Molecular photoacoustic imaging is an exciting new field that promises to visualize molecular indicators of disease. The objective of this dissertation is to progress molecular imaging by providing a photoacoustic microscopy platform to better validate in vivo molecular photoacoustic imaging, diagnose disease, and study fundamental photoacoustic processes. Initially, a custom photoacoustic microscope was developed to provide high-sensitivity and high-resolution of both endogenous and exogenous contrast agents in thin cell or tissue samples. After characterization, the photoacoustc microscope was first used to image the hemoglobin distribution in the spleen and liver. The photoacoustic microscope was then used to image nanoparticles in injured and diseased cell and tissues samples. These images can be used for in vivo photoacoustic image validation or, independently, as a diagnostic tool for disease. To enhance the utility of photoacoustic microscopy, a quantitation technique was developed for nanoparticles in cells and tissues. Quantitative photoacoustic imaging has the potential to replace mass spectrometry and histology for a wide array of molecular imaging and targeting studies. Finally, photoacoustic microscopy was used to study the nonlinear dependence of the photoacoustic pressure with laser fluence of nanoparticle-loaded cells. New discoveries about the nonlinear dependence with nanoparticle concentration and cell type are presented. These new discoveries may provide the framework for a new type of photoacoustic imaging with contrast that is cell-type specific. Overall, the work described in this dissertation can be used to improve diagnosis and accelerate clinical translation of new and emerging molecular imaging techniques. / text

Regenerative medicine is an interdisciplinary field which has advanced with the use of biotechnologies related to biomaterials, growth factors, and stem cells to replace or restore damaged cells, tissues, and organs. Among various therapeutic approaches, cell-based therapy is most challenging and exciting for both scientists and clinicians pursuing regenerative medicine. Specifically, stem cells, including mesenchymal stem cells and adipose-derived stem cells, are promising candidate cell types for cell-based therapy because they can differentiate into multiple cell types for tissue regeneration and stimulate other cells through neovascularization or paracrine signaling. Also, for effective treatment using stem cells, the tissue engineered constructs, such as bioactive degradable scaffolds, that provide the physical and chemical cues to guide their differentiation are incorporated with stem cells before implantation. Also, it was previously demonstrated that tissue-engineered matrices can promote tubulogenesis and differentiation of stem cells to vascular cell phenotypes. Hence, during tissue regeneration after stem cell therapy, there are numerous factors that need to be monitored. As a result, imaging-based stem cell tracking is essential to evaluate the distribution of stem cells as well as to monitor proliferation, differentiation, and interaction with the microenvironment. Therefore, there is a need for a stem cell imaging technique that is not only noninvasive, sensitive, and easy to operate, but also capable of quantitatively assessing stem cell behaviors in the long term with high spatial resolution. Therefore, the overall goal of this research is to demonstrate a novel imaging method capable of continuous in vitro assessment of stem cells as prepared with tissue engineered constructs and noninvasive longitudinal in vivo monitoring of stem cell behaviors and tissue regeneration after stem cell implantation. In order to accomplish this, gold nanoparticles are demonstrated as photoacoustic imaging contrasts to label stem cells. In addition, ultrasound and photoacoustic imaging was utilized to monitor stem cells and neovascularization in the injured rat tissue. Therefore, using these methods, tissue regeneration can be promoted and noninvasively monitored, resulting in a better understanding of the tissue repair mechanisms following tissue injury. / text

