The availability and demand for proton therapy is rapidly expanding across the globe. One of the key decisions that must be made in the procurement process by all new centres is which proton treatment planning system (TPS) to purchase. The first topic of this thesis is therefore to evaluate the performance of three different proton TPSs in the planning of ten meningioma patients. The comparison is built upwards from the beam commissioning and attempts are made to make as many variables as possible consistent between systems. Few statistically significant differences were found between the plans, although differences between the systems (such as layer spacing and spot positioning) are discussed. It is hoped this work will be of general use to the whole proton physics community and will encourage further development of proton TPSs from vendors. One of the major sources of range uncertainty in current proton treatment planning is due to the necessary conversion of the patient’s X-ray computed tomography (CT) dataset from CT numbers to relative stopping powers (RSPs). The remainder of the thesis looks to address this. The stoichiometric procedure is considered the most accurate method to generate the X-ray CT to RSP calibration curve. In the third chapter of the thesis an investigation is made into the errors of this procedure: specifically, the theoretical calculation of the RSP, step four of the process. The impact of these errors on the proton beam range is calculated for both phantom and patient cases. It has been suggested that proton radiography could offer a solution to the uncertainty in this calibration curve. The fourth chapter of the thesis therefore looks at a novel method of proton radiography, which involves taking the dose ratio of two pristine Bragg peaks. The investigation proceeds with a theoretical analysis of the application limits of the technique, together with an experimental validation of the theoretical approach. The fifth chapter of the thesis demonstrates an approach that uses proton radiography to improve the calibration curve. Assuming the information in the proton radiograph to be correct, the calibration curve can be optimised by comparison with a digitally reconstructed radiograph through the X-ray CT. The function of this optimiser is validated on synthetic datasets and its application is demonstrated with real measurements on plastic and real tissue phantoms. The technique is also shown to offer an improvement in the water-equivalent path length prediction at a therapeutic depth.

The inflammatory bowel diseases (IBD), Crohn’s disease and ulcerative colitis, are chronic inflammatory diseases predominantly affecting the gastrointestinal tract. They are of unknown aetiology; however, there is a significant genetic component. A multitude of studies have identified common genetic variants associated with these diseases; however, they only account for a proportion of the disease heritability. The study of large families with many affected individuals is theoretically a powerful method for identifying rare disease causing variants that might contribute to this ‘missing heritability’. To date, this approach has not been successfully employed in IBD owing, in part, to the moderate size of families identified. This thesis describes the ascertainment, phenotypic and genomic characterisation of two extended multi-plex Ashkenazi Jewish (AJ) kindreds with over 50 and 25 cases of IBD, respectively. An interrogation of the possible mechanisms underlying this familial aggregation highlighted a partial role for common disease associated variants. Linkage analysis was employed in an attempt to identify loci segregating with the disease. A novel method for reconstructing haplotype flow information across extended pedigrees is described and implemented in these kindreds demonstrating the absence of a single locus shared by all affected individuals. Exome sequencing a cohort of unrelated AJ individuals and cases from the families permitted the prioritisation of a number of candidate variants however, these did not reach an empirical significance thresh-old as defined using a Monte Carlo gene-dropping approach. It is possible that the analyses of these data in combination with those from additional families may yield significance. However it is clear that even in extended families with many affected individuals, the genetics of IBD remains complex.

A qualitative study was conducted amongst 57 practising Catholics in Spain: students, priests, and contemplative monks and nuns. Through semi-structured interviews, participant observation and ethnography their understanding of severe sadness and the difference with pathological sadness, coping mechanisms and help-seeking behaviour were explored. The participants clearly differentiated between sadness in response to a cause, sadness that “made sense”, and cases where sadness was not explained by the context, sadness that “did not make sense”. The former was seen as a normal reaction to adversity which should be resolved by the individuals’ social, cultural and religious resources, while the latter was likely to be conceptualised as pathological, along the lines of depression, warranting psychiatric consultation. It was also found that religion played a crucial role in the way sadness was understood and resolved: symptoms that otherwise might have been described as evidence of a depressive episode were often understood in those more religiously committed within the framework of the “Dark Night of the Soul” narrative, an active transformation of emotional distress into a process of self-reflection, attribution of religious meaning and spiritual growth. A complex portrayal of the role of the spiritual director and the parish priest in helping those undergoing sadness and depression emerged, containing positive aspects and criticisms of some priests’ lack of commitment and mental health training. This study emphasises the importance of taking into account the context of depressive symptoms, as the absence of an appropriate context is seemingly what made participants conceptualise them as abnormal. It also warns about the risks of medicalising normal episodes of sadness and raises questions about the lack of face validity of the current diagnostic classification for depressive disorder, which exclusively uses descriptive criteria. The thesis concludes by making some suggestions regarding differentiating normal from pathological sadness and how to incorporate existential issues into clinical practice.

