iii Abstract Cells have evolved multiple mechanisms of cellular motility ranging from the migration of large cell cohorts to specialized migration of individual cells. The wide range of invasion modes has been exploited by cancer cells to their advantage, which has rendered the metastatic process so difficult to defeat. To allow for a better understanding of cancer invasion plasticity, we have employed studies on cancer cells that adopt the proteolytically active, adhesion-dependent, elongated mesenchymal invasion mode, the protease-independent, low adhesion, rounded amoeboid invasion mode, or combination of both. To study invasion plasticity directly, we have established two model systems of the mesenchymal- amoeboid transition (MAT) that allow for regulated induction of MAT in 3D in vitro environments. Using these systems, MAT was induced in HT1080 fibrosarcoma cells and the acquisition of a motile, invasive amoeboid phenotype was confirmed. We then observed the mesenchymal and amoeboid invasion strategies within 3D collagen in more detail using a digital holographic microscope. Further, HT1080 cells before and after MAT were subject to high throughput proteomic and transcriptomic studies. Comparison of gene expression and protein levels of mesenchymal and amoeboid cells disclosed an inflammatory-like...

Malaria is a disease caused by the protozoan parasite Plasmodium where the species that causes the most severe form of malaria in humans is known as Plasmodium falciparum. At least 40% of the global population is at risk of contracting malaria with 627 000 people dying as a result of this disease in 2012. Approximately 90% of all malaria deaths occur in sub-Saharan Africa, where approximately every 30 seconds a young child dies, making malaria the leading cause of death in children under the age of five years old. The malaria parasite has a complex life cycle utilising both invertebrate and vertebrate hosts across sexual and asexual stages. The erythrocyte invasion stage of the life cycle in the human whereby the invasive merozoite form of the parasite enters the erythrocyte is a central and essential step, and it is during this stage that the clinical symptoms of malaria manifest themselves. Merozoites invade erythrocytes utilising multiple, highly specific receptor-ligand interactions in a series of co-ordinated events. The aim of this study was to better understand the interactions occurring between the merozoite and erythrocyte during invasion by using modern, cutting-edge proteomic techniques. This was done in the hope of laying the foundation for the discovery of new key therapeutic targets for antimalarial drug and vaccine-based strategies, as there is currently no commercially available antimalarial vaccine and no drug to which the parasite has not at least started showing resistance. In this study healthy human erythrocytes were treated separately with different protein-altering enzymes and chemicals being trypsin, the potent oxidant sodium periodate (NaIO4), the amine cross-linker tris(2-chloroethyl)amine hydrochloride (TCEA) and the thiol cross-linker 1,11-bis(maleimido)triethylene glycol (BM(PEG)3). The resulting erythrocyte protein alterations were visualised as protein band differences on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE), where treated and untreated control erythrocyte ghost protein fingerprints were visually compared to one another. The protein bands showing differences between treated and control samples were in-gel digested using trypsin then sequenced by liquid chromatography tandem mass spectrometry (LC-MS/MS) and identified using proteomics-based software. In this way, the erythrocyte proteins altered by each enzyme/chemical treatment were identified. Malaria invasion assays were performed where each treatment group of erythrocytes as well as the control erythrocytes were incubated separately with schizont stage malaria parasites for the duration of one complete life cycle. Using fluorescent staining and flow cytometry, the invasion inhibition efficiency for each treatment group was evaluated. By utilising these methods, the identification and the relative importance of the erythrocyte proteins involved in the invasion process were determined. Protein fingerprints of control and treated erythrocyte ghosts were visualised and optimised on SDS PAGE where induced protein band differences were successfully identified by LC-MS/MS. It was found that each treatment altered erythrocyte proteins with changes found in Band 3, actin, phosphoglycerate kinase 1, spectrin alpha, spectrin beta, ankyrin, haemoglobin, Bands 4.1 and 4.2, glycophorin A and stomatin. The invasion assays revealed that TCEA inhibited invasion to the greatest extent as compared to the other treatments, followed by BM(PEG)3 and trypsin. Sodium periodate-treated erythrocytes could not be assessed using the invasion assay due to auto-haemolysis. Band 3, glycophorin A, Band 4.1 and stomatin appear to be of higher relative importance in the invasion process as compared to the other altered erythrocyte proteins. These results confirmed the known roles of spectrin alpha, spectrin beta, glycophorin A, Band 3 and Band 4.1 in invasion, and suggested that ankyrin, Band 4.2 and stomatin may also be involved. This study highlighted the potential that modern, cutting-edge proteomic techniques provide when applied to previous comparative studies found in older literature, as previously unidentified proteins that can be involved in invasion were revealed. These results can be used as a foundation in future studies in order to identify new key targets for the development of new antimalarial drug- and vaccine-based strategies, with the hope of preventing the suffering of the millions of malaria-inflicted people worldwide, and ultimately eradicating this deadly disease. / Dissertation (MSc)--University of Pretoria, 2014. / tm2015 / Pharmacology / MSc / Unrestricted