Recent clinical studies have demonstrated that photoacoustic imaging (PAI) provides important diagnostic information for breast cancer staging. Despite these promising studies, PAI remains an unfeasible option for clinics due to the cost to implement, the required large modification in user conduct and the inflexibility of the hardware to accommodate other applications for the incremental enhancement in diagnostic information. The research described in this dissertation addresses these issues by designing attachments to clinical ultrasound probes and incorporating custom detectors into commercial ultrasound scanners. The ultimate benefit of these handheld devices is to expand the capability of current ultrasound systems and facilitate the translation of PAI to enhance cancer diagnostics and neurosurgical outcomes. Photoacoustic enabling devices (PEDs) were designed as attachments to two clinical ultrasound probes optimized for breast cancer diagnostics. PAI uses pulsed laser excitation to create transient heating (<1°C) and thermoelastic expansion that is detected as an ultrasonic emission. These ultrasonic emissions are remotely sensed to construct noninvasive images with optical contrast at depths much greater than other optical modalities. The PEDs are feasible in terms of cost, user familiarity and flexibility for various applications. Another possible application for PAI is in assisting neurosurgeons treating aneurysms. Aneurysms are often treated by placing a clip to prevent blood flow into the aneurysm. However, this procedure has risks associated with damaging nearby vessels. One of the developed PEDs demonstrated the feasibility to three-dimensionally image tiny microvasculature (<0.3mm) beyond large blood occlusions (>2.4mm) in a phantom model. The capability to use this during surgery would suggest decreasing the risks associated with these treatments. However, clinical ultrasound arrays are not clinically feasible for microsurgical applications due to their bulky size and linear scanning requirements for 3D. Therefore, capacitive micromachined ultrasound transducer (CMUT) two-dimensional arrays compatible with standard ultrasound scanners were used to generate real-time 3D photoacoustic images. Future probes, designed incorporating CMUT arrays, would be relatively simple to fabricate and a convenient upgrade to existing clinical ultrasound equipment. Eventually, a handheld tool with the ability to visualize, in real-time 3D, the desired microvasculature, would assist surgical procedures. The potential implications of PAI devices compatible with standard ultrasound equipment would be a streamlined cost efficient solution for translating photoacoustics into clinical practice. The practitioner could then explore the benefits of the enhanced contrast adjunctive to current ultrasound applications. Clinical availability of PAI could enhance breast cancer diagnostics and cerebrovascular surgical outcomes.

Made available in DSpace on 2014-06-11T19:25:32Z (GMT). No. of bitstreams: 0 Previous issue date: 2002-12-18Bitstream added on 2014-06-13T19:53:22Z : No. of bitstreams: 1 fonseca_jrl_me_rcla.pdf: 850383 bytes, checksum: bcd0aa3efb5190b07e8211dc3c5cdbca (MD5) / A espectrometria fotoacústica sofreu um grande avanço nos últimos anos, como técnica de análise de materiais e sistemas. Os calorímetros que utilizam o fenômeno fotoacústico se desenvolveram para análises que não podem ser feitas por outros procedimentos, como, a reflectância difusa, reflectância total atenuada e espalhamento Raman. O espectrômetro elaborado nesse trabalho pode ser utilizado para a determinação de energia de laser e calores específicos de materiais biológicos sólidos. Foi confeccionado com materiais de baixo custo e existentes no mercado nacional. Foi testado com sucesso para medidas de energia de laser. / Photoacoustic spectrometry has undergone a significant development in the last years as a technic of material and system analyses. Calorimeters using photoacoustic phenomenon were developed to provide analyses that cannot be done by other procedures such as diffuse reflectance, total atenuated reflectance and Raman’s scattering about. The spectometer constructed in this work can be used to determine laser energy and specific heat of biological solid materials. It was constructed with cheap material easily found in national market. The photoacoustic spectometer was sucessfully tested in measuring laser energy.

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No presente trabalho, realizado com hemoproteínas na forma de pó, o efeito da hidratação foi observado através de espectroscopia fotoacústica. Amostras de carboxi-hemoglobina e carboxi-mioglobina mantidas em diferentes ambientes de umidade relativa (UR), mostraram variações em seus espectros na região da banda de Soret. Para amostras mantidas em baixa hidratação característico do derivado carboxi, em alta hidratação (acima de aproximadamente 90% UR) o espectro era característico do derivado carboxi e na região intermediária o espectro era de uma mistura dos dois derivados. Essa mudança de ligante observada em alta hidratação pode ser explicada supondo que a proteína tem flexibilidade e atinge um estado conformacional que possibilita a entrada e saída do ligante. Em baixa hidratação a estrutura da proteína é rígida e tal que o acesso ao grupo heme está fechado, impossibilitando a troca do ligante. Essa explicação é coerente com vários resultados experimentais que indicam a existência de duas estruturas para essas hemoproteínas em solução / At the present work accomplished with powder of hemoproteins, the hydration effect was observed through photoacoustic spectroscopy. Samples of caroxyhemoglobin and carboxymyglobin kept at different relative humidity (RH) environments, showed variations in their spectra in the Soret bad region. For the samples which were kept at low hydration (Bellow about 33% RH) the spectrum was characteristic of carboxy derivative, whereas at high hydration (above about 93% RH) the spectrum was characteristic of oxy derivative, and in the the intermediate region the spectrum was a mixture of both derivatives. This ligand change observed at high hydration, may be explained assuming that the protein has flexibility, and reach a conformational state which enables the ligand to GO in and out. At low hydrations the protein structure is rigid and such that the access to the heme group is closed becoming impossible the ligand change. This explanation agrees with several experimental results that point to the existence of two structures to these hemoproteins in solution