Background: Pakistanis are the second largest ethnic minority group in the UK. However, there is a paucity of sexual health research among this group. In contrast to their parents’ generation, other South Asians are beginning to have premarital (sexual) relationships. This change may be occurring among young Pakistanis. In this thesis I explore social and cultural influences on the relationships and sexual behaviours of Pakistani youth, in order to determine whether there is unmet sexual health need. Methods: I analysed data from the second National Survey of Sexual Attitudes & Lifestyles (Natsal-2), exploring attitudes, learning about sex and first sexual experiences among Pakistanis (n=365). I then carried out thirty in-depth interviews with young Pakistanis and community workers to explore key life issues and implications for relationships and sexual behaviour. Findings: In Natsal-2, Pakistanis reported more conservative sexual attitudes than other groups, e.g. many saw premarital sex as wrong. They were also more likely to be married at first sex. Yet two-thirds of Pakistani men also reported being in non-marital relationships at this time, suggesting attitudes are not necessarily consonant with behaviour. The in-depth interviews found that premarital relationships are commonplace among youth, but due to social and cultural constraints, are conducted in secrecy. Young Pakistanis strive to maintain individual and cultural reputation. There were gender differences in perceptions, relationship types and vulnerability to sexual health risk. Sexual health knowledge was poor and few had access to support. Conclusions: Contrary to faith and cultural norms, premarital relationships do occur among Pakistani youth. The secrecy of these relationships and the pressures and complex gender roles they negotiate mean that some young Pakistanis are at risk of poor sexual health. This has implications for the delivery of appropriate preventative and curative services, which should encompass a broad understanding of sexual health and risk.

Microglia have neurotoxic and neuroprotective effects. The aim of the current project was to investigate the effects of perineuronal microglial activation on axonal regeneration in adult rats and the effects of prolonged neuroinflammation on foetal mouse brain. In contrast to the PNS, the CNS only displays limited axonal regeneration after injury and little perineuronal inflammation. Inflammation around the cell bodies of axotomised neurons has been demonstrated to promote CNS regeneration. Polyinosinic:polycytidylic acid (Poly I:C) is an inflammatory agent. Following delivery of Poly I:C into the motor cortex and a concomitant C4 dorsal corticospinal tract (CST) injury, rats exhibited more CST axons in the cervical spinal cord and less retraction from the injury site than controls. Following facial nerve axotomy, Poly I:C injections adjacent to the facial nucleus accelerated functional recovery. Viral vectors carrying Granulocyte Macrophage Colony Stimulating Factor (GM-CSF) were injected into motor cortex. GM-CSF virus reduced retraction of corticospinal axons from a spinal cord injury site. Behavioural studies of forelimb movements showed that C4 injury had a greater impact on fine distal movements, particularly reaching and grasping, which are known to be controlled by the CST. Treating rats with GM-CSF virus showed a trend towards improved forelimb sub-movements and significantly aided the reaching function recovery. Perinatal activation of periventricular phagocytes has been suggested to result in white matter damage, causing persistent motor disabilities. Transuterine injections of control or GM-CSF virus targeting the lateral ventricles of mice at gestational day 14, resulted in a widespread virally-transduced cells. Greater numbers of phagocytic and activated microglia were present in the areas of viral transduction. There was increased inflammation in the periventricular white matter. Although, the level of transduction remained relatively constant with increasing time, inflammation decreased, suggesting that GM-CSF toxicity is high before or around birth.

