In the presented research we propose to improve on historically accepted radiobiological models via Monte Carlo simulation of radiation tracks passing through a cell nucleus modeled with up-to-date subnuclear structures. This is performed by generating a radiation track database using the Monte Carlo code, Geant4-DNA, that simulates radiation interactions at the nanometer scale of DNA. These tracks are called upon from the database and intersected with a cell nucleus model that incorporates DNA-containing structures. This allows for a Monte Carlo simulation of how DNA double strand breaks are produced by radiation. The results can be used to correlate to many experimentally observed biological endpoints, e.g. chromosome aberrations as well as cell death.

No combate às neoplasias, diferentes técnicas radioterápicas têm surgido, apoiadas em avanços tecnológicos, com o objetivo de otimizar a eliminação das células tumorais produzindo o menor dano a tecidos sadios dos pacientes. Os planejamentos de tratamentos radioterápicos visam o estabelecimento de parâmetros técnicos de irradiação de forma que as doses prescritas nos volumes de tratamento sejam atingidas. Enquanto que a prescrição das doses se baseiam em considerações biológicas de radiosensibilidade dos tecidos, os cálculos físicos do planejamento levam em conta parâmetros dosimétricos associados aos feixes de radiação e às características físicas dos tecidos irradiados. A incorporação de informações de sensibilidade de tecidos aos cálculos radioterápicos pode auxiliar na particularização de tratamentos e no estabelecimento de critérios de comparação e escolha de técnicas radioterápicas, contribuindo para o controle tumoral e sucesso do tratamento. Para tanto, modelos biológicos de resposta celular à radiação ionizante devem ser bem estabelecidos. Este trabalho visou estudar a aplicabilidade do uso de modelos biológicos em cálculos de planejamento radioterápico com objetivo de auxiliar na avaliação de técnicas radioterápicas. A probabilidade de controle tumoral (TCP) foi estudada para duas formulações do modelo linear-quadrático, com e sem consideração de repopulação celular, em função de parâmetros de planejamento, como dose por fração, e de parâmetros radiobiológicos, como a razão ?/?. Além disso, o uso de critérios biológicos para comparação de técnicas radioterápicas foi testado através da simulação de um planejamento de próstata utilizando simulação Monte Carlo com o código PENELOPE. Posteriormente, planejamentos radioterápicos de tumores de próstata de cinco pacientes do Hospital das Clínicas da Faculdade de Medicina de Ribeirão Preto, USP, utilizando-se três técnicas de irradiação diferentes, foram comparados através do critério de probabilidade de controle tumoral. Para tanto, matrizes de dose obtidas do sistema de planejamento radioterápico XiO foram utilizadas para obter as distribuições de TCP e histogramas TCP-volume. Os estudos realizados permitiram concluir que variações em parâmetros radiobiológicos podem influenciar significativamente cálculos de controle tumoral e que a análise de histogramas TCP-volume pode fornecer informações importantes para avaliação de tratamentos radioterápicos. Entretanto, o estabelecimento de fatores quantitativos de comparação através de critérios radiobiológicos passa pelo estabelecimento de protocolos de prescrição clínica baseados nesses critérios. Além disto, valores radiobiológicos sofreram grandes alterações na literatura recentemente e, portanto, a inclusão destes parâmetros nos cálculos de planejamentos requer grandes cuidados. / In the fight against cancer, different irradiation techniques have been developed based on technological advances and aiming to optimize the elimination of tumor cells with the lowest damage to healthy tissues. The radiotherapy planning goal is to establish irradiation technical parameters in order to achieve the prescribed dose distribution over the treatment volumes. While dose prescription is based on radiosensitivity of the irradiated tissues, the physical calculations on treatment planning take into account dosimetric parameters related to the radiation beam and the physical characteristics of the irradiated tissues. To incorporate tissue\'s radiosensitivity into radiotherapy planning calculations can help particularize treatments and establish criteria to compare and elect radiation techniques, contributing to the tumor control and the success of the treatment. Accordingly, biological models of cellular response to radiation have to be well established. This work aimed to study the applicability of using biological models in radiotherapy planning calculations to aid evaluating radiotherapy techniques. Tumor control probability (TCP) was studied for two formulations of the linear-quadratic model, with and without repopulation, as a function of planning parameters, as dose per fraction, and of radiobiological parameters, as the ?/? ratio. Besides, the usage of biological criteria to compare radiotherapy techniques was tested using a prostate planning simulated with Monte Carlo code PENELOPE. Afterwards, prostate plannings for five patients from the Hospital das Clínicas da Faculdadede Medicina de Ribeirão Preto, USP, using three different techniques were compared using the tumor control probability. In that order, dose matrices from the XiO treatment planning system were converted to TCP distributions and TCP-volume histograms. The studies performed allow the conclusions that radiobiological parameters can significantly influence tumor control calculations and that the TCP-volume histograms can provide important information for treatment techniques evaluation. However, the establishment of quantitative comparison parameters using radiobiological criteria demands the establishment of prescription protocols based on these same parameters. Also, the literature recently showed large variations in radiobiological parameters, meaning that the inclusion of those in treatment planning calculations should require a careful endeavor.

