Les métastases sont responsables de 90 % des décès causés par le cancer. Les métastases sont des foyers cancéreux secondaires qui se forment à distance de la tumeur d’origine. Des cellules cancéreuses quittent la tumeur primaire, rejoignent la circulation sanguine puis colonisent des organes voisins par migration à travers l’endothélium vasculaire. Ce phénomène d’adhésion à l’endothélium et de migration à travers l’endothélium appelé l’extravasation est une étape clé du processus métastatique. L’identification des molécules impliquées constitue une priorité dans le but d’élaborer de nouvelles drogues anticancéreuses. Nous avons précédemment montré que la molécule d’adhésion cellulaire InterCellular Adhesion Molecule-1 (ICAM-1) exprimée par les cellules endothéliales, est impliquée dans l’interaction des cellules de cancer de la vessie (BCs) avec l’endothélium. Cependant les ligands d’ICAM-1 n’ont pas été étudiés. Dans cette étude, nous utilisons des tests d'adhésion cellulaire et la microscopie à force atomique (AFM) afin d’identifier les ligands d’ICAM-1 et de mesurer les forces impliquées dans l’interaction ligand-ICAM-1. Nous avons identifié que les protéines MUC1 et CD43 exprimées par les BCs les plus invasives se lient à ICAM-1 en développant des forces d’intensité différente selon le couple considéré. Une analyse détaillée des événements de rupture suggère que CD43 est fortement lié au cytosquelette et que son interaction avec ICAM-1 correspond principalement à des sauts brusques. Au contraire, MUC1 semble être lié faiblement au cytosquelette et ses interactions avec ICAM-1 sont principalement associées à la formation de filaments membranaires ou « tethers ». Les forces mises en jeu lors de la migration des cellules cancéreuses à travers l'endothélium ont été étudiées par microscopie de forces de traction (TFM). Les résultats préliminaires montrent que les tractions exercées par les cellules cancéreuses lors de l’extravasation sont mesurables par TFM. / Cancer metastasis is associated with 90% cancer-associated deaths, when cancer cells escape from the primary tumor and form metastatic colonies in secondary sites. Extravasation is an important step in cancer metastasis, where cancer cells carried in blood, adhere and transmigrate through the endothelium. Therefore identifying the key molecules involved during the adhesion process could enable to develop new anticancer cancer drugs able to inhibit the adhesion of cancer cells to the endothelium. We have previously shown that InterCellular Adhesion Molecule-1 (ICAM-1) expressed by endothelial cells is involved in the interactions of bladder cancer cells (BCs) with the endothelium. However the ICAM-1 ligands have never been investigated. In this study, we combined adhesion assays and Atomic Force Microscopy (AFM) to identify the ligands involved and to quantify the forces relevant in such interactions. We report the expression of MUC1 and CD43 on BCs and demonstrate that these ligands interact with ICAM-1 to mediate cancer cell-endothelial cell adhesion in the case of the more invasive BCs. AFM experiments were performed to quantify the force ranges involved by MUC1 and CD43 during their interaction with ICAM-1. AFM measurements combined with a Gaussian Mixture Model showed distinct force ranges for the interaction of ICAM-1 with MUC1 and ICAM-1 with CD43. Furthermore, a detailed analysis of the rupture events suggests that CD43 is strongly connected to the cytoskeleton and that its interaction with ICAM-1 mainly corresponds to force ramps followed by sudden jumps. On the contrary, MUC1 seems to be weakly connected to the cytoskeleton as its interactions with ICAM-1 are mainly associated with the formation of tethers. The forces involved during the transmigration of cancer cells through the endothelium was investigated using Traction Force Microscopy (TFM). Preliminary results showed that tractions exerted by cancer cells during transmigration can be studied and quantified using TFM.

