Since the recognition (DSM-IV; American Psychiatric Association, 1994) of life-threatening illnesses as a stressor that can precipitate posttraumatic stress disorder (PTSD), research has focussed on the issue of PTSD following cancer. Although the utility of a trauma framework has been questioned, understanding symptoms associated with PTSD such as intrusive cognitions may be critical in understanding psychological distress in cancer patients. Research has found that cancer patients experience negative intrusive thoughts, which are associated with marked distress. However, studies have rarely explored the content or nature of intrusions. In addition to verbal intrusions, intrusive memories of illness have been reported in cancer patients. More recently, intrusive imagery has been found in populations of anxious patients and reported to have a causal role in the maintenance of anxiety. Based on the recognition of cancer as a protracted experience involving multiple stressors, future-oriented visual intrusions, as well as intrusive memories and thoughts may play a role in psychological functioning. Chapter 1 is an overview of the literature assessing the presence of posttraumatic stress and posttraumatic stress symptoms in cancer patients. Chapter 2 is a cross-sectional study which showed that anxious prostate cancer patients (N=65) were significantly more likely to report intrusive cognitions compared to matched non-anxious (N=65) prostate cancer patients. Intrusive cognitions were frequent, uncontrollable and associated with significant distress and maladaptive adjustment. Chapter 3 is a cross-sectional study (N=139), which showed that factors such as how patients appraise intrusive cognitions affects anxiety severity and intrusion-related distress, after controlling for intrusion frequency. Chapter 4 provides a discussion of the use of imagery in psychological therapy and how imagery has been used with cancer patients in therapy. Chapter 5 presents two single-case studies of cancer patients completing a short therapy for anxiety, imagery rescripting, aimed at reducing negative properties of intrusive cognitions whilst also alleviating anxiety and depression. The final chapter provides a general discussion of the thesis and presents ideas for future research.

This thesis focuses on the protein kinase N family of serine/threonine protein kinases. This comprises three isoforms, with kinase domains highly related to the protein kinase C family, but with distinctive Rho/Rac dependent regulation. The critical roles played by Rho/Rac in regulating the cytoskeleton and cell migration/invasion led to an investigation into the role of PKNs in this process. Initial work in the thesis derived from the observation that PKN1 translocated to an insoluble cytosolic compartment in response to hyperosmotic stress. This had been shown to be dependent upon Racl and 3-phosphoinositide dependent kinase (PDK1). The thesis describes the characterisation of the domain in PKN 1 responsible for the Rac-dependent hyperosmotic response. Through the use of PKNl/PKCzeta chimeras a 49 amino acid sequence within the kinase domain was shown to be necessary and sufficient for the observed osmotic behaviour. In developing a cell model for migration/invasion, a breast cancer cell model was identified that displayed high PKN expression. siRNA mediated depletion of PKN isoforms or inhibition of kinase activity, revealed a requirement for PKN in both migration and invasion. Comparison with other transformed cells indicated that the relative contribution each isoform makes towards these processes appears to be cell type dependent. As a candidate downstream target, PLD has been implicated in migration and invasion of breast cancer cells and so the previously described interaction between PKN and phospholipaseD in migrating breast cancer cells was investigated. Results suggest that the activity of PLD contributes to the migration of MDAMB-468 cells and that the production of phosphatidic acid in migrating cells is stimulated by PKN.