This work describes the development and application of a novel combination of single-cell Raman spectroscopy (RS), automated data processing, and principal component analysis (PCA) for investigating radiation induced biochemical responses in human tumour cells. The developed techniques are first validated for the analysis of large data sets (~200 spectra) obtained from single cells. The effectiveness and robustness of the automated data processing methods is demonstrated, and potential pitfalls that may arise during the implementation of such methods are identified. The techniques are first applied to investigate the inherent sources of spectral variability between single cells of a human prostate tumour cell line (DU145) cultured {\it in vitro}. PCA is used to identify spectral differences that correlate with cell cycle progression and the changing confluency of a cell culture during the first 3-4 days after sub-culturing. Spectral variability arising from cell cycle progression is (i) expressed as varying intensities of protein and nucleic acid features relative to lipid features, (ii) well correlated with known biochemical changes in cells as they progress through the cell cycle, and (iii) shown to be the most significant source of inherent spectral variability between cells. This characterization provides a foundation for interpreting spectral variability in subsequent studies. The techniques are then applied to study the effects of ionizing radiation on human tumour cells. DU145 cells are cultured in vitro and irradiated to doses between 15 and 50 Gy with single fractions of 6 MV photons from a medical linear accelerator. Raman spectra are acquired from irradiated and unirradiated cells, up to 5 days post-irradiation. PCA is used to distinguish radiation induced spectral changes from inherent sources of spectral variability, such as those arising from cell cycle. Radiation induced spectral changes are found to correlate with both the irradiated dose and the incubation time post-irradiation, and to arise from biochemical differences in lipids, nucleic acids, amino acids, and conformational protein structures between irradiated and unirradiated cells. This study is the first use of RS to observe radiation induced biochemical effects in single cells, and is the first use of vibrational spectroscopy to observe such effects independent from cell cycle or cell death related processes. The same methods are then applied to a panel of human tumour cell lines, derived from prostate (DU145, PC3, LNCaP and PacMet), breast (MDA-MB-231 and MCF7) and lung (H460), which vary by p53 gene status and intrinsic radiosensitivity. One radiation induced PCA component is detected for each cell line by statistically significant changes in the PCA score distributions for irradiated samples, as compared to unirradiated samples, in the first 24 to 72 hours post-irradiation. These RS response signatures arise from radiation induced changes in cellular concentrations of aromatic amino acids, conformational protein structures, and certain nucleic acid and lipid functional groups. Correlation analysis between the radiation induced PCA components separates the cell lines into three unique RS response categories: R1 (H460, MCF7 and PacMet), R2 (MDA-MB-231 and PC3), and R3 (DU145 and LNCaP). These RS categories partially segregate according to radiosensitivity; the R1 and R2 cell lines are radioresistant and the R3 cell lines are radiosensitive (PacMet radiosensitivity (R1) unknown). The R1 and R2 cell lines further segregate according to p53 gene status, corroborated by cell cycle analysis post-irradiation. Preliminary results obtained from a mouse prostate tumour cell line (TRAMP-C2), irradiated both in vitro and in vivo, indicate that RS signatures of radiation response may also be detectable from tumour cells obtained from an in vivo system during radiation therapy treatment. These results indicate the potential for future RS studies designed to investigate, monitor, or predict radiation response. / Graduate

Thesis (DR. P.H.)--University of Michigan. / No collective t.p.; title from cover.

