Biological materials are complex hierarchical systems subjected to external stimuli like mechanical forces, chemical potentials and electrical signals. A deeper understanding of the behavior of these materials is required for the response characterization of healthy and diseased conditions. The primary aim of this dissertation is to study the mechanics of biological materials like cells and tissues from a computational perspective and relate its behavior with experimental works, so as to provide a framework for the identification and treatment of pathological conditions like cancer and vascular diseases. The first step towards understanding the behavior of a biological cell is to comprehend its response to external mechanical stimuli. Experimentally derived material properties of cells have found to vary by orders of magnitude even for the same cell type. The primary cause of such disparity is attributed to the stimulation process, and the theoretical models used to interpret the experimental data. The variations in mechanical properties obtained from the experimental and theoretical studies can be overcome only through the development of a sound mathematical framework correlating the derived mechanical property with the cellular structure. Such a formulation accounting for the inhomogeneity of the cytoplasm due to stress fibers and actin cortex is developed in this work using Mori-Tanaka method of homogenization. Mechanical modeling of single cells would be extremely useful in understanding its behavior in an experimental setup. Characterization of in-vivo response of cells requires mathematical modeling of the embedding environment like fibers and fluids, which forms the extra cellular matrix. Studies on fluid-tissue interactions in biomechanics have primarily relied on either an iterative solution of the individual solid or tissue phases or a sequential solution of the entire domain using a coupled algorithm. In this dissertation, a new computational methodology for the analysis of fluid-tissue interaction problem is presented. The modeling procedure is based on a biphasic representation of fluid and tissue domain, consisting of fluid and solid phases. The biphasic-fluid interaction model is also implemented to study the transfer of low-density lipoprotein from the blood to the arterial wall, and also the nutrient transfer in the tissue scaffolds of a bioreactor.

Pluchea indica (L.) Less. is a shrub of the family Compositae and is widespread along the western coast in Taiwan. Previous studies indicated that the components of Pluchea indica have potent anti-inflammatory, anti-ulcer and antimicrobial activities. In the present study, the effects of aqueous extract of roots and leaves from P. indica on the cancer cell lines were investigated. Various experimental approaches including cell growth curves, MTT assay, MTS assay, focus formation assay and cell migration assay were performed on the aqueous extract-treated cancer cells. Our results demonstrate the aqueous extract of P. indica induced anti-proliferation activity on GBM8401 and HeLa cancer cell lines.

The present study explored the effect of three anti-tumor drugs (cisplatin, fluorouracil, and temozolomide) on viability and cytosolic free Ca2+ concentrations ([Ca2+]i) in OC2 human oral cancer cells. The effect of cisplatin related mitogen-activated protein kinases (MAPKs) phosphorylation was also examined. Cisplatin at concentration of 25-150 £gM decreased viability in a concentration-dependent manner, and so did fluorouracil (50-1000 £gM) and temozolomide (50-600 £gM). The three anti-tumor drugs all failed to induce a [Ca2+]i increase; thus it seemed that these drugs induced cell death via Ca2+-independent pathways. Immunoblotting showed that OC2 cells have background phospho-ERK, phospho-JNK and phospho-p38 MAPKs. It was found that cisplatin influenced the phosphorylation of ERK, JNK and p38 MAPKs at different time points.

Micro (mi)RNAs have recently been found to play an important role in cancer biology. In order to further understand how miRNAs affect prostate tumour progression, we evaluated miRNA expression in two invasive prostate tumour lines, PC3 and DU145. We then focused our evaluation on a novel miRNA, miR-105, whose levels were significantly decreased in both tumour cell lines as compared to normal prostate epithelial cells. As miR-105 levels were reduced in prostate tumour cell lines, we restored its expression following transfection of cells with mimic constructs to over-express miR-105 in both cell lines, in order to determine its effect on various tumourigenic properties. Over-expression caused decreased tumour cell proliferation, anchorage-independent growth and invasion in vitro and inhibited tumour growth in vivo. We further identified CDK6 as a putative target of miR-105, which likely contributed to its inhibition of tumour cell growth. Our results suggest that miR-105 inhibits tumour cell proliferation and may be an interesting target to regulate tumour growth or potentially used as a biomarker to differentiate between less and more aggressive tumours in patients.