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Ces travaux de thèse sont motivés par le développement de techniques d’imagerie alternatives pour le diagnostic précoce du cancer du sein. Parmi celles-ci, l’imagerie photoacoustique couple potentiellement les avantages de deux modalités d’imagerie non-invasives, à savoir la quantification de contrastes physiologiques du fait de l’excitation optique et la haute résolution du fait d’un sondage acoustique.Le but de ces travaux est de proposer une modélisation multiondes du phénomène photoacoustique, et d’incorporer ce modèle dans un algorithme de reconstruction efficace pour résoudre le problème inverse. Celui-ci se rapporte à la reconstruction de cartes de propriétés physiques (optique et/ou acoustiques) de l’intérieur du sein. La Méthode des Eléments Finis (MEF) a été retenue pour résoudre l’équation de propagation optique. Pour la résolution de l’équation de propagation acoustique, une méthode semi-analytique, basée sur des calculs par transformées de Fourier (méthod k-space), a été choisie. Pour la résolution du problème inverse, deux approches ont été étudiées : i) un sondage passif, permettant de remonter à la distribution de pression initiale, à l’aide de la méthode de retournement temporel ; ii) un sondage actif, où l’on interroge le milieu sélectivement sous différentes excitations, permettant de remonter quantitativement aux propriétés optiques du milieu. On appelle cette dernière approche Tomographie PhotoAcoustique Quantitative (TPAQ). Une étude spécifique sur le protocole d’illumination/détection a été conduite, prenant également en compte les contraintes expérimentales. / The present work was motivated by the development of alternative imaging techniques for breast cancer early diagnosis, that is photoacoustic imaging, which potentially couples the merits of optical imaging and ultrasound imaging, that is high optical functional contrasts brought by optical probing and high spatial resolution by ultrasound detection. Our work aims at modeling the photoacoustic multiwave phenomenon and incorporate it in an efficient reconstruction algorithm to solve the inverse problem. The inverse problem consists in the recovery of interior maps of physical properties of the breast. The forward model couples optical and acoustic propagations. The Finite Element Method (FEM) was chosen for solving the optical propagation equation, while a semi-analytical method based on Fourier transforms calculations (k-space method) was preferred for solving the acoustic propagation equation. For the inverse model, time reversal method was adopted to reconstruct the initial pressure distribution, an active approach of the inverse problem was also achieved, which decoupled the optical properties from measured photoacoustic pressure, this approach is called quantitative photoacoustic tomography (QPAT), in this approach, illumination/detection protocol was studied, and the experimental set up is also take into consideration. In the last step, photoacoustic pressure measurements obtained from experiment and simulation are studied and compared.

