The relationship between Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL) is not entirely elucidated and a clonal relation may be present more often than previously believed. Mechanisms of tumour progression and resistance to therapy are poorly understood. Between 1974 and 1994 all individuals in Sweden with both HL and NHL were identified. Thirty-two cases were studied using clinical, histopathological and immunohistochemical methods. The second lymphoma often appeared in an aggressive clinical form and a significant correlation between the expression of p53 and LMP-1 in the first and second lymphoma was demonstrated. The treatment outcome for 307 patients with advanced stages of HL, in an unselected population was in accordance with the treatment results of large centres world-wide. Some patients were successfully selected for a shorter chemotherapy-regimen without inferior treatment results. In 124 patients with relapse, the survival of those primarily treated with radiotherapy according to the National guidelines was in accordance with the survival of patients of initially advanced stages. A worse outcome was found for those who received both chemotherapy and radiotherapy initially, probably because of a higher frequency of bulky disease in this group. Immunohistochemical analysis of the tumour suppressor protein p53 and retinoblastoma protein (Rb) of paired samples at diagnosis and at relapse in 81 patients did not reveal any specific staining pattern affecting survival. A novel B-cell line (U-2932) was established from a patient with a diffuse large B-cell lymphoma previously treated for advanced stage and subsequent relapses of HL. An identical rearranged IgH gene was demonstrated in tumour cells from the patient and in U-2932. A p53 point mutation was detected and over-expression of the p53 protein was found. A complex karyotype with high-level amplifications of the chromosomal regions 18q21 and 3q27, i.e. the loci for bcl-2 and bcl-6 were demonstrated.

Chlamydiae are important human bacterial pathogens with an intracellular life cycle that consists of two distinct bacterial forms, an infectious form (EB) that infects the eukaryotic host cell, and a non-infectious form (RB) that allows intracellular proliferation. To be successful, chlamydiae need to alternate between EB and RB to generate infectious EB’s which are competent to infect new host cells. Chemical genetics is an attractive approach to study bacterial pathogenesis; in principal this approach relies on an inhibitory compound that specifically inhibits a protein of interest. An obstacle in using this approach is target identification, however whole genome sequencing (WGS) of spontaneous mutants resistant to novel inhibitory compounds has significantly extended the utility of chemical genetic approaches by allowing the identification of their target proteins and/or biological pathways. In this thesis, a chemical genetics approach is used, I have found that heme and glucose metabolism of C. trachomatis is specifically important for the transition from the RB form to the infectious EB form. Heme and glucose metabolism are both coupled to energy metabolism, which suggests a common link between the RB-to-EB transitions. In connection with the above findings I have developed strategies that enable the isolation of isogenic C. trachomatis mutant strains. These strategies are based on WGS of spontaneous mutant populations and subsequent genotyping of clonal strains isolated from these mutant populations. Experiments with the mutant strains suggest that the uptake of glucose-6-phosphate (G-6-P) regulates the RB-to-EB transition, representing one of the first examples where genetics has been used to study C. trachomatis pathogenesis. Additional experiments with the mutant strains indicate that G-6-P promotes bacterial growth during metabolic stress. In concert with other findings presented in this thesis, I have fine-tuned methods that could be employed to reveal how novel inhibitory chemical compounds affect chlamydiae. In a broader context, I suggest that C. trachomatis could be used as a model organism to understand how new inhibitory drugs affect other bacterial pathogens. In addition, I observed that C. pneumoniae infections resulted in generalized bone loss in mice and that these mice display a cytokine profile similar to infected bone cells in vitro. Thus, this study indicates that C. pneumoniae potentially can infect bone cells in vivo, resulting in bone loss, alternatively, the inflammatory responses seen in vivo could be the causative factor of the bone loss observed.

