Chi-Keung Leung. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (p. 138-146). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Six male long-distance runners participated in this study to evaluate the responses to exercise at 90% and 100% vV02max. Subjects participated in five maximal exercise tests: one incremental, three tests at 90% vV02max, and one test at 100% vV02max. The results of this study demonstrate that V02max can be elicited in a constant-velocity test at 90% vV02max.

Vaccination and the use of immune stimulants are two important ways to mitigate the costs of disease in fish aquaculture. A vaccine to Enteric redmouth disease (ERM) was the first licensed fish vaccine in the world. Although effective in protecting fish from the motile bacterial (Yersinia ruckeri) infection, ERM can occur in ERM vaccinated fish due to the rise of non-motile Y. ruckeri that does not express flagellin. This highlighted the need for continual improvement of vaccine efficacy and the importance of flagellin in fish immune responses. In this thesis the immune response to the ERM vaccine was studied first to give insights for vaccine development. A recombinant flagellin from Y. ruckeri (YRF) was then produced and its bioactivities were investigated in vitro and in vivo. The immune response to ERM vaccination was studied in rainbow trout in two major and relevant immune organs, the spleen and gills. Intraperitoneal injection of the ERM vaccine induces an early balanced expression of pro- and anti-inflammatory cytokines and adaptive cytokines in the spleen, with a heightened expression of acute phase proteins (APPs) and anti-microbial peptides (AMPs) in both spleen and gills. The analysis suggests that ERM vaccination activates host innate immunity and the expression of specific IL-12 and IL-23 isoforms leading to a Th1 and Th17 biased immune responses. This study has increased our understanding of the host immune response to ERM vaccination and the adaptive pathways involved. The early responses of a set of genes established in this study may prove useful as biomarkers in future vaccine development in aquaculture. YRF was next produced in a bacterial system, and purified. Its bioactivity was investigated first in the trout macrophage cell line RTS-11 and head kidney primary cell cultures. YRF is a potent activator of pro-inflammatory cytokines, APPs, AMPs and subunits of the IL-12 cytokine family in vitro. This property was further confirm in vivo in multiple tissues after intraperitoneal injection of YRF. These results suggest that flagellins are important pathogen-associated molecular patterns (PAMPs) that can activate an inflammatory response in fish not only in vitro but also in vivo. Furthermore, YRF was shown to be the most potent PAMP in vitro, in terms of activation of an inflammatory response, compared to pure LPS and peptidoglycan. In addition, YRF mixed with complete Freund's adjuvant can induce YRF-specific IgM antibodies in rainbow trout. These antibodies are able to neutralize YRF bioactivity, and react against the middle domain of YRF, as assessed in Western blot analysis. When YRF was fused with a protein antigen, it increased the antigen-specific IgM antibody response. This analysis reveals that YRF is a potent activator of host immune responses and can be used as an immune stimulant and adjuvant to improve vaccine efficacy.

A fundamental question in the study of memory is: Why do we remember some events but forget others? It has been proposed that people preferentially remember motivationally relevant information, as these memories may be useful in guiding choices in the future, a framework called adaptive memory. This dissertation examined the brain mechanisms that support adaptive memory, specifically focusing on how memory is shaped by rewards and dopamine, using a combination of pharmacological manipulations and behavioral assays. First, we found that rewards retroactively prioritize memory for preceding neutral events, and consistent with models of hippocampal replay, two periods of consolidation are necessary for this effect: a period of rest immediately following encoding and overnight consolidation. Second, motivated by research showing that neurotransmitters, such as dopamine, potentiate motivationally relevant memories to endure over long durations, we administered d-amphetamine (a dopamine agonist) before encoding. We found that when hippocampus dependent memory is tested after a short delay, working memory best accounts for memory performance, but when tested after a long delay, d-amphetamine level directly predicts memory performance. And third, we tested how d-amphetamine modulates different memory systems after a delay, using two different behavioral paradigms in which participants learned about overlapping associations using either stimulus-response learning or deliberate associative encoding. In both experiments, we found that d-amphetamine during encoding enhanced test performance on the trained items a week later; however, we did not detect any evidence that d-amphetamine modulates the integration of the overlapping pairs. Together, the work reported in this dissertation suggests that memory for motivationally relevant information is prioritized, dopamine enhances performance across different memory and learning systems, the effect of both reward and dopamine on memory and learning emerge after consolidation, and dopamine does not bias the hippocampus to encode memories in an integrated manner.