Tumour cell invasion is one of the most critical steps in malignant progression. It includes a broad spectrum of mechanisms, including both individual and collective cell migration, which enables them to spread towards adjacent tissue, and form new metastases. Understanding the mechanisms of cell spreading, and invasion, is crucial for effective anticancer therapy. Two modes of individual migration of tumour cells have been established in a three-dimensional environment. Mesenchymally migrating cells use proteases to cleave collagen bundles, and thus overcome the ECM barriers. Recently described protease-independent amoeboid mode of invasion has been discovered in studies of cancer cells with protease inhibitors. During my PhD study, I have focused on determining the molecular mechanisms involved in amoeboid invasion of tumour cells. We have examined invasive abilities in non-metastatic K2 and highly metastatic A3 rat sarcoma cell lines. We have shown that even though highly metastatic A3 rat sarcoma cells are of mesenchymal origin, they have upregulated Rho/ROCK signalling pathway. Moreover, A3 cells generate actomyosin-based mechanical forces at their leading edges to physically squeeze through the collagen fibrils by adopting an amoeboid phenotype. Amoeboid invasiveness is also less dependent on...

One theory concerning the invasiveness of exotic plants suggests that they exude allelopathic compounds that are novel in areas being invaded. Here, I tested for allelopathic effects of root exudates of the invasive plant Heracleum mantegazzianum (giant hogweed) in a series of experiments both in vitro and in the garden, and compared them with effects of its native congener H. sphondylium (common hogweed) and two less related native species. Moreover, I addressed intraspecific variability of allelopathy by comparing effects of 41 different maternal lines of H. mantegazzianum sampled from several areas and populations within the Czech Republic. Not only the effects, but also the identity of allelopathic compounds was investigated. In order to test also for the interaction of allelochemicals with soil microorganisms, I analysed the composition of soil microbial communities expressed as phospholipid and neutral lipid fatty acids (PLFA and NLFA, respectively) together with nutrient conditions and light availability at invaded sites in the Czech Republic. The results show that H. mantegazzianum is able to exert phytotoxic effects in vitro as well as in the garden experiment; however, its effects did not differ from the inhibition caused by the native species tested. Variation partitioning among areas,...

Riparian vegetation along the water courses is one of the places that are highly vulnerable to the spread of invasive alien plants. Water corridors allow their easy and rapid spread to new area. The diploma thesis occupies with changes in the distribution of the invasive neophytes in the riparian vegetation of the Berounka river during the years 2015-2018. The objective of this study was to find out how changes in the course of each year will be reflected and whether a certain trend of population development can be traced. In connection with this, the thesis occupies with the possible influence of extremely dry years 2015 and 2018 on the occurrence of invasive neophytes. Field research took place on the banks of the Berounka river from the village of Čilá near Skryje to the confluence of the Berounka and Vltava river in Prague-Lahovice during the summer months in 2015, 2016, 2017 and 2018. The riparian vegetation was divided into 500 ± 150 m long segments. There were 90 segments where the occurrence and abundance of 17 invasive neophytes were mapped. In total, occurrence of 14 taxons was registered. The most frequent taxons were Robinia pseudacacia, Impatiens glandulifera and Solidago sp. in the Berounka riverbank vegetation. The highest number of specimen was recorded for invasive neophyte...

What is democracy? Is it something that suits all countries. The western world encourages the outside world to democratize according to their guidelines. The purpose of the essay is to present two reasons, one about Iraq's failure to democratize, and the other about Tunisia's democratic success. The method in this essay include qualitative methods, where a democratized comparative study as well as the most similar systems design approach will be used. These studies will attempt to determine the composition of democratization outcomes, using three of the dimensions internal and external democratization factors as suggested by Larry Diamond in his book, as well as like the regime transition of Daniel Silander. The results of this study indicate that Iraq had more political dissent as well as ethnic and extremist groups during the invasion and less international support for its democratization beside the United States. On the other hand, Tunisia has a more unified civil society and thanks to the democratic support of the European Union, a peaceful transition to a democratic system is possible. In this essay, a deeper analysis of democratization will be highlighted, including how these essential factors have caused different outcomes in each country.