Introduction: The true incidence of radiation toxicity to the bowel remains unknown; in the UK, it has been reported that about 90% of patients who receive pelvic radiotherapy will have some change in their bowel function, and in up to 50% this affects their quality of life significantly (Andreyev, 2007). It is unclear why some cancer survivors develop significant symptoms that arise as a result of multiple functional, structural and physiological deficiencies related to radiation injury. Aims: There is a need to identify tissue specific biomarkers of normal tissue injury and identify those patients who might be at risk of severe injury to the bowel. In this thesis, I sought to investigate the true incidence and presentation of RIBI in a London Cancer Centre. I then developed a template for a scoring model to explore how reporting of symptoms in the clinical setting might be improved. After investigating the use of cell-cycle markers as a marker of (chemo)-radiosensitivity, I then utilised these markers in colonic crypt cells to attempt to link the proliferative status after exposure to radiation to symptom presentation and severity. Methods and Results: A retrospective cohort study revealed 152 women treated for cervical and endometrial cancer with symptoms of RIBI, which were clustered into 3 groups using factor analysis. Exploratory and Confirmatory Factor Analysis was used to test a novel scoring model template. Immunostaining in 35 cervical tumour samples with the cell-cycle markers Mcm2, Geminin, and Ki67 did not find expression of these markers were linked to (chemo)-radiosensitivity and tumour response. These markers were used to assess proliferation in colo-rectal crypt cells and showed decreased expression in all layers suggesting a loss of proliferative capacity after radiation. Conclusions: Young patients with cervical cancer are more likely to develop significant symptoms of RIBI. Our simple scoring tool validated on a prospective cohort could provide invaluable data to improve management of women with bowel symptoms after radiation. Further work exploring proliferation in colonic crypt cells after radiation exposure could identify women at greater risk of radiation injury.

X-ray imaging is a non-destructive and preferred method in paleopathology to reconstruct the history of ancient diseases. Sophisticated imaging technologies such as computed tomography (CT) have become common for the investigation of skeletal disorders in human remains. The effects of CT exposure on ancient cells have not been quantitatively examined in the past and may be important for subsequent genetic analysis. To remedy this shortcoming, different Monte Carlo models were developed to simulate X-ray irradiation on ancient cells. Effects of mummification and physical processes were considered by using two sizes of cells and three different phantom tissues that enclosed the investigated cell cluster. This cluster was positioned at the isocentre of a CT scanner model, where the cell hit probabilities P(0,1,…, n) were calculated according to the Poisson distribution. To study the impact of the dominant physical process, CT scans for X-ray spectra of 80 and 120 kVp were simulated. The calculated risk of DNA damage according to the multi-hit, multi-target model revealed that the probability of two DNA hits was pNT = 0.001 with cell size 6x6x10 μm3 (NT6610) and pNT = 0.00033 with cell size 4x4x6 μm3 (NT446) for normal tissue (NT) at 80 kVp. A further decrease in DNA damage was observed with pNT = 0.0006 (NT6610) and pNT = 0.00009 (NT446) at 120 kVp. All values of p are in good agreement with those given by the X-ray risk of cancer. It is concluded that the probability of ancient DNA (aDNA) damage following CT imaging depends on the number and volume of fragments m with paDNA < pNT^m (m ≥ 1). Increasing the number of aDNA fragments m is associated with rapidly decreasing aDNA damage through X-ray imaging.

Stereotactic radiosurgery is an increasingly common treatment modality that uses very small photon fields. This technique imposes high dosimetric standards and complexities that remain unsolved. In this work the dosimetric performance of CMOS active pixel sensors is presented for the measurement of small-photons beams. A novel CMOS active pixel sensor called Vanilla developed for scientific applications was used. The detector is an array of 520 × 520 pixels on a 25 μm pitch which allows up to six dynamically reconfigurable regions of interest (ROI) down to 6 × 6 pixels. Full frame readout of over 100 frame/s and a ROI frame rate of over 20000 frame/s are available. Dosimetric parameters measured with this sensor were compared with data collected with ionization chambers, film detectors and GEANT4 Monte Carlo simulations. The sensor performance for the measurement of cross-beam profiles was evaluated for field sizes of 0.5 × 0.5 cm2. The high spatial resolution achieved with this sensor allowed the accurate measurement of profiles from one single row of pixels. The problem of volume averaging is solved by the high spatial resolution provided by the sensor allowing for accurate measurements of beam penumbrae and field size under lateral electronic disequilibrium. Film width and penumbrae agreed within 2.1% and 1.8%, respectively, with film measurement and better than 1.0% with Monte Carlo calculations. Agreements with ionization chambers better than 1.0% were obtained when measuring tissue-phantom ratios. The data obtained from this imaging sensor can be easily analyzed to extract dosimetric information. The results presented in this work are promising for the development and implementation of CMOS active pixel sensors for dosimetry applications.