Lee Ka Wai Sharon. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 227-260). / Abstracts in English and Chinese. / ACKNOWLEDGEMENTS --- p.I / ABBREVIATIONS --- p.III / ABSTRACT --- p.VI / 摘要 --- p.XI / CONFERENCE PUBLICATIONS --- p.XVII / TABLE OF CONTENTS --- p.XVIII / Chapter Part I - --- General Introduction / Chapter Chapter 1: --- Introduction --- p.1 / Chapter 1.1. --- The Search of Immunomodulatory Agents --- p.1 / Chapter 1.2. --- Cordyceps sinensis (Dong Cong Xia Cao) as an Immunomodulatory Agent --- p.2 / Chapter 1.2.1. --- General Aspects --- p.2 / Chapter 1.2.2. --- Evidence from the Traditional Chinese Medicine Concepts --- p.2 / Chapter 1.2.3. --- Evidence from the Traditional Chinese Medicine Classics --- p.4 / Chapter 1.2.4. --- Evidence from the Modern Research Literature --- p.4 / Chapter 1.2.4.1. --- lmmunomodulation --- p.4 / Chapter 1.2.4.2. --- Anti-tumor Effects --- p.7 / Chapter 1.2.4.3. --- Other Activities Related to the Immune System --- p.8 / Chapter 1.2.4.4. --- Potential Active Ingredients: Cordycepin and Polysaccharides --- p.8 / Chapter 1.2.5. --- Prescription and Usage: Single Vs Concocted --- p.11 / Chapter 1.2.5.1. --- Single Form as an Immunoactivating Agent --- p.11 / Chapter 1.2.5.2. --- Concocted as an Anti-asthmatic Agent --- p.12 / Chapter 1.3. --- Our Hypothesis and Rationale --- p.13 / Chapter Chapter 2: --- Experimental Design --- p.24 / Chapter 2.1. --- General Aspects of the Human Immune System --- p.24 / Chapter 2.2. --- Designing the In vitro Study on Cell-mediated Immunity --- p.24 / Chapter 2.2.1. --- T Cells --- p.25 / Chapter 2.2.2. --- Macrophages --- p.26 / Chapter 2.3. --- Designing the In vitro and In vivo Study of Anti-tumor Activities --- p.29 / Chapter 2.3.1. --- Tumor Biology --- p.29 / Chapter 2.3.2. --- Tumor and Immunity --- p.29 / Chapter 2.3.2.1. --- T-Cell-Mediated Cytolysis (Tc Cells) --- p.30 / Chapter 2.3.2.2. --- Delayed-Type Hypersensitivity (TDth Cells) --- p.30 / Chapter 2.3.2.3. --- Natural Killer (NK) Cells --- p.30 / Chapter 2.3.2.4. --- Lymphokine-Activated Killer (LAK) Cells --- p.31 / Chapter 2.3.2.5. --- Antibody-Dependent Cell-Mediated Cytotoxic (ADCC) Cells --- p.31 / Chapter 2.3.2.6. --- Activated Macrophages (AMΦ) --- p.31 / Chapter 2.3.3. --- Mechanism of Tumor Engulfment --- p.32 / Chapter 2.3.4. --- The Experimental Plan --- p.33 / Chapter 2.4. --- Designing the In vitro Study and Clinical Trials on Anti-asthmatic Activities --- p.36 / Chapter Part II - --- Methodology / Chapter Chapter 3: --- Materials and Methods / Chapter 3.1. --- List of Materials and Their Origin --- p.39 / Chapter 3.1.1. --- Traditional Chinese Medicine --- p.39 / Chapter 3.1.2. --- Cells for In vitro Experiments --- p.39 / Chapter 3.1.3. --- Mice for In vivo Experiments --- p.40 / Chapter 