Blood vessel patterning during angiogenesis is a guided process. Endothelial tip cells located at the end of vascular sprouts are important for vessel guidance. One of the aims of my studies is to determine how endothelial tip cell formation is regulated. Previous studies have shown that the Notch ligand, Delta-like 4 (Dll4), is prominently expressed in tip cells, therefore suggesting a role of Notch signalling in its regulation. In the mouse, suppression of Notch signalling by pharmacological inhibition of γ-secretase or genetic deletion of one Dll4 allele resulted in excessive endothelial tip cell formation and a poorly-patterned, hyperdense vessel network. Induction of Notch signalling by Jagged1 peptide treatment led to a decrease in tip cell formation and a sparse vessel network. Thus, the Notch pathway negatively regulates tip cell formation to promote a non-sprouting endothelial stalk cell. The Notch-regulated ankyrin repeat protein, Nrarp, is a Notch target gene that is highly expressed in stalk cells and endothelial cells located at vessel branchpoints. Deletion of Nrarp in mouse and morpholino knockdown of nrarp-a and nrarp-b in zebrafish resulted in decreased endothelial cell proliferation and junctional instability, leading to ectopic vessel regression and the formation of a sparse vessel network. In endothelial cells, loss of Nrarp function increased Notch signalling but decreased Wnt signalling, thereby confirming its role as a negative and positive modulator of Notch and Wnt signalling, respectively. Mice null for Lymphoid enhancer factor 1 (Lef1) or deficient in endothelial Ctnnb1 also displayed increased vessel regression and decreased vessel density, suggesting a role of canonical Wnt/β-catenin signalling in maintaining vessel stability. In conclusion, my studies show that Notch signalling regulates endothelial tip cell formation and vessel stability to fine-tune vessel patterning. Furthermore, I demonstrate that Nrarp provides a molecular link to Wnt signalling to regulate vessel stability.

Protein Kinase C (PKC) comprises a family of phospholipid-dependent Ser/Thr kinases, implicated in a broad array of cellular processes. PKC activity is subject to a complex network of regulatory inputs, including co-factor binding, phosphorylation and protein-protein interaction. In addition, the chronic activation of PKC frequently leads to its down-regulation this process may be intrinsic to the tumour promoting activity of the phorbol esters. The aim of this work was to investigate the regulation of the novel PKC-e isoform by binding partners and phosphorylation, with a particular focus on the process of agonist-induced degradation. A yeast 2-hybrid screen was performed using a PKC-e bait and two novel binding partners were identified, both with associations to the ubiquitin/proteasome system VHL Binding Protein 1 (VBP1) and F-box WD40 protein 7 (Fbw7). Interactions were verified in mammalian cells and mapped to the catalytic domain of PKC-e. Extensive studies revealed that neither partner influenced the process of PKC-e down-regulation. However, Fbw7a was demonstrated to represent an in vitro PKC substrate. The site of phosphorylation was mapped to Ser-18 and, using phospho-specific antibodies, was shown to be phosphorylated in the cell. PKC-e activity is required for its agonist-induced degradation. Studies were therefore undertaken to investigate autophosphorylation, which may be implicated in this process. Serine residues 234, 316 and 368 were identified as novel PKC-e autophosphorylation sites. Using phospho- specific antibodies, all three sites were shown to be occupied in response to PKC-e activation. Phosphorylation at these sites was found not to influence agonist-induced PKC-e down-regulation. However, a critical role was established for phosphorylated Ser-368, in the recruitment of the PKC-e binding partner, 14-3-3(3. Together these findings provide insight into the mechanisms controlling PKC-e activity and demonstrate a relationship between regulation through phosphorylation and protein-protein interaction.