Thesis (DR. P.H.)--University of Michigan. / No collective t.p.; title from cover.

Thesis (M. Sc.)--Carleton University, 2001. / Includes bibliographical references (p. 79-84). Also available in electronic format on the Internet.

Thesis (Ph. D.)--University of Wisconsin-Madison, 1989. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves: 108-122).

Made available in DSpace on 2014-10-09T12:23:29Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T13:56:41Z (GMT). No. of bitstreams: 1 01114.pdf: 3545903 bytes, checksum: 16ffa3cd108f92ce30100ddc3a442ad9 (MD5) / Tese (Doutoramento) / IEA/T / Faculdade de Medicina, Universidade de Sao Paulo - FM/USP

Orientador: Solange Maria de Almeida / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Odontologia de Piracicaba / Made available in DSpace on 2018-08-05T09:03:13Z (GMT). No. of bitstreams: 1 Domingos_AndreadeCastro_D.pdf: 10036948 bytes, checksum: 7cd917fe62aa2831d762e0ac4943e2b4 (MD5) Previous issue date: 2005 / Resumo: O objetivo do presente trabalho foi avaliar morfologicamente e imunohistoquimicamente os efeitos agudos da radiação gama em glândulas parótidas de ratos. Vinte e um animais foram divididos em 2 grupos experimentais, controle e irradiado, sendo o primeiro grupo composto por animais não expostos à radiação e o segundo formado for animais que tiveram a região de cabeça e pescoço irradiada com uma dose única de 15 Gy. As glândulas parótidas dos animais irradiados foram removidas nos tempos 4, 8, 12, 24, 48 e 72 horas após sua exposição e, posteriormente, submetidas ao processamento tecidual por meio de métodos imunohistoquímicos, para a avaliação da proteína laminina, e pela coloração por hematoxilina-eosina, para avaliação morfológica. Não foram observadas diferenças estatisticamente significantes entre a intensidade demarcação da laminina do grupo controle e do grupo irradiado. Por outro lado, verificou-se o espessamento e a presença de conglomerados de laminina nas membranas basais acinares dos subgrupos irradiados, tendo sido detectadas diferenças significativas entre o grupo controle e o subgrupo 72 horas (p = 0,0315). A análise morfológica revelou a presença de figuras de necrose nuclear, vacuolizações e severa atrofia dos ácinos, especialmente nos grupos 8 e 12 horas. O início do processo de reparo se deu no tempo 24 horas, mas não foi observada recuperação completa nos últimos tempos avaliados. Os resultados sugeriram que os efeitos deletérios da radiação ionizante podem ser um dos fatores responsáveis pelo remodelamento da matriz extracelular / Abstract: The aim of this study was to evaluate the morphological and immunohistochemical characteristics of gamma radiation acute effects on rat parotid glands. Twenty-one animals were divided into two groups: control and irradiated ones. The former was composed by non-exposed animals, while the latter was formed by rats which had their necks and heads irradiated with 15 Gy. The parotid glands were excised at 4, 8, 12, 24, 48 and 72 hours after irradiation and, then, were submitted to immunohistochemical methods, for the evaluation of the laminin, and to hematoxylin-eosin staining, for the analysis of the morphology. Laminin staining intensities between irradiated and non-irradiated salivary glands showed no statistically significant differences. Nevertheless, it was observed that laminin thickening and laminin-positive conglomerations demonstrated significant differences between the control and the 72-hour-subgroup (p= 0,0315). The morphological analysis revealed the presence of vacuolation, acinar atrophy and necrosis, especially by 8 and 12 hours after irradiation. Regeneration of the tissue started by 24 hours, but it was not complete at 48 and 72 hours after exposure. The results showed that radiation may be one of the factors that induce extracellular matrix remodeling / Doutorado / Radiologia Odontologica / Doutor em Radiologia Odontológica

Made available in DSpace on 2014-10-09T12:23:29Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T13:56:41Z (GMT). No. of bitstreams: 1 01114.pdf: 3545903 bytes, checksum: 16ffa3cd108f92ce30100ddc3a442ad9 (MD5) / Tese (Doutoramento) / IEA/T / Faculdade de Medicina, Universidade de Sao Paulo - FM/USP