Thesis (Ph.D.)--Memorial University of Newfoundland, 2002. / Restricted until May 2005. Bibliography: leaves162-192.

Le processus de migration est un processus moléculaire intégré qui contribue in vivo à de nombreux processus physiologiques de motilité, comme le développement, la surveillance immunitaire et les métastases du cancer. Pour comprendre la migration cellulaire, il est nécessaire de considérer l'environnement de la cellule, le type de cellules et la morphologie ainsi que l'organisation interne, i.e. son cytosquelette et ses adhérences focales. Ce travail se concentre sur l'étude de la migration de cellules cancéreuses de la vessie sur des supports déformables. L'analyse de trois lignées cellulaires de capacité métastatique différente est présentée. La capacité des cellules cancéreuses à réagir à leur environnement est analysée et le processus de migration est décrit en termes de contraintes de traction. La réorganisation interne de la structure des cellules est étudiée par l'observation microscopique des filaments d'actine, des moteurs de myosine et des sites de transmission de force, i.e. les adhésions focales. On s'intéresse à la relation de complémentarité entre les différentes capacités invasives des cellules cancéreuses, les forces de traction ainsi que les forces exercées sur le lamellipode et les structures internes des cellules. Il est constaté que plusieurs paramètres peuvent être utilisés pour discriminer la capacité métastatique, comme le type de migration, les forces de traction, les zones focales d'adhésion, ainsi que l'indice de diffusivité de migration. Cette étude constitue donc une première tentative pour différencier diverses cellules invasives en utilisant la migration sur des substrats mous. / The migration process is a physically integrated molecular phenomena that contributes to many physiological motility in vivo processes such as development, immune surveillance and cancer metastasis. To understand cell migration it is necessary to consider the cell's environment, cell type and morphology as well as the internal organization of the cells, i.e. its cytoskeleton and focal adhesions. This work focuses on the study of migrating bladder cancer cells on two--dimensional deformable substrates. The analysis of a panel of three cell lines with different invasive capacity is presented. The ability of cancer cells to respond to their environment is analysed and the migration process is described in terms of traction stresses. The internal reorganization of cells structure is studied by microscopic observation of the actin filaments, the myosin motors and the sites of force transmission, i.e. the focal adhesions. The complementary relationship among different invasive capacities of cancer cells, traction stresses as well as the forces exerted on the lamellipodium and internal structures of cells are discussed. It is found that several parameters can be used for discriminating invasiveness, such as migration type, traction forces, focal adhesion areas, as well as the migration diffusivity index. This study therefore constitutes a first attempt to differentiate various invasive cells using migration on soft substrates.