Resumo: O sistema de microscopia de força atômica se tornou um instrumento popular e útil para medir as forças intermoleculares com resolução atômica que pode ser aplicado em eletrônica, análises biológicas, engenharia de materiais, semicondutores, etc. Este trabalho estuda o comportamento da dinâmica não-linear da ponta da sonda causada pelo tipo da amostra e os modos de funcionamento de um microscópio de força atômica. Utilizando-se de simulações numéricas, busca-se uma solução aproximada, através do método de perturbação de múltiplas escalas e teoria de controle linear ótimo consegue-se um bom entendimento do trabalho feito e explicado a seguir. Este trabalho está dividido em três partes, na primeira apresentou-se o problema, mostrando a necessidade de se controlar o comportamento caótico no sistema a ser estudado. Mostrou-se o funcionamento do microscópio atômico com todas suas variáveis de funcionamento. Foram geradas as equações de movimento e os resultados são obtidos através de integrações numéricas das equações de movimento, obteve-se oscilações regulares e irregulares (caóticos), os quais dependem da escolha dos parâmetros do sistema. Na segunda parte do trabalho, utilizou-se o método das múltiplas escalas, efetuou-se a busca de uma solução analítica aproximada para o movimento estacionário do sistema, que foi obtida através de técnicas de perturbações. Este método foi desenvolvido foi desenvolvido por [10] para controlar estes sistemas / Abstract: The atomic force microscope system has become a popular and useful instrument to measure the intermolecular forces with atomic-resolution that can be applied in electronics, biological analysis, materials, semiconductors etc. This work studies the complex nonlinear dynamic behavior of the probe tip between the sample and cantilever of an atomic force microscope using numeral simulations, method of multiple scales, and optimal linear control. This work concerns of three parts, in the first we will make the presentation of the AFM, showing various models of AFM. In second part, regular and irregular (chaotic) behaviors depend of the physical parameters and can be observed when a numerical integration is performed. When the dynamic system of the AFM becomes a chaotic oscillator a computational and analytical study of the nonlinear dynamic behavior of the AFM oscillator is proposed and it is obtained by perturbations method. The third part is dedicated to the application and performance of the linear feedback control for the suppressing of the chaotic motion of a non ideal system, theses systems are numerically studied. We use the method developed by [10] to control both the non-ideal system. This method seeks to find an optimal linear feedback control where they find - if conditions for the application of linear control in non-linear, ensuring the stability of the problem / Orientador: José Manoel Balthazar / Coorientador: Bento Rodrigues de Pontes / Banca: Átila Madureira Bueno / Banca: Angelo Marcelo Tusset / Mestre

Ce travail porte sur l'étude du comportement rhéologique de fluide en milieu confiné. Pour cela le levier d'un microscope à force atomique (AFM) est utilisé pour sonder les propriétés rhéologiques d'un fluide confiné entre deux surfaces : la surface d'une sphère collée à l'extrémité du levier et une surface plane sur lequel le fluide est déposé. Le dispositif expérimental est constitué du système de mesure d'un AFM et d'un piézoélectrique permettant d'approcher ou d'éloigner de la sphère la surface plane. Un modèle analytique permet d'extraire les propriétés rhéologiques du fluide confiné à partir de la déflexion du levier induite par le pincement du fluide. Cette méthode a été validée pour les fluides newtoniens. Par contre pour les fluides non-newtoniens comme par exemple la solution de polyacrylami de nous avons trouvé que la viscosité dépend de la distance D et que le cisaillement n'est pas le seul paramètre pertinent pour interpréter les propriétés rhéologiques. / This thesis focuses on the study of the rheological behavior of confined fluids. For this purpose, the microcantilever of an atomic force microscope (AFM) is used to probe the rheological properties of a fluid confined between two surfaces, the surface of a sphere glued to the free-end of the AFM microcantilever and a flat solid surface on which the fluid is deposited. The set-up consists of an AFM, an electrical system for the deflection measurement and a piezoelectric device to move the solid surface (approach, oscillation, etc.). An analytical model allows to determine the rheological properties of the confined fluid from the measurement of the microcantilever deflection due to the hydrodynamic force exerted by the fluid on the sphere.This method has been validated for Newtonian fluids. For non-Newtonian fluids, such as polyacrylamide solution, we found that the viscosity depends on the distance D between the sphere and the plane surface and the shear rate is not the only relevant parameter for interpreting the rheological properties.