The treatment of pancreatic cancer is challenging. Patients are often beyond curative surgical therapy and palliative treatment with chemotherapy provides limited benefit. New markers of cancer activity and therapeutic targets are required. This thesis has firstly reviewed the available literature on Tumour M2-PK, a dimeric form of M2 isoenzyme of pyruvate kinase, in GI cancer and carried out a meta-analysis of the clinical data on pancreatic cancer. Experimental work evaluated the measurement of M2-pyruvate kinase in human pancreatic cancer cell lines with altered microenvironment (hypoxia, acidic pH or glucose-deprived condition). Tumour M2-PK level was measured using ELISA, total M2-PK by immunoblotting and pyruvate kinase activity by spectrophotometric analysis. Apoptosis or necrosis was detected by measuring active Caspase 3/7 and 8, Bcl-2, Bax and Annexin V staining. Localisation of M2-PK in pancreatic cancer cell was studied by immunocytochemistry. The clinical review has shown that Tumour M2-PK is not an organ-specific marker of GI cancer but is elevated with positive predictive value of 86–88% in gastro-oesophageal and colorectal cancers. In pancreatic cancer the diagnostic odds ratio (DOR) of an elevated Tumour M2-PK was similar to those of CA19-9 with overall sensitivity of 94% and specificity of 55%. Higher levels of Tumour derived M2-pyruvate kinase were observed in Colo 357 cell lines compared to Panc-1 cells. Exposure of Colo 357 cells to altered culture conditions resulted in decreased cell proliferation accompanied by elevated Tumour M2-PK levels with unchanged total M2-PK levels suggesting tetramerdimer switch-over, which was confirmed by the corresponding change in the pyruvate kinase activity. No correlation of Tumour M2-PK level or PK activity with apoptotic or anti-apoptotic markers was observed. Immunocytochemistry suggested M2-PK localisation to intracellular membrane-bound structures with no translocation to nucleus or mitochondria under altered tumour microenvironment. Conclusion: Tumour M2-PK is a potential marker of pancreatic cancer. Altering the tumour microenvironment causes a switch to measured M2-PK levels. This may allow cells to overcome cell apoptosis and could be a pathway facilitating tumour survival.

Survival from cancer is becoming a reality for more people in the world each year. Survival rate from colorectal cancer disease is approximately 80% one year after diagnosis, but falls to 62% at 5 years from diagnosis. Quality of life research in colorectal cancer to date has focused on investigating patients' experience during the diagnostic or treatment phase while the experiences of those who have survived this cancer have been ignored. Based on the concept of Health-Related Quality of Life (HRQOL) this study was focused on understanding and assessing the impact of colorectal cancer disease and its treatment on Greek patients' HRQOL over time. Also, this study sought to identify multiple factors (related either to patient or disease characteristics) that contributed to patients' HRQOL in both specific and general domains. Age, gender, stage at diagnosis, time elapse since diagnosis, income, education, colostomy appliance, disease recurrence, depression and communication between couples were examined for their effect on HRQOL over time. 145 Greek outpatients (male 87, female 58) completed the Functional Assessment of Cancer Therapy-Colorectal (FACT-C) Quality of Life Instrument and the Mental Component of the Short Form 36 Health Survey questionnaire measuring both generic and disease-specific HRQOL, as well as the Center for Epidemiologic Studies Depression Scale (CES-D) to detect depressive symptoms, and the Enriching & Nurturing Relationship Issues, Communication & Happiness (ENRICH) scale to assess communication between couples, at an interval of either one year or more than 5 years since diagnosis. Statistical significance was set at (p<0.05) and data were analysed using the Statistical Package for the Social Sciences (SPSS). The findings suggest that patients who survived colorectal cancer experienced an overall high quality of life independent of the stage of disease at diagnosis or time since diagnosis. Among those factors that had a negative effect on patients overall HRQOL over time depression was the most prevalent. Stoma patients experienced an overall lower HRQOL than non-stoma patients. Most domains of HRQOL assessment such as physical, emotional and role functioning of stoma patients were negatively affected, but these did not reach statistical significance. Notably, stoma patients in this sample showed significantly more dissatisfaction with body image than patients without a stoma – a finding that was more prevalent in women. This may suggest that stoma formation negatively affects sexual function and body image. Finally, patients with lower incomes and a recurrence or metastatic disease also experienced a poorer HRQOL. It is recommended that a practice-based strategy is developed in Greece to assess the HRQOL and psychosocial functioning of these patients as well as the recommendation that in the preoperative stage, after surgery and in the rehabilitation phase for stoma patients to be assessed and supported by a specialist Stoma Care nurse. Other suggestions for future research are also proposed.

No description available.

No description available.

No description available.

No description available.