3.1.4. --- "Medium, Buffer, Supplements and Reagents for Cell Culture" --- p.40 / Chapter 3.1.5. --- Dye for Cellular Staining --- p.40 / Chapter 3.1.6. --- Cell Mitogens and Activator --- p.41 / Chapter 3.1.7. --- Reagents for Flow Cytometric Analysis --- p.41 / Chapter 3.1.8. --- Reagent Kits --- p.41 / Chapter 3.1.9. --- ELISA Kits --- p.42 / Chapter 3.1.10. --- Antibodies --- p.43 / Chapter 3.1.11. --- Reagents for RNA Extraction --- p.44 / Chapter 3.1.12. --- Reagents for Gel Electrophoresis --- p.44 / Chapter 3.1.13. --- Reagents for cDNA Expression Array --- p.44 / Chapter 3.1.14. --- Other Reagents --- p.45 / Chapter 3.1.15. --- Special Equipment and Apparatus --- p.45 / Chapter 3.2. --- Details of Materials --- p.46 / Chapter 3.2.1. --- Traditional Chinese Medicine --- p.46 / Chapter 3.2.1.1. --- Natural Cordyceps sinensis --- p.46 / Chapter 3.2.1.2. --- HERBSnSENSEŚёØ Cordyceps --- p.46 / Chapter 3.2.1.3. --- Wheeze-Relief Formula --- p.46 / Chapter 3.2.2. --- "Media, Supplements and Reagents for Cell Culture" --- p.47 / Chapter 3.2.2.1. --- Cell Culture Media --- p.47 / Chapter 3.2.2.2. --- Serum Supplements --- p.47 / Chapter 3.2.2.3. --- Anti-CD16 Magnetic Microbeads --- p.47 / Chapter 3.2.2.4. --- Fico´HёØ-Paque Plus Solution --- p.47 / Chapter 3.2.2.5. --- PercolĺёØ Solution --- p.48 / Chapter 3.2.2.6. --- Phosphate Buffered Saline (PBS) --- p.48 / Chapter 3.2.2.7. --- Water --- p.48 / Chapter 3.2.3. --- Dye for Cellular Staining --- p.48 / Chapter 3.2.3.1. --- HemacoloŕёØ for Microscopy --- p.48 / Chapter 3.2.3.2. --- Trypan Blue Dye --- p.49 / Chapter 3.2.4. --- Reagents for Flow Cytometry --- p.49 / Chapter 3.2.4.1. --- FACS Flow Sheath Fluid --- p.49 / Chapter 3.2.4.2. --- FACS Wash Medium --- p.49 / Chapter 3.2.4.3. --- Paraformaldehyde --- p.49 / Chapter 3.2.5. --- Special Equipments and Apparatus --- p.49 / Chapter 3.2.5.1. --- Magnetic Cell Sorting System (MACS) --- p.49 / Chapter 3.3. --- Human Subjects --- p.51 / Chapter 3.3.1. --- Inclusion Criteria --- p.51 / Chapter 3.3.2. --- Exclusion Crtieria --- p.51 / Chapter 3.3.3. --- Medication --- p.52 / Chapter 3.3.4. --- Informed Consent and Patient Information --- p.52 / Chapter 3.4. --- Animals --- p.53 / Chapter 3.4.1. --- Maintenance --- p.53 / Chapter 3.4.2. --- Survival Experiment Using Erhlich Ascites Tumor Bearing ICR Mice --- p.53 / Chapter 3.4.3. --- Experiments of Immunomodulatory activity in Sarcoma 180 Bearing BALB/c Mice --- p.54 / Chapter 3.5. --- Methodology --- p.55 / Chapter 3.5.1. --- Preparation of the Traditional Chinese Medicine --- p.55 / Chapter 3.5.1.1. --- Hot Water Extraction of Water Soluble Fraction of Natural Cordyceps sinensis --- p.55 / Chapter 3.5.1.2. --- Hot Water Extraction of Water Soluble Fraction of HERBSnSENSEŚёØ Corydceps and the Wheeze-relief Formula for In vitro Experiments --- p.55 / Chapter 3.5.1.3. --- HERBSnSENSEŚёØ Corydceps for the In Vivo Experiments --- p.56 / Chapter 3.5.1.4. --- Extraction Efficiency of the Hot Water Extracts --- p.56 / Chapter 3.5.2. --- Limulus Ameobocyte Lysate Test --- p.56 / Chapter 3.5.3. --- Cell Preparation --- p.57 / Chapter 3.5.3.1. --- "Isolation of Human Peripheral Blood Mononuclear Cells, Lymphocytes and Monocytes" --- p.57 / Chapter 3.5.3.2. --- Isolation of Eosinophils --- p.58 / Chapter 3.5.3.3. --- Isolation of Spleen Cells from BALB/c Mice --- p.58 / Chapter 3.5.3.4. --- "Murine Ehrlich Ascites Tumor (EAT), PU5-18, and Sarcoma 180 (SC-180) Cell Lines" --- p.59 / Chapter 3.5.3.5. --- Human Eosinophilic Leukemic Cell Line (EoL-1) --- p.59 / Chapter 3.5.3.6. --- Human Hepatocarcinoma Hep-3B Cell Line --- p.59 / Chapter 3.5.3.7. --- Human Leukemic Cell Line (HL-60) --- p.59 / Chapter 3.5.3.8. --- Human Mast Cell Line (HMC-1) --- p.60 / Chapter 3.5.4. --- Collection of Mouse Serum and Human Plasma --- p.60 / Chapter 3.5.5. --- Collection of Culture Supernatant --- p.60 / Chapter 3.5.6. --- The Trypan Blue Exclusion Assay --- p.61 / Chapter 3.5.7. --- Colorimetric 5-bromo-2'-deoxyuridine (BrdU) Cell Proliferation Enzyme Linked Immunosorbent Assay (ELISA) --- p.61 / Chapter 3.5.8. --- Immunophenotyping --- p.62 / Chapter 3.5.9. --- The Cytometric Bead Array (CBA) Kits --- p.62 / Chapter 3.5.10. --- Intracellular Florescence Staining for Reactive Oxygen Species --- p.63 / Chapter 3.5.11. --- The Intracellular Zymosan Florescence Assay --- p.64 / Chapter 3.5.12. --- Total Cellular RNA Extraction --- p.64 / Chapter 3.5.13. --- Gel Electrophoresis of RNA Integrity --- p.65 / Chapter 3.5.14. --- cDNA Expression Array --- p.65 / Chapter 3.5.15. --- Cell Staining Using Cytospin --- p.66 / Chapter 3.5.16. --- Annexin V-FITC/Propidium Iodide Apoptosis Detection --- p.66 / Chapter 3.5.17. --- Weighing the Spleen and Tumor --- p.67 / Chapter 3.5.18. --- Preparing Samples for the Eosinophilic Cationic Protein Fluoroenzymeimmunoassay --- p.67 / Chapter 3.5.19. --- Statistical Analysis --- p.67 / Chapter Part III - --- Results: Pre-functional Assays / Chapter Chapter 4: --- "Extraction, Endotoxin Measurement, In vitro Cytotoxicity Testing, and the Selection of Optimal Concentration" / Chapter 4.1. --- Extraction efficiency --- p.68 / Chapter 4.1.1. --- Introduction --- p.68 / Chapter 4.1.2. --- Results --- p.68 / Chapter 4.2. --- Endotoxin Level --- p.69 / Chapter 4.2.1. --- Introduction --- p.69 / Chapter 4.2.2. --- Results and Interpretation --- p.69 / Chapter 4.3. --- Cytotoxicity --- p.70 / Chapter 4.3.1. --- Introduction --- p.70 / Chapter 4.3.2. --- Results and Interpretation --- p.71 / Chapter 4.3.2.1. --- Peripheral Blood Mononuclear Cells (PBMC) --- p.71 / Chapter 4.3.2.2. --- Eosinophils --- p.72 / Chapter 4.4. --- The Optimal Concentration (OC) --- p.76 / Chapter 4.4.1. --- Introduction --- p.76 / Chapter 4.4.2. --- Results and Interpretation --- p.76 / Chapter Part IV- --- Results: Immunomodulatory Activities of Cordyceps sinensis as a Single Herb / Chapter Chapter 5: --- Mitogenic Activity --- p.80 / Chapter 5.1. --- Introduction --- p.80 / Chapter 5.2. --- Results --- p.80 / Chapter 5.3. --- Discussion --- p.81 / Chapter Chapter 6: --- Cytokines and Cytokine Receptors --- p.84 / Chapter 6.1. --- Introduction --- p.84 / Chapter 6.2. --- Results --- p.84 / Chapter 6.2.1. --- Effects of Natural Cordyceps sinensis and HERBSnSENSEŚёØ Cordyceps on the Induction of Cytokines from Lymphocytes --- p.84 / Chapter 6.2.1.1. --- TNFa --- p.84 / Chapter 6.2.1.2. --- IL-6 --- p.85 / Chapter 6.2.1.3. --- IL-10 --- p.85 / Chapter 6.2.2. --- Effects of Natural Cordyceps sinensis and HERBSnSENSEŚёØ Cordyceps on the Induction of Cytokines from Monocytes --- p.92 / Chapter 6.2.2.1. --- IL-1β --- p.92 / Chapter 6.2.2.2. --- IL-6 --- p.92 / Chapter 6.2.2.3. --- IL-10 --- p.97 / Chapter 6.2.2.4. --- TNFα --- p.97 / Chapter 6.2.3. --- Effects of Natural Cordyceps sinensis and HERBSnSENSEŚёØ Cordyceps on the Expression of Cytokine Receptor --- p.102 / Chapter 6.2.4. --- Effects of Natural Cordyceps sinensis and HERBSnSENSEŚёØ Cordyceps on the Gene Expression of Cytokines and Cytokine Receptors in Peripheral Blood Mononuclear Cells --- p.105 / Chapter 6.3. --- Discussion --- p.112 / Chapter Chapter 7: --- Macrophage Functions: Phagocytosis and Release of Reactive Oxygen Species (ROS) --- p.116 / Chapter 7.1 --- Introduction --- p.116 / Chapter 7.2. --- Results --- p.117 / Chapter 7.2.1. --- Phagocytosis --- p.117 / Chapter 7.2.2. --- Release of Reactive Oxygen Species (ROS) --- p.117 / Chapter 7.3. --- Discussion --- p.124 / Chapter Chapter 8: --- Apoptosis of Selected Cancer Cell Lines --- p.126 / Chapter 8.1. --- Introduction --- p.126 / Chapter 8.2. --- Results --- p.127 / Chapter 8.2.1. --- Differential Cytotoxic Effects of natural Cordyceps sinensis and HERBSnSENSEŚёØ Cordyceps on Various Cancer Cell Lines In vitro --- p.127 / Chapter 8.2.2. --- Differential Anti-Proliferative Effects of natural Cordyceps sinensis and HERBSnSENSEŚёØ Cordyceps on Various Cancer Cell Lines In vitro --- p.129 / Chapter 8.2.3. --- Differential Apoptotic Effects of Natural Cordyceps sinensis and HERBSnSENSEŚёØ Cordyceps on Various Cancer Cell Lines In vitro --- p.131 / Chapter 8.2.3.1. --- Peripheral Blood Mononuclear Cells --- p.131 / Chapter 8.2.3.2. --- Hepatocarcinoma Hep-3B --- p.131 / Chapter 8.2.3.3. --- Human Eosinophilic Leukemic Cell Line --- p.134 / Chapter 8.2.3.4. --- Human Mast Cell Line --- p.134 / Chapter 8.2.3.5. --- Human Leukemic Cell Line (HL-60) --- p.138 / Chapter 8.2.3.6. --- Murine