This thesis studies the effect of hypoxia (at rest and during exercise) on the arterial and venous cerebral circulation, investigating the venous system role in high altitude headache. Methods: 1) Hypobaric hypoxic studies investigated 198 trekkers and 24 Investigators to 5300m, 14 to 6400m and 8 to 8848m. 2) Normobaric hypoxic studies used Magnetic Resonance Imaging (MRI)) at sea-level. Four domains were addressed: i. Arterial: Hypobaric hypoxia: (n=24) Transcranial Doppler (TCD) measured middle cerebral artery diameter (MCAD) and blood velocity (MCAv). Sea-Level normobaric hypoxia: (n=7) A hypoxicator (FiO2 = 11%) for 3 hours with a 3Tesla MRI scan measured MCAD and MCAv. ii. Brain Oxygenation: Near Infrared Spectroscopy (NIRS) monitored Regional Brain Oxygenation (rSO2). iii. Venous: Retinal imaging at altitude and MRI at sea-level assessed the venous system. iv. Headache: A daily diary recorded headache burden. Results: Arterial: Hypobaric and normobaric hypoxia induced MCA dilatation. Mean (±(SEM)) MCAD increased in hypoxia (from 5.23(±0.23)mm (at 5300m) to 9.34(±0.88)mm (at 7950m)(p<0.001) (TCD). At sea-level, (after 3 hours FiO2 = 11%) MCAD increased from 3.04(±0.13)mm to 3.27(±0.13)mm (MRI). Brain Oxygenation: rSO2 decreased more than peripheral arterial saturation (SaO2), especially during exercise. The relative percentage reduction in resting SaO2 and rSO2 from 75m to 5300m was -22.23 ±0.56% and -30.61 ±1.28% (p<0.001) respectively. Venous: Hypoxia induced retinal and cerebral venous distension. Twenty-three of 24 subjects exhibited retinal venous distension (range 5 to 44%). Degree of distension correlated with headache (r = 0.553, p=0.005). Possession of a narrow transverse sinus strongly related to retinal and cerebral venous distension and headache. Headache: Headache Severity Index (HSI) (headache score x duration) correlated inversely to both lateral and third ventricular volumes summed (r = -0.5, p = 0.005) and pericerebellar CSF volume (r = -0.56, p = 0.03). Conclusions: Large cerebral arteries dilate and veins distend with hypoxia. This suggests an important influence of cerebral venous anatomy and physiology on headache, with implications for pathophysiological states and their management.

Coagulation proteinases, such as thrombin, exert a plethora of cellular effects via activation of proteinase-activated receptors (PARs). Abnormal activation of the coagulation cascade has been widely implicated in the pathology of fibroproliferative lung disease, including the most common form idiopathic pulmonary fibrosis (IPF). The high-affinity thrombin receptor PAR-1 is over-expressed in the lungs of IPF patients, particularly by the hyperplastic epithelium and fibroblasts within fibrotic foci. Moreover, viral infections are implicated in aberrant coagulation activation and accumulating evidence points to viruses as contributory factors to acute exacerbations that lead to deterioration of lung function in IPF patients. This thesis focuses on investigating and extending our current understanding of the role of PAR-1 in driving lung injury and fibrosis. The data show that primary human lung fibroblasts and microvascular endothelial cells express abundant PAR-1 and elicit robust calcium signalling responses following thrombin stimulation. In contrast, normal human alveolar and bronchial epithelial cells express low levels of PAR-1 but high levels of PAR-2. Exposure of the alveolar epithelial cell line, A549, to TGFβ increases PAR-1 expression and functional responses but this was not seen in freshly isolated primary human alveolar and bronchial epithelial cells. The role of TGFβ and pro-coagulant signalling responses was also investigated in an in vivo model of viral infection on a background of pulmonary fibrosis. Herpesvirus infection in the fibrotic lung promoted robust inflammatory responses characterized by extensive pulmonary consolidation but did not lead to an increase in lung collagen deposition. Importantly, viral infection was associated with the local upregulation of key components of the extrinsic coagulation pathway. Although PAR-1 is known to contribute to lung injury and fibrosis in a single hit experimental model of pulmonary fibrosis by promoting the activation of latent TGFβ, systemic targeting of TGFβ and the coagulation cascade does not impact on inflammation induced by viral infection in the fibrotic lung.