The 87Rb-86Sr system is a widely used long-lived isotope geochronometer. 87Rb, the naturally occurring radioactive isotope of Rb, undergoes beta-decay to stable 87Sr with a half-life of approximately 50 Ga. Decay of 87Rb to 87Sr results in variable 87Sr/86Sr in minerals with different Rb/Sr, and measurement of 87Rb/86Sr and 87Sr/86Sr allows for the determination of the age of the rock. Accurate ages depend both on the quality of the isotopic analysis and on the accuracy of the 87Rb decay constant, lambda87. Although the currently accepted value for lambda87 of 1.42 × 10-11a-1 has been in use for over 30 years, there is growing evidence that it is not accurate. Recent attempts to refine lambda87 and its precision have not reached a consensus. This thesis describes a new experiment to measure lambda87 by 87Sr accumulation over a period of about 30 years, and the preparation of a 84-86Sr double-spike in conjunction with that experiment. Radiogenic 87Sr produced in aliquots of a RbClO4 salt was measured by isotope dilution thermal ionization mass spectrometry. An average of 31 measurements yields a value of 1.398 ± 0.003 × 10-11a-1 . This requires a substantial revision from the previously accepted decay constant and makes Rb-Sr ages calculated with it 1.5% older. A Rb-Sr and U-Pb isotopic chronometry study was carried out on thirteen ordinary chondrites – the most common type of meteorite, the origin and history of which are still unclear. Some meteorites appear disturbed, possibly by recent shock during breakup of the parent body, whereas others yielded accurate and precise U-Pb and Pb-Pb ages. For example, L5 Elenovka yielded distinct ages for silicates (4555 Ma) and phosphates (4535 Ma), allowing the cooling rate of this meteorite from approximately 1055 K to 759 K to be constrained to 15 ± 3 K/Ma. Rb-Sr yielded less precise ages than U-Pb, but using the new decay constant allows accurate comparison between the two methods. This study creates a firm foundation for future studies in thermal history of chondrites and terrestrial metamorphic complexes using Rb-Sr together with other isotopic chronometers.

The retinoblastoma tumor suppressor (pRb) is thought to orchestrate terminal differentiation by inhibiting cell proliferation and apoptosis and stimulating lineage-specific transcription factors. In this thesis I have shown that in the absence of pRb, differentiating primary myoblasts fused to form short myotubes that never twitched and degenerated via a non-apoptotic mechanism. The shortened myotubes exhibited an impaired mitochondrial network, mitochondrial perinuclear aggregation, autophagic degradation and reduced ATP production. Bcl-2 and autophagy inhibitors restored mitochondrial function and rescued muscle degeneration, leading to twitching myotubes that expressed normal levels of muscle-specific proteins and eventually exited the cell-cycle. A hypoxia-induced glycolytic switch also rescued the myogenic defect after chronic or acute inactivation of Rb in a HIF-1-dependent manner. These results demonstrate that pRb is required to inhibit apoptosis in myoblasts and autophagy in myotubes but not to activate the differentiation program. I next tested the effect of retinoblastoma protein family members – p107 and p130 – on skeletal myogenesis in the absence of Rb. Chronic or acute inactivation of Rb plus p130 or Rb plus p107 increased myoblast cell death and reduced myotube formation, yet expression of Bcl-2, treatment with autophagy antagonist or exposure to hypoxia extended myotube survival, leading to long, contracting myotubes that appeared indistinguishable from control myotubes. Triple mutations in Rb family genes further accelerated cell death and led to elongated myocytes or myotubes containing two nuclei, some of which survived and twitched under hypoxia. Whereas nuclei in Rb-/- myotubes were unable to stably exit the cell-cycle, myotubes lacking both p107/p130 became permanently post-mitotic, suggesting that pRb, but not p107 or p130 may be lost in cancer because of the unique requirement for cell-cycle exit during terminal differentiation. This thesis demonstrates that pRb is required to inhibit apoptosis in myoblasts and autophagy in myotubes but not to activate the differentiation program, and reveal a novel link between pRb and cell metabolism.