Made available in DSpace on 2014-06-11T19:27:18Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-03-18Bitstream added on 2014-06-13T19:35:13Z : No. of bitstreams: 1 pereira_rs_me_araca.pdf: 790587 bytes, checksum: 194f37c505d12a98d7b75720d14a4e5f (MD5) / A causa dos tumores da glândula perianal é desconhecida, entretanto eles podem ser modulados por influência de hormônios gonadais, basicamente a testosterona, uma vez que há uma alta prevalência (até 95%) de regressão da forma benigna dessa neoplasia após a castração dos machos. A proteína Ki-67 foi descrita pela primeira vez em 1983 é uma proteína nãohistona, que não é expressa em células na fase G0, mas que pode ser detectada nas fases ativas do ciclo celular, G1, S, G2 e mitose Verificou-se que o Ki-67 está associado com a proliferação celular, estando a população de células tumorais positivas ao Ki-67 correlacionada com a evolução clínica da doença. Foram estudados 42 casos de neoplasias da glândula perineal, sendo 15 adenomas, 15 epiteliomas, 12 carcinomas. Foram utilizadas 13 amostras de tecido de glândula perianal normal, como controle. De cada animal foram obtidas informações como: sexo, idade, raça, presença de outras neoplasias, se castrado ou não, se em caso de fêmea havia o uso de anticoncepcional, tempo de evolução, recidiva e tempo de sobrevida. Dos 42 casos de neoplasias de glândulas perianais, 34 (80,95%) foram provenientes de cães machos e 8 (19,05%) de fêmeas. A média de idade dos machos com a neoplasia foi de 9,8 anos, e das fêmeas 7,4 anos, sendo que para os dois sexos a idade variou de 5 a 15 anos. Em relação às raças, 11 (26,19%) eram S.R.D. (sem raça definida), 9 (21,42%) Poodle, 4 (9,52%) Cocker Spaniel e Husky Siberiano. O histórico de diagnóstico de outras neoplasias, foi constatado em 23,80% dos animais deste estudo. Em relação à castração, 32 (94,11%) dos machos não eram castrados e apenas 8 (19,05%) fêmeas eram. O tempo de evolução da neoplasia apresentou média de 1,2 anos. Histórico de recidiva da neoplasia foram relatados em 14 animais, sendo maior... / The cause of perianal gland tumors is unknown, though they may be modulated by the influence of gonadal hormones, primarily testosterone, since there is a high prevalence (up to 95%) of regression of the benign tumor after males castration. Protein Ki-67 was first described in 1983, and it is a nonhistone protein, which is not expressed in G0 phase cells, but can be detected in the active phases of the cell cycle, G1, S, G2 and mitosis. It was found that Ki-67 is associated with cell proliferation, and the population of tumor cells positive for Ki-67 correlated with clinical outcome. In this present work we studied 42 cases of perineal gland neoplasms, being 15 adenomas, 15 epitheliomas and 12 carcinomas. We used 13 tissue samples of normal perianal gland as control. From each animal we obtained information such as gender, age, race, presence of other malignancies, neutering status, whether if there were female contraceptive use, duration, recurrence and survival time. Of the 42 cases of perianal gland neoplasms, 34 (80.95%) were from males and eight dogs (19.05%) females. The males average age with cancer was 9.8 years and 7.4 years for females and for both sexes aged between 5 and 15 years. In regard to race, 11 (26.19%) were no breed, 9 (21.42%) were Poodle, and 4 (9.52%) were Siberian Husky and Cocker Spaniels. The history of diagnosis of other cancers, was found in 23.80% of the animals in this study. In relation to castration, 32 (94.11%) of males were not castrated and only 8 (19.05%) were females. The progression of the tumor showed an average of 1.2 years. History of recurrence of cancer were reported in 14 animals, and was higher (66.66%) for carcinomas. Ki-67 staining revealed that the carcinomas showed higher proliferation rate (9.87%) compared to groups of epitheliomas... (Complete abstract click electronic access below)

Micro (mi)RNAs have recently been found to play an important role in cancer biology. In order to further understand how miRNAs affect prostate tumour progression, we evaluated miRNA expression in two invasive prostate tumour lines, PC3 and DU145. We then focused our evaluation on a novel miRNA, miR-105, whose levels were significantly decreased in both tumour cell lines as compared to normal prostate epithelial cells. As miR-105 levels were reduced in prostate tumour cell lines, we restored its expression following transfection of cells with mimic constructs to over-express miR-105 in both cell lines, in order to determine its effect on various tumourigenic properties. Over-expression caused decreased tumour cell proliferation, anchorage-independent growth and invasion in vitro and inhibited tumour growth in vivo. We further identified CDK6 as a putative target of miR-105, which likely contributed to its inhibition of tumour cell growth. Our results suggest that miR-105 inhibits tumour cell proliferation and may be an interesting target to regulate tumour growth or potentially used as a biomarker to differentiate between less and more aggressive tumours in patients.