O estudo das não linearidades das séries temporais da microviga de um microscópio de força atômica tem sido essencial para o desenvolvimento e aperfeiçoamento deste equipamento de ampliação de imagens. Não linearidades podem aparecer com frequência nos experimentos, afetando significativamente a resposta prevista de forma que estas instabilidades se concretizam em imagens ruins. Esta tese apresenta resultados obtidos através de uma abordagem experimental e teórica. Buscou-se aperfeiçoar modelos clássicos de osciladores do microscópio de força atômica melhorando seu comportamento caótico através da observação dos resultados dos experimentos. A identificação de sistemas é feita pelo método de espaço de estados. Outra abordagem de séries temporais obtidas através de um microscópio de força atômica torna possível a reconstrução de espaço de estados, utilizando-se de técnicas como informação mútua, falsos vizinhos e defasagem de tempo. Analisa-se também o comportamento caótico das séries temporais usando o teste 0-1 e escala indexada em quatro experimentos que resultam em um mapa que relaciona a altura que a microviga vibra com o coeficiente do teste 0-1 e com a escala indexada. / The study of time series nonlinearities of the cantilever\'s atomic force microscope has been essential to the development and improvement of this image magnification equipment. Nonlinearities may appear frequently in the experiments, significantly affecting expected response in a way that these instabilities generate bad images. This thesis presents results obtained through an experimental and theoretical approach accordingly. We attempted to improve the classical models of oscillators atomic force microscope improving its chaotic behavior by observing the results of the experiments. The system identification is made by method space state. Another approach to time series obtained through an atomic force microscope makes it possible to reconstruct space phase, using techniques such as mutual information and false neighbors delay. It is analyzed chaotic behavior time series by using the 0-1 test and scale index in four experiments resulting in a map that relates the height of cantilever deflections with the 0-1 test coefficient and the indexed scale.

Avec l'intégration de circuits intégrés de plus en plus denses, le besoin d'outils de caractérisation adaptés à ces échelles se fait ressentir. Identifier et analyser les problèmes de fiabilité survenant dans ces structures à des dimensions inférieures à 100 nm demande la mise au point d'instruments innovants. Ce travail de thèse a consisté dans un premier temps à développer un appareil à champs proches sensible aux propriétés mécaniques de surface, et dans un second temps à analyser les résultats expérimentaux en s'appuyant sur des approches analytiques et/ou numériques. Désigné sous le nom de microscope à force atomique à résonance de contact (CR-AFM), cet appareil est sensible à la rigidité effective de films minces sur substrat, ce qui lui permet de cartographier la rigidité mécanique de films minces. Nous avons mené un important travail de développement instrumental afin d'obtenir des résultats expérimentaux répétables et fiables, condition indispensable à une analyse quantitative. Puis nous avons utilisé le CR-AFM sur divers échantillons : empilements modèles (films de silice sur silicium, avec épaisseurs variables de silice), films de silice avec porosité variable, structures damascènes d'interconnexion cuivre,… Des images traduisant les variations d'élasticité de surface ont ainsi pu être construites. Pour quantifier ces variations, nous avons analysé nos résultats à l'aide de différents modèles (approches analytiques et numériques). Des simulations par éléments finis ont été réalisées pour étayer ces résultats. / The reduction of feature size in integrated circuits has raised an increasing need for characterization tools displaying small-scale resolution. Reliability issues taking place in these structures with dimensions below 100 nm require the development of innovative instruments. This thesis has first focused on the development of near field apparatus displaying sensitivity to surface mechanical properties. Afterwards analytical and numerical modelings have been developed to analyze the obtained experimental data. Known as contact resonance atomic force microscope (CR-AFM), this apparatus is sensitive to the effective stiffness of thin film on substrate, allowing the mapping of the mechanical stiffness. A significant work on the apparatus setting-up and procedure has been done to obtain repeatable and reliable experimental data, which is a prerequisite for quantitative analysis. Then CR-AFM experiments have been done on various samples: model stacks (silica thin films on silicon with varying silica thickness), silica films with tuned porosity, Damascene copper interconnect structures,… The mapping of elastic stiffness of such samples has been built-up. In order to quantify these contrasts, our experimental results have been analyzed through different models (analytical and numerical). Finite element simulations were also performed to support these results.