The 87Rb-86Sr system is a widely used long-lived isotope geochronometer. 87Rb, the naturally occurring radioactive isotope of Rb, undergoes beta-decay to stable 87Sr with a half-life of approximately 50 Ga. Decay of 87Rb to 87Sr results in variable 87Sr/86Sr in minerals with different Rb/Sr, and measurement of 87Rb/86Sr and 87Sr/86Sr allows for the determination of the age of the rock. Accurate ages depend both on the quality of the isotopic analysis and on the accuracy of the 87Rb decay constant, lambda87. Although the currently accepted value for lambda87 of 1.42 × 10-11a-1 has been in use for over 30 years, there is growing evidence that it is not accurate. Recent attempts to refine lambda87 and its precision have not reached a consensus. This thesis describes a new experiment to measure lambda87 by 87Sr accumulation over a period of about 30 years, and the preparation of a 84-86Sr double-spike in conjunction with that experiment. Radiogenic 87Sr produced in aliquots of a RbClO4 salt was measured by isotope dilution thermal ionization mass spectrometry. An average of 31 measurements yields a value of 1.398 ± 0.003 × 10-11a-1 . This requires a substantial revision from the previously accepted decay constant and makes Rb-Sr ages calculated with it 1.5% older. A Rb-Sr and U-Pb isotopic chronometry study was carried out on thirteen ordinary chondrites – the most common type of meteorite, the origin and history of which are still unclear. Some meteorites appear disturbed, possibly by recent shock during breakup of the parent body, whereas others yielded accurate and precise U-Pb and Pb-Pb ages. For example, L5 Elenovka yielded distinct ages for silicates (4555 Ma) and phosphates (4535 Ma), allowing the cooling rate of this meteorite from approximately 1055 K to 759 K to be constrained to 15 ± 3 K/Ma. Rb-Sr yielded less precise ages than U-Pb, but using the new decay constant allows accurate comparison between the two methods. This study creates a firm foundation for future studies in thermal history of chondrites and terrestrial metamorphic complexes using Rb-Sr together with other isotopic chronometers.

The retinoblastoma tumor suppressor (pRb) is thought to orchestrate terminal differentiation by inhibiting cell proliferation and apoptosis and stimulating lineage-specific transcription factors. In this thesis I have shown that in the absence of pRb, differentiating primary myoblasts fused to form short myotubes that never twitched and degenerated via a non-apoptotic mechanism. The shortened myotubes exhibited an impaired mitochondrial network, mitochondrial perinuclear aggregation, autophagic degradation and reduced ATP production. Bcl-2 and autophagy inhibitors restored mitochondrial function and rescued muscle degeneration, leading to twitching myotubes that expressed normal levels of muscle-specific proteins and eventually exited the cell-cycle. A hypoxia-induced glycolytic switch also rescued the myogenic defect after chronic or acute inactivation of Rb in a HIF-1-dependent manner. These results demonstrate that pRb is required to inhibit apoptosis in myoblasts and autophagy in myotubes but not to activate the differentiation program. I next tested the effect of retinoblastoma protein family members – p107 and p130 – on skeletal myogenesis in the absence of Rb. Chronic or acute inactivation of Rb plus p130 or Rb plus p107 increased myoblast cell death and reduced myotube formation, yet expression of Bcl-2, treatment with autophagy antagonist or exposure to hypoxia extended myotube survival, leading to long, contracting myotubes that appeared indistinguishable from control myotubes. Triple mutations in Rb family genes further accelerated cell death and led to elongated myocytes or myotubes containing two nuclei, some of which survived and twitched under hypoxia. Whereas nuclei in Rb-/- myotubes were unable to stably exit the cell-cycle, myotubes lacking both p107/p130 became permanently post-mitotic, suggesting that pRb, but not p107 or p130 may be lost in cancer because of the unique requirement for cell-cycle exit during terminal differentiation. This thesis demonstrates that pRb is required to inhibit apoptosis in myoblasts and autophagy in myotubes but not to activate the differentiation program, and reveal a novel link between pRb and cell metabolism.

Biological hydrogen production through integration of dark and photo-fermentation by using biomass is a promising alternative for energy supply problems. The main purpose of this study was to investigate photobiological H2 production by the purple non-sulfur (PNS) bacteria Rb. capsulatus on dark fermentation effluent of sugar beet thick juice (DFESBTJ). Presence of NH4+ in effluents is an important parameter since NH4+ inhibit the nitrogenase enzyme activity. Therefore, the influence of different NH4+ concentrations in the DFESBTJ by removing using natural zeolite clinoptilolite on photofermentative H2 production were studied using Rb. capsulatus DSM1710 and Rb. capsulatus YO3 (hup-). Also, the effect of EtOH concentrations (between 6.25 and 200 mM) in the defined medium on H2 production were studied using both bacterial strains since EtOH is a possible by-product of dark fermentation process. The experiments were carried out in small scale bottle photobioreactors (PBRs) and outdoor panel PBR (4 L). H2 productivity of 1.12 mmol/Lc/h was attained over 15 days of operation for panel PBR. The results showed that the zeolite was effective in removing NH4+ from the DFESBTJ as its concentration decreased by 95% after treatment. In both bacterial strains, an increase in the maximum productivities and molar H2 yields was observed with the decrease in NH4+concentrations. There was no significant effect of EtOH on H2 production except the inhibition at 200 mM. The main conclusions were that both bacterial strains could effectively utilize the DFESBTJ for growth and H2 production, therefore facilitating the integration of the dark and photo-fermentation for sustainable biohydrogen production.