This dissertation explores the differential effects of fires and rodent communities on native and invasive desert plant communities. Chapter one examines the impacts of fire and repeat fires on fuel loads in two different Utah desert sites, one in the cool Great Basin Desert and one in the hyper-arid Mojave Desert, over the course of four years. We found that both desert sites responded with varying intensities to a single burn, but the effects of a reburn were not as pronounced. We also found that our Great Basin Desert site had a stronger response to fire than our Mojave Desert site, producing a higher plant fuel loads that could potentially exacerbate the changing fire regimes. These data can be used to help map the effects that climate change may have on fuel loads and the fire potential of these deserts. Chapter two uses a full factorial experimental design to compare the effects of rodent herbivory and fire on native versus invasive seedlings at our Mojave Desert field site. We found that rodent herbivory has a more negative impact on the survival of native seedlings than invasive seedlings. This could be because the invasives are grasses that tend to tolerate herbivory better than the native shrubs and forbs. Chapter three again uses a full factorial experimental design to assess the impacts of rodents and fire on the fate of native and invasive seeds at our field site in the Mojave Desert. We found that rodents removed seeds, and they did show preferences for some species of seed over others, but these preferences were not different between native and invasive seeds or seed mass. We propose that the preferences may be influenced by other seed traits such as water content, handling time or physical and chemical defenses. As with the seedlings in chapter two, fire did not have any impact on rodent seed preferences. These data highlight the importance of considering rodent effects on seeds used in restoration effects following wildfires.

Understanding cancer biomarkers for diagnosis and prognosis leads to improved patient treatments and care. This thesis addresses the relevance of thymidine kinase 1 (TK1) as a cancer biomarker and the role of TK1 in cancer progression. Worldwide, cancer leads to more than 12 million deaths annually. In the United States alone, each year over 1.5 million cases will be diagnosed and over half a million persons will die. The most prevalent cancer types include skin, lung, breast, prostate, and colon. TK1 is known to be present in the serum of patients with multiple cancer types, including lung, breast, colon and prostate. In fact, it is shown to be detectable in cancer patients even before they manifest clinical symptoms. Additionally, the levels of TK1 increase progressively with increasing tumor grade; meaning that levels of TK1 can indicate tumor grade. Cellular proliferation markers such as p53 and Ki-67 have been compared to TK1 in cancer diagnosis and prognosis. TK1 has potential as both a prognostic and diagnostic biomarker in various cancer types including breast. Breast cancer is one of the most aggressive cancer types with 20-30% of diagnosed tumors becoming metastatic. Recent findings have identified additional involvement of TK1 downstream of cellular proliferation in cancer progression, including cellular invasion which is a part of cancer metastasis. These findings while efficacious, fail to identify the individual contribution of TK1 in downstream processes that aid in cancer progression. As mentioned previously, TK1 is upregulated in several different cancer types. We propose that there is an advantage to upregulated levels of TK1 in cancer progression and seek to explore its role specifically in cell invasion and survival. Based on our current understanding of TK1, we first wanted to review the history of TK1 and show the importance of understanding this crucial enzyme. Finally, we report our results from experiments exploring the influence of TK1 in vitro on breast cancer cell invasion and survival.

Bc. Daniel Samek Abstrakt The diploma thesis "The august invasion of Soviet troops in 1968 in Brno and the reaction of local law enforcement agency, government, media and general public" aims for more complex and multiple perspectives approach the invasion and the subsequent occupation of Brno with the units of Sowiet Union. Klíčová slova : Brno, 1968, Pražské jaro, Sovětský svaz, invaze, okupace

Glucocorticoids are effective immunosuppressive and anti-inflammatory agents, but some aspects of stress appear to be proinflammatory. This study investigates this apparent paradox as it applies to stress exposure and the development of arthritis in a rat strain that has subnormal hypothalamic-pituitary-adrenal (HPA) responsiveness. Female Lewis rats were subjected to 1 week of rotating, psychological stressors for 5 h daily, beginning 7 days following inoculation with type [I collagen. The collagen-induced arthritis (CIA) group exposed lo stress showed reduced ankle width increase (p < 0.001) and decreased hindlimb severity scores (p < 0.001). At sacrifice, 2 days following stress termination, no differences in either measure remained and there was no difference in hind paw volume. However, the area of the tibia invaded by stroma, as quantitated by image analysis, was reduced in the stressed rats (p < 0.05). In animals exposed to stress, adrenal weights were increased (p < 0.005) and plasma corticosterone levels were elevated at sacrifice (p < 0.02). Both injected groups had significantly larger adrenal (p < 0.005) and lower thymus weights (p < 0.05) than did uninjected controls. Likewise, both CIA groups had reduced glucocorticoid receptor immunoreactivity in synovial membranes compared to controls (p < 0.001), suggesting that the Lewis rat's HPA deficiency may be intensified by glucocorticoid receptor downregulation during the induction of CIA. These data indicate that the responsiveness of the HPA axis to psychological stress in this strain is sufficient to alter disease progression.