Made available in DSpace on 2014-06-11T19:28:33Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-11-25Bitstream added on 2014-06-13T19:37:14Z : No. of bitstreams: 1 nozaki_r_me_bauru.pdf: 1471745 bytes, checksum: 002a3fa9cff00fe97414905826f120e6 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O sistema de microscopia de força atômica se tornou um instrumento popular e útil para medir as forças intermoleculares com resolução atômica que pode ser aplicado em eletrônica, análises biológicas, engenharia de materiais, semicondutores, etc. Este trabalho estuda o comportamento da dinâmica não-linear da ponta da sonda causada pelo tipo da amostra e os modos de funcionamento de um microscópio de força atômica. Utilizando-se de simulações numéricas, busca-se uma solução aproximada, através do método de perturbação de múltiplas escalas e teoria de controle linear ótimo consegue-se um bom entendimento do trabalho feito e explicado a seguir. Este trabalho está dividido em três partes, na primeira apresentou-se o problema, mostrando a necessidade de se controlar o comportamento caótico no sistema a ser estudado. Mostrou-se o funcionamento do microscópio atômico com todas suas variáveis de funcionamento. Foram geradas as equações de movimento e os resultados são obtidos através de integrações numéricas das equações de movimento, obteve-se oscilações regulares e irregulares (caóticos), os quais dependem da escolha dos parâmetros do sistema. Na segunda parte do trabalho, utilizou-se o método das múltiplas escalas, efetuou-se a busca de uma solução analítica aproximada para o movimento estacionário do sistema, que foi obtida através de técnicas de perturbações. Este método foi desenvolvido foi desenvolvido por [10] para controlar estes sistemas / The atomic force microscope system has become a popular and useful instrument to measure the intermolecular forces with atomic-resolution that can be applied in electronics, biological analysis, materials, semiconductors etc. This work studies the complex nonlinear dynamic behavior of the probe tip between the sample and cantilever of an atomic force microscope using numeral simulations, method of multiple scales, and optimal linear control. This work concerns of three parts, in the first we will make the presentation of the AFM, showing various models of AFM. In second part, regular and irregular (chaotic) behaviors depend of the physical parameters and can be observed when a numerical integration is performed. When the dynamic system of the AFM becomes a chaotic oscillator a computational and analytical study of the nonlinear dynamic behavior of the AFM oscillator is proposed and it is obtained by perturbations method. The third part is dedicated to the application and performance of the linear feedback control for the suppressing of the chaotic motion of a non ideal system, theses systems are numerically studied. We use the method developed by [10] to control both the non-ideal system. This method seeks to find an optimal linear feedback control where they find - if conditions for the application of linear control in non-linear, ensuring the stability of the problem

A Restauração de Imagens é uma técnica que possui aplicações em várias áreas, por exemplo, medicina, biologia, eletrônica, e outras, onde um dos objetivos da restauração de imagens é melhorar o aspecto final de imagens de amostras que por algum motivo apresentam imperfeições ou borramentos. As imagens obtidas pelo Microscópio de Força Atômica apresentam borramentos causados pela interação de forças entre a ponteira do microscópio e a amostra em estudo. Além disso apresentam ruídos aditivos causados pelo ambiente. Neste trabalho é proposta uma forma de paralelização em GPU de um algoritmo de natureza serial que tem por fim a Restauração de Imagens de Microscopia de Força Atômica baseado na Regularização de Tikhonov. / Image Restoration is a technique which has applications in several areas, e.g., medicine, biology, electronics, and others, where one of the goals is to improve the final appearance of the images of samples, that have for some reason, imperfections or blurring. The images obtained by Atomic Force Microscope have blurring caused by the interaction forces between the tip of the microscope and the sample under study. Moreover exhibit additive noise caused by the environment. This thesis proposes a way to make a parallelization on a GPU of a serial algorithm of which is a Image Restoration of Images from Atomic Force Microscopy using Tikhonov Regularization.