Enterohaemorrhagic E. coli (EHEC) are important gastrointestinal pathogens of humans. E. coli serotype O157:H7 is the EHEC most commonly associated with human illness. E. coli O157:H7 is carried asymptomatically by cattle which form an important reservoir for the bacterium. E. coli O157:H7 has been found to colonise at the terminal rectum of cattle in preference to other sites in the bovine gastrointestinal tract. The first objective of this work was to characterise the roles of bacterial secreted components responsible for key functions in the modulation of host defences against EHEC. Data presented here reaffirms the role of flagellin in the elicitation of a proinflammatory response in a cultured human epithelial cell line; however, the response of a bovine epithelial cell line to bacterial secreted products was not affected by the presence or absence of flagellin. A role in the modulation of the host response for the StcE protease was also investigated: although its role in interaction with the bovine host was not established, bovine secretory antibodies to StcE were detected in rectal mucosal scrapings from an E. coli O157:H7-challenged calf, suggesting that StcE is expressed and recognised in vivo. The second key objective was to isolate E. coli from the bovine intestinal tract in order to define the colonisation patterns of E. coli within the bovine intestinal tract and relate this to bacterial genotype and to provide bovine E. coli isolates to test for inhibitory activity against E. coli O157:H7 which may yield bacteria with potential as probiotic agents with a view to reducing the prevalence of EHEC in cattle. Genotypic analysis of bovine resident E. coli confirmed that these strains carry a variety of virulence factor-encoding genes; however, certain dominant genotypes were identified and the genomic structure of representative isolates was predicted by genomic microarray. EHEC-related genotypes were found to be positively associated with colonisation at the rectum, whereas non-EHEC genotypes were found to colonise multiple intestinal sites without showing any apparent site-specificity. The third and final objective of this analysis was to carry out genotypic analysis of Scottish EHEC strains in order to predict whether increased incidence of EHEC infection in Scotland may be related to the presence of EHEC strains carrying altered complement of virulence factor-encoding genes. The analysis of EHEC isolated in Scotland revealed that these strains exhibit a genomic profile which is largely typical of EHEC isolated elsewhere, although there were certain differences in the carriage of a certain genomic elements. The results presented here support the proposal that bacteriophages are the key mediators of genetic variability among E. coli isolates.

