The bulk modulus presents values between 3-4 GPa, while the compressibility is 2.7 − 3.2 ∙ 10-10 𝑃𝑎-1 within the frequency range studied. Compared to water and certain MBs, both possessing a lower bulk modulus, the droplets are less compressible. To conclude, the droplets have similar phase velocity profiles with the same dependencies on frequency and concentration as MBs, resulting in similar behavior of these droplets when exposed to ultrasound. Hence, affecting the wave similarly to MBs in terms of spreading. The droplet are, however, not as compressible. This most likely affects their oscillation and they, hence, might not have equally beneficial scattering ability. This could reduce their utilization as contrast agents. Some of the potential error sources present during the laboratory work and the development of the post-processing code were not achieving perfect optimization of the transducer alignment, vaporization of the droplets resulting in reduced concentration, possible diffraction, not optimal processing of data and inadequate correction for 2πn ambiguity.

No description available.

The pursuit of more suitable drugs intended for possible biological applications are a continuously growing topic of research within the scientific community. One of these suitable qualities includes the need for hydrophilicity and or some appropriate delivery system for the drug to enter into biological systems. A system of analyzing and following these compounds would then, however, be necessary to conduct any kind of mechanistic or interaction studies for he said drug within the biological system. Just to name a few, fluorescence and X-ray computed tomography (CT) methods allow for imaging of biological systems but require the need of compounds with specific qualities. Finally, even with a means of entering and following a oaded drug, it would not be complete without a way of targeting its intended location. Herein, the first chapter reports the synthesis and characterization of a fluorene-based pyridil bis-?-diketone compound with suitable one- and two-photon fluorescent properties and its encapsulation into Pluronic F127 micelles for the possible application of tracking lysosomes. Next the synthesis and characterization of a BODIPY-based fluorophore with excellent fluorescence ability is reported. This compound was conjugated to two triphenylphosphine (TPP) groups and is shown as a potential mitochondria probe within HCT-116 cells. Finally, the synthesis and characterization of diatrizoic acid (DA) based derivatives conjugated to silica nanoparticles, as well as unconjugated, are reported as potential CT contrast agents. The derivatives were also functionalized with maleimide moieties facilitating subsequent potential bioconjugation of a targeting protein via a thiol group.

No description available.