A significant body of literature has reported strategies and techniques to assess the mechanical properties of biological samples such as proteins, cellular and tissue systems. Atomic force microscopy has been used to detect elasticity changes of cancer cells. However, only a few studies have provided a detailed and complete protocol of the experimental procedures and data analysis methods for non-adherent blood cancer cells. In this work, the elasticity of NB4 cells derived from acute promyelocytic leukemia (APL) was probed by AFM indentation measurements to investigate the effects of the disease on cellular biomechanics. Understanding how leukemia influences the nanomechanical properties of cells is expected to provide a better understanding of the cellular mechanisms associated to cancer, and promises to become a valuable new tool for cancer detection and staging. In this context, the quantification of the mechanical properties of APL cells requires a systematic and optimized approach for data collection and analysis, in order to generate reproducible and comparative data. This Thesis elucidates the automated data analysis process that integrates programming, force curve collection and analysis optimization to assess variations of cell elasticity in response to processing criteria. A processing algorithm was developed by using the IGOR Pro software to automatically analyze large numbers of AFM data sets in an efficient and accurate manner. In fact, since the analysis involves multiple steps that must be repeated for many individual cells, an automated and un-biased processing approach is essential to precisely determine cell elasticity. Different fitting models for extracting the Young’s modulus have been systematically applied to validate the process, and the best fitting criteria, such as the contact point location and indentation length, have been determined in order to obtain consistent results. The designed automated processing code described in this Thesis was used to correlate alterations in cellular biomechanics of cancer cells as they undergo drug treatments. In order to fully assess drug effects on NB4 cells, viability assays were first performed using Trypan Blue staining for primary insights before initiating thorough microplate fluorescence intensity readings using a LIVE/DEAD viability kit involving ethidium and calcein AM labelling components. From 0 to 24 h after treatment using 30 µM arsenic trioxide, relative live cell populations increased until 36 h. From 0 to 12 h post-treatment, relative populations of dead cells increased until 24 h post-treatment. Furthermore, a drastic drop in dead cell count has been observed between 12 and 24 h. Additionally, arsenic trioxide drug induced alterations in elasticity of NB4 cells can be correlated to the cell viability tests. With respect to cell mechanics, trapping of the non-adherent NB4 cells within fabricated SU8-10 microwell arrays, allowed consistent AFM indentation measurements up to 48 h after treatment. Results revealed an increase in cell elasticity up to 12 h post-treatment and a drastic decrease between 12 and 24 h. Furthermore, arsenic trioxide drug induced alterations in elasticity of NB4 cells can be correlated to the cell viability tests. In addition to these indentation and viability testing approaches, morphological appearances were monitored, in order to track the apoptosis process of the affected cells. Relationships found between viability and elasticity assays in conjunction with morphology alterations revealed distinguish stages of apoptosis throughout treatment. 24 h after initial treatment, most cells were observed to have burst or displayed obvious blebbing. These relations between different measurement methods may reveal a potential drug screening approach, for understanding specific physical and biological of drug effects on the cancer cells.

Background: Oesophagogastric cancers generally present late in life with advanced disease and carry a poor prognosis. Few patients receive curative treatment. Polo-like Kinase 1 (PLK1) is a mitotic kinase with regulatory functions at the G2/M cell cycle phase transition. In mammalian cells, PLK1 phosphorylates and activates FOXM1, a forkhead transcription factor at the G2/M cell cycle phase transition. FOXM1 then promotes transcription of multiple gene products, including PLK1 and CCNB1, which then act individually or in complexes to further phosphorylate FOXM1 generating a positive feedback loop driving the cell into M phase. Aims: We aimed to assess the expression of PLK1 and FOXM1 in oesophageal and gastric cancer patients. Secondly we aimed to investigate the expression and inter- relationship of PLK1 and FOXM1 in oesophageal cell lines during the cell cycle. Results: FOXM1 and PLK1 are expressed in oesophageal cell lines and demonstrate cross-regulatory interactions. Inhibition of PLK1 leads to the decreased expression of FOXM1 and it’s target gene in oesophageal cell lines. FOXM1 and PLK1 are also concomitantly overexpressed in a large proportion of oesophageal and gastric carcinoma’s at both the protein and mRNA level. Other FOXM1 target genes including, CCBN1, AURKB and CKS1 are co-expressed in a similar manner. In a homogenous cohort of patients who underwent surgery, the expression of PLK1 and AURKB was prognostic for overall survival. Conclusions: This study has demonstrated that FOXM1 and a number of target genes including PLK1 are coordinately expressed in a proportion of oesophageal and gastric carcinomas. This suggests that chemotherapeutic treatments that target this pathway may be of clinical utility.