Depression is one of the most common causes of periods of disability. There is evidence suggesting that the serotonin system is involved in the pathophysiology of depression. It has been suggested that synaptic serotonin levels are reduced in depressed patients, and that pharmacological blockade with antidepressants of the serotonin transporter (SERT) would result in alleviated symptoms of depression by enhancing serotonin neurotransmission. Since depression can be treated with antidepressants that target SERT, and a recently discovered 5-HTT gene-linked polymorphic region (5-HTTLPR) of the SERT gene has been shown to predispose to depression, the SERT assumes a key role in depression. Traditionally, depression severity was assessed using psychological testing of patients. However in the last 20 years, neuroimaging techniques using magnetic resonance imaging (MRI) of brain structures and molecular single photon emission computed tomography (SPECT) evolved which appear promising to better understand the pathophysiology at the tissue level. However, preclinical data on abnormalities that involve the serotonin system are limited. The studies presented in this thesis attempt to shed more light on the feasibility of using the novel MRI technique diffusion tensor imaging (DTI), and SPECT to detect disturbances of the serotonin system. Firstly, in order to elucidate the capabilities of DTI as a research tool in the detection of conceivably mild changes in white matter involving the serotonin system, a mouse model of life-long SERT deficiency was studied. Secondly, in order to validate DTI image processing methodology, a mouse model with reportedly profound myelin dysfunction was examined. Histology techniques were applied to the same mouse brains in order to explore the tissue correlate of the DTI signal changes. Thirdly, as myelin was hypothesised to interact with the serotonin system, in vitro autoradiography of SERT in mice with widespread hypomyelination was conducted in order to test this hypothesis. Lastly, in a rat model of SERT depletion, the relative abilities of a well established SPECT radioligand, [125I]βCIT (2β-carbomethoxy-3β-(4-iodophenyl)tropane), and a relatively novel SERT tracer, [123I]ADAM (2-((2-((dimethylamino)methyl)phenyl)thio)-5-iodophenylamine) were examined using micro-SPECT. The data demonstrate that DTI did not detect any changes in white matter organisation in SERT-deficient mice. Surprisingly, subtle changes in white matter microstructure were detected in mice that were haploinsufficient for SERT, i.e. heterozygous null mice, displaying a 50 % SERT reduction compared to WT as detected using DTI. On the other hand, profound hypomyelination was detected using DTI in another mouse model with white matter pathology, and correlations between DTI and histopathological markers were present, indicating that this technology provides good indications of severe pathology, while small changes, if present, may be missed. In addition, the SERT availability appeared not to be affected in mice with widespread hypomyelination. While post mortem autoradiography of SERT-depleted rats showed widespread reductions in SERT binding using dedicated specific SERT ligands, micro-SPECT using [125I]βCIT and [123I]ADAM did not show any differences. [125I]βCIT delivered good quality brain SPECT images, however analysis of [123I]ADAM scans was hampered by the poor definition of structures. Thus this thesis provides important information on the feasibility, and sensitivity of current neuroimaging modalities. In addition, methodological flaws and uncertainties in the current literature were identified, which underpins the need for improving and standardising methodological approaches, particularly in SPECT imaging.

The Large Eddy Simulation (LES) technique with the Smagorinsky-Lilly dynamic subgrid model and two-equation Standard k-ω Transitional turbulence model are applied to investigate non-spiral and spiral blood flow through three dimensional models of arterial stenosis and aneurysm. A spiral pattern of blood flow is thought to have many beneficial effects on hemodynamics. Previous computational studies on spiral blood flow involve only steady spiral flow in a straight stenosed pipe without considering an upstream curved section of the artery. But a spiral pattern in the blood flow may exist due to the presence of an upstream curved section in the artery. On the other hand, pressure is generally considered a constant quantity in studies on pulsatile flow through either arterial stenosis or aneurysm; however, blood pressure is a waveform in a physiological flow. Although cosine-type or smooth regular stenoses are generally taken in investigations of blood flow in a three-dimensional model of arterial stenosis, in reality, stenoses are of irregular shape. Besides stenosis and aneurysm, another abnormal condition of the artery is the presence of stenosis with an adjacent aneurysm in the same arterial segment, especially in the posterior circulation. A study on (steady or pulsatile) flow through such arterial stenosis with an adjacent aneurysm in the same arterial segment is not available so far. Therefore, taking above things into consideration, thorough investigations of steady and unsteady pulsatile non-spiral and spiral blood flow in three-dimensional models of stenosis and aneurysm are needed to give a sound understanding of the transition-to-turbulence of blood flow due to stenosis and aneurysm and to study the the effects of spiral velocity on the transition-to-turbulence. The LES technique has mostly been used to investigate turbulent flow in engineering fields other than bio-fluid mechanics. In the last decade, LES has seen its excellent potential for studying the transition-to-turbulence of physiological flow in bio-fluid