Cette thèse porte sur l'étude de la méthode d'imagerie photoacoustique, une nouvelle modalité hybride permettant de combiner la haute résolution de l'imagerie par ultrasons et le contraste de l'imagerie optique. Nous y étudions en particulier le problème inverse associé et sa résolution : il se décompose en l'inversion de l'équation d'ondes et en celle de l'équation de diffusion optique, dont le but est de retrouver les paramètres optiques du milieu. Dans la première partie de cette étude nous développons un modèle permettant de prendre en compte les variations de la vitesse acoustique dans le milieu biologique. En effet, la plupart des méthodes d'inversion supposent une vitesse acoustique constante, ce qui est à l'origine d'erreurs dans les reconstructions. La deuxième partie de la thèse porte sur une étude mathématique du phénomène de limitation de la profondeur de l'imagerie photoacoustique. Nous calculons une estimation de stabilité du problème inverse dans le cas d'un milieu stratifié et nous montrons que la reconstruction se dégrade avec la profondeur. Nous étudions dans la dernière partie le phénomène photoacoustique en présence de nanoparticules métalliques : ces marqueurs permettent d'amplifier par des résonances le signal photoacoustique généré autour d'elles. Elles permettent ainsi une meilleure visibilité des tissus en profondeur. Nous explicitons ici le modèle mathématique de génération du signal photoacoustique, ainsi que la résolution théorique du problème inverse photoacoustique dans ce contexte. / This thesis work is related to photoacoustic imaging techniques which are new multiwave modalities in medical imaging that combine both high resolution of ultrasounds and contrast of optical methods. Weprecisely studied the inverse problem that consists of determining the optical coefficients of biologicaltissues from measurement of acoustic waves generated by the photoacoustic effect. The photoacoustic inverse problem proceeds in two steps.We first retrieve the initial pressure from the measurement of the pressure wave on a part of the boundary of the sample. The first inversion takes then the form of a linear inverse source problem and provides internal data for the optical waves that are more sensitive to the contrast of the absorption and diffusion coefficients. In a second step we recover the optical coefficients from the acquired internal data.The aim of this work is to study the two inversions in different contexts. In the first part, we develop a model that takes into account the variations of the acoustic speed in the medium. Indeed, most of the inversion methods suppose that the acoustic speed is constant, and this assumption can lead to errors in the reconstruction of the optical coefficients. The second part of this work is the derivation of stability estimates for the photoacoustic inverse problem in a layered medium. We prove that the reconstruction is getting worse with depth. This is one of the main drawbacks of the photacoustic method, the imaging depth is limited to a few centimeters. The last part is about photoacoustic generation with plasmonic nanoparticles. They enhance the photoacoustic signal around them, so that we can investigate the tissue more deeply. We derive the mathematical model of the photoacoustic generation by heating nanoparticles, and we solve the photoacoustic inverse problem in this context.

Abstract Determination of blood glucose level is a frequently occurring procedure in diabetes care. As the most common method involves collecting blood drops for chemical analysis, it is invasive and liable to afflict a degree of pain and cause a skin injury. To eliminate these disadvantages, this thesis focuses on pulsed photoacoustic techniques, which have potential ability in non-invasive blood glucose measurement. The fundamental theory of photoacoustics in liquid and soft tissue was studied systematically. The distributions of photoacoustic sources in a near-infrared optical skin model were simulated by the Monte Carlo method. Expansion coefficient and specific heat of glucose solution were measured by thermodynamic method, while the sound velocity in it was determined by photoacoustic approach. The effect of glucose on blood optical scattering was studied by a picosecond pulsed laser together with a streak camera. A photoacoustic apparatus comprising a pulsed laser diode and a piezoelectric transducer was built and applied to measure glucose concentration in water and scattering media. Moreover, this apparatus was also used to non-invasive experiment on human fingers. The measurements showed that the expansion coefficient, specific heat and acoustic velocity change by 1.2%, -0.6% and 0.28%, respectively, in response to a 1% change in glucose concentration. The sum effect of these parameters to photoacoustic signal was much larger than that of optical absorption of glucose in near infrared wavelengths, which provided photoacoustic technique a higher degree of sensitivity than offered by the optical absorption method. At the wavelength of 905 nm, the measured glucose detection sensitivity in a 3% milk solution, a tissue sample and whole human blood was 5.4%, 2.5% and 14%, respectively. Each figure is higher than that of glucose in water, about 2%, for a one percent change in glucose concentration. This was supported by the temporal dispersion curves of glucose in blood samples, which demonstrated that glucose decreased the optical scattering of tissues. The currently photoacoustic apparatus could detect the minimal glucose concentration of 100 mg/dl in whole blood samples. It is sensitive to physiological changes in non-invasive measurement, but insufficient for evaluating change in the physiological glucose concentration. Current photoacoustic techniques have apparently advantages in study of scattering media and made great progress in tissue imaging and diagnosis. However, in non-invasive blood glucose measurement they met similar problems as optical approaches based on scattering effect.