O tratamento clareador pode resultar em alterações na rugosidade superficial do esmalte dental, as quais podem ser vistas a olho nu como alterações de cor, opacidade e perda de brilho. O objetivo deste estudo foi analisar in vitro a alteração de rugosidade, cor e brilho de superfícies de esmalte dental humano após clareamento com um produto a base de peróxido de hidrogênio (PH) 38% e após polimento com fluoreto de sódio 2% neutro (FS2%) ou pasta dentifrícia contendo partículas de nano-hidroxiapatita. Para isso, coroas de incisivos centrais humanos foram preparadas e tiveram suas faces vestibulares analisadas em microscópio de força atômica (MFA), o qual determinou os valores de rugosidade [Ra, RMS, Z range e Power Spectral Density (PSD)] iniciais. A cor e o brilho destas amostras foram determinados por um espectrofotômetro. Todas as amostras foram clareadas com PH 38% por 135 minutos. Os valores de Ra, RMS, Z range e PSD foram estabelecidos pelo MFA e a cor e o brilho determinados pelo espectrofotômetro. As amostras foram divididas em dois grupos: Flúor, que recebeu um polimento com gel de FS2%; e nHA, que recebeu um polimento com pasta dentifrícia contendo partículas de nano-hidroxiapatita. Os valores de Ra, RMS, Z range, PSD, cor e brilho pós-polimento foram determinados pelo MFA e espectrofotômetro. Após o clareamento, os valores de Ra, RMS, Z range e PSD não apresentaram alteração estatisticamente significante, os valores de L* aumentaram e C* diminuiu significativamente em todas as amostras. O brilho superficial não apresentou alteração significativa após o clareamento. O grupo Flúor não apresentou alteração de rugosidade significativa para Ra, RMS, Z range e PSD após o polimento. Os valores de L*, C* e brilho superficial também não apresentaram alterações significativas após o polimento das superfícies clareadas. No grupo nHA, os valores de Ra e RMS não apresentaram alteração significativa após o polimento das superfícies clareadas, porém os valores de Z range e PSD apresentaram alteração estatisticamente significante. Em comparação à superfície clareada, L* e C* não apresentaram alteração significante após o polimento, porém o brilho superficial foi significativamente maior. O tratamento clareador de consultório mostrou ser eficiente quanto à alteração de cor e não gerou aumento na rugosidade superficial do esmalte dental. Um polimento com pasta dentifrícia contendo partículas de nano-hidroxiapatita realizado após o clareamento é capaz de aumentar o brilho e diminuir a rugosidade superficial do esmalte dental. / Bleaching treatment may result in dental enamel surface roughness alterations, which might be seen as color alterations, opacity and gloss loss. The aim of this research was to in vitro investigate roughness, color and gloss alterations of human dental enamel surfaces after bleaching treatment with 38% hydrogen peroxide (HP) and after polishing with neutral 2% sodium fluoride or dental tooth paste containing nano-hydroxiapatite particles. Human central incisors were prepared and buccal surfaces were analyzed by an atomic force microscope (AFM), which determined roughness values [Ra, RMS, Z range and Power Spectral Density (PSD)]. Surface color and gloss were determined by a spectrophotometer. All samples were bleached with 38% HP for 135 minutes. AFM determined Ra, RMS, Z range and PSD after bleaching. Color and gloss were determined by the spectrophotometer. Samples were distributed in two different groups: Fluor, which was polished with neutral 2% sodium fluoride, and nHA, which was polished with dental tooth paste containing nano-hydroxyapatite particles. Ra, RMS, Z range, PSD, color and gloss values were determined by AFM and spectrophotometer after polishing. After bleaching, Ra, RMS, Z range and PSD didnt show any statistical difference. L* increased and C* decreased significantly in all samples after bleaching. Surface gloss didnt show significant alteration after bleaching. Fluor group didnt show surface roughness alteration considering Ra, RMS, Z range and PSD after polishing. L*, C* and gloss didnt change after bleached surfaces are polished with neutral 2% sodium fluoride. Ra and RMS of nHA group didnt change significantly after polishing, but Z range and PSD showed significant alteration. Despite L* and C* didnt change, gloss increased significantly after polishing with dental tooth paste containing nano-hydroxyapatite. In-office bleaching treatment showed to be efficient as for color enhancement and didnt cause roughness alteration in dental enamel surface. An after-bleaching polishing with dental tooth paste containing nano-hydroxyapatite particles may increase gloss and decrease surface roughness of dental enamel.