mechanics. Though the k-ω Transitional model is used in few instances, mainly LES is applied in this study. Firstly, investigations of steady non-spiral and spiral blood flow through threedimensionalmodels of cosine-type regular stenosed tube without and with upstream curved segment of varying angles of curvature are performed by using the k-ω Transitional model and LES. A fully developed Poiseuille velocity profile for blood is introduced at the inlets of the models. To introduce a spiral effect at the inlet, onesixth of the bulk velocity is taken as the tangential velocity at the inlet along with the axial velocity profile there. Secondly, physiological pulsatile non-spiral and spiral blood flow through a three-dimensional model of a straight tube having cosine-type regular stenosis are investigated by using mainly LES. A two-equation k-ω Transitional model is also used in one non-spiral flow case. The first four harmonics of the Fourier series of pressure pulse are used to generate physiological velocity profiles at the inlet. At the outlet, a pressure waveform is introduced. The effects of percentage of area reduction in the stenosis, length of the stenosis, amplitude of pulsation and Womersley number are also examined. Thirdly, transient pulsatile non-spiral and spiral blood flow through a threedimensional model of irregular stenosis are investigated by applying LES and comparison is drawn between non-spiral flow through a regular stenosis and that through an irregular stenosis. Lastly, pulsatile non-spiral and spiral blood flow through a three-dimensional model of irregular stenosis with an adjacent post-stenotic irregular aneurysm in the same arterial segment are studied by applying LES and the k-ω Transitional model. The effects of variation in spiral velocity are also examined. The results presented in this thesis are analysed with relevant pathophysioloical consequences. In steady flow through the straight stenosed tube, excellent agreement between LES results for Re = 1000 and 2000 and the corresponding experimental results are found when the appropriate inlet perturbations are introduced. In the models with an upstream curved segment, no significant effect of spiral flow on any flow property is found for the investigated Reynolds numbers; spiral pattern disappears before the stenosis – which may be due the rigid wall used in the models and/or a steady flow at the inlet. The effects of the curved upstream model can be seen mainly in the maximum turbulent kinetic energy (TKE), the maximum pressure drop and the maximum wall shear stress (WSS), which in the curved upstream models generally increase significantly compared with the corresponding results in the straight stenosed tube. The maximumcontributions of the SGS motion to the large-scale motion in both non-spiral and spiral flow through a regular stenosis, an irregular stenosis and an irregular stenosis with an adjacent post-stenotic irregular aneurysm are 50%, 55%and 25%, respectively, for the highest Reynolds number investigated in each model. Although the wall pressure and shear stress obtained from the k-ω Transitional model agree quite well with the corresponding LES results, the turbulent results obtained from the k-ω Transitional model differ significantly from the corresponding LES results – this shows unsuitability of the k-ω model for pulsatile flow simulation. Large permanent recirculation regions are observed right after the stenosis throat in both non-spiral and spiral flow, which in the model of a stenosis with an adjacent post-stenotic aneurysm are stretched beyond the aneurysm and the length of the recirculation regions increases with spiral velocity. This study shows that, in both steady and unsteady pulsatile flow through the straight tube model having either a stenosis (regular or irregular) or an irregular stenosis with an adjacent post-stenotic irregular aneurysm, the TKE rises significantly at some locations and phases if a spiral effect is introduced at the inlet of the model. However, the maximum value of the TKE in a high spiral flow drops considerably compared with that in a low spiral flow. The maximum wall pressure drop and shear stress occur around the stenosis throat during all the phases of the pulsatile cycle. In the model of a stenosis only, the wall pressure rises in the immediate post-stenotic region after its drop at the stenosis throat. However, in the model of a stenosis with an adjacent aneurysm, the wall pressure does not rise to regain its undisturbed value before the start of the last quarter of the aneurysm. The effects of the spiral flow on the wall pressure and WSS are visible only in the downstream region where they take oscillatory pattern. The break frequencies of energy spectra for velocity and pressure fluctuations from −5/3 power slope to −10/3 power slope and −7/3 power slope, respectively, are observed in the downstream transition-to-turbulence region in both the non-spiral and spiral flow. At some locations in the transition region, the velocity spectra in the spiral flow has larger inertial subrange region than that in non-spiral flow. The effects of the spiral flow on the pressure spectra is insignificant. Also, the maximum wall pressure drop, the maximum WSS and the maximum TKE in the non-spiral flow through the irregular stenosis rise significantly compared with the corresponding results in the non-spiral flow through the regular stenosis. When the area reduction in the stenosis is increased, the maximum pressure drop, the maximumWSS and the TKE rise sharply. As for the effects of the length of the stenosis, the maximum WSS falls significantly and the maximum TKE rises sharply due to the increase in the length of the stenosis; but the maximum pressure drop is almost unaffected by the increase in the stenosis length.