[pt] Nesta dissertação de mestrado, nós desenvolvemos um processo de litografia baseado na técnica de microscopia de força atômica. O estudo do processo de litografia aqui utilizado inicia-se com a deposição e caracterização de filmes finos de sulfeto de arsênio amorfo (a-As2S3) em substratos de silício e a deposição de uma camada metálica de alumínio, utilizada como máscara, sobre a superfície do a-As2S3. O microscópio de força atômica é utilizado para escrever os padrões de forma controlada na camada metálica, e para tal, a influencia dos parâmetros de controle do microscópio na realização da litografia foi analisada. Para a transferência do padrão litografado realiza-se um posterior processo de fotossensibilização e dissolução química do a-As2S3 com uma solução de K2CO3. Após a dissolução, uma camada de ouro foi depositada por erosão catódica DC, seguido de uma nova dissolução, desta vez com NaOH resultando na transferência de nanoestruturas de Au para o substrato de silício. / [en] In this dissertation, we have developed a lithography process based on the atomic force microscopy of technique. The study of the lithography process starts with the deposition and characterization of amorphous arsenic sulfide thin films (a-As2S3) in silicon substrates and the deposition of a metallic aluminum layer, used as mask, on the surface of the a-As2S3. An atomic force microscope was used to write patterns in a controlled way on the metallic layer. Therefore, the influence of microscope feedback system on the accomplishment of the lithography was analyzed. In order to transfer the lithographed pattern to a silicon substrate, the a- As2S3 was exposed to a UV light source and was dissolved with a K2CO3 solution. Then, a thin gold layer was deposited by sputtering DC, and a new dissolution, now with NaOH was performed, leading to the deposition of Au nanostructures onto the silicon substrate.

Because cells are machines, their structure determines their function (health). But their structure also determines cells’ mechanical properties. So if we can understand how cells’ mechanical properties are influenced by specific structures, then we can observe what’s happening inside of cells via mechanical measurements. The Atomic Force Microscope (AFM) has become a standard tool for investigating the mechanical properties of cells. In many experiments, an AFM is used to ‘poke’ adherent cells with nanonewton forces, and the resulting deformation observed via, e.g. Laser Scanning Confocal Microscopy. Results of such experiments are often interpreted in terms of continuum mechanical models which characterize the cell as a linear viscoelastic solid. This “top-down” approach of poking an intact cell —complete with cytoskeleton, organelles etc.— can be problematic when trying to measure the mechanical properties and response of a single cell component. Moreover, how are we to know the sensitivity of the cell’s mechanical properties to partial modification of a single component (e.g. reducing the degree of cross- linking in the actin cortex)? In contrast, the approach taken here —studying the deformation and relaxation of lipid bilayer vesicles— might be called a “bottom-up” approach to cell mechanics. Using Coarse- Grained Molecular Dynamics simulations, we study the deformation and relaxation of bilayer vesicles, when poked with constant force. The relaxation time, equilibrium area expansion, and surface tension of the vesicle membrane are studied over a range of applied forces. Interestingly, the relaxation time exhibits a strong force-dependence. Force-compression curves for our simulated vesicle show a strong similarity to recent experiments where giant unilamellar vesicles were compressed in a manner nearly identical to that of our simulations.