Macrophages/Monocytes Cell Line PU5-18 --- p.138 / Chapter 8.2.3.7. --- Murine Erhlich Ascites Tumor (EAT) --- p.142 / Chapter 8.2.3.8. --- Murine Sarcoma 180 (SC-180) --- p.142 / Chapter 8.3. --- Discussion --- p.145 / Chapter Part V- --- Results: Immunomodulatory Activities of Cordyceps sinensis in Concoction / Chapter Chapter 9: --- The In vivo Animal Model --- p.147 / Chapter 9.1. --- introduction --- p.147 / Chapter 9.2. --- Results --- p.148 / Chapter 9.2.1. --- The ICR Mice Model --- p.148 / Chapter 9.2.1.1. --- In vivo Effects of Natural C. sinensis and HERBSnSENSEŚёØ Cordyceps on the Ascitic Fluid Production of ICR Mice --- p.148 / Chapter 9.2.1.2. --- Effects of Natural C. sinensis and HERBSnSENSEŚёØ Cordyceps on the Survival of Tumor-bearing ICR Mice --- p.149 / Chapter 9.3. --- The BALB/c Mice Model --- p.153 / Chapter 9.3.1. --- In vivo Effects of HERBSnSENSEŚёØ Cordyceps on Spleen and Tumor Weight --- p.153 / Chapter 9.3.2. --- Effects of HERBSnSENSEŚёØ Cordyceps on the Mitogenic Activities of Spleen Cells --- p.154 / Chapter 9.3.3. --- "In vivo Effects of HERBSnSENSEŚёØ Cordyceps on the Cell Surface Expression of CD3, CD4, and CD8" --- p.157 / Chapter 9.3.4. --- Effects of HERBSnSENSEŚёØ Cordyceps on the Cytokine Release from Cultured Spleen Cells --- p.161 / Chapter 9.3.4.1. --- TNFα --- p.161 / Chapter 9.3.4.2. --- IFNγ --- p.163 / Chapter 9.3.4.3. --- IL-2 --- p.163 / Chapter 9.3.4.4. --- IL-4 --- p.163 / Chapter 9.3.4.5. --- IL-6 --- p.167 / Chapter 9.3.4.6. --- IL-10 --- p.167 / Chapter 9.3.4.7. --- IL-12p70 --- p.167 / Chapter 9.3.4.8. --- Monocyte Chemoattractant Protein(MCP)-1 --- p.167 / Chapter 9.3.5. --- In vivo Effects of HERBSnSENSEŚёØ Cordyceps on the Cytokine Synthesis --- p.172 / Chapter 9.4. --- Discussion --- p.174 / Chapter Chapter 10: --- In vitro Studies on Eosinophils and Peripheral Blood Mononuclear Cells --- p.178 / Chapter 10.1. --- Introduction --- p.178 / Chapter 10.2. --- Results --- p.180 / Chapter 10.2.1. --- In vitro Effects of Wheeze-Relief Formula on the Survival of IL-5 Enhanced Eosinophils --- p.180 / Chapter 10.2.2. --- In vitro Effects of Wheeze-Relief Formula on the Degranulation of Eosinophils --- p.180 / Chapter 10.2.3. --- In vitro Effects of Wheeze-Relief Formula on the Surface Expression of Adhesion Molecules and Chemokine Receptors on Eosinophils --- p.183 / Chapter 10.2.4. --- In vitro Effects of Wheeze-Relief Formula on the Surface Expression of Adhesion Molecules on Eosinophils --- p.183 / Chapter 10.2.5. --- In vitro Effects of Wheeze-Relief Formula on the Cytokine Release from Peripheral Blood Mononuclear Cells --- p.187 / Chapter 10.2.6. --- In vitro Effects of Wheeze-Relief Formula on the Gene Expression Profile of Cytokines and Cytokine Receptors of Peripheral Blood Mononuclear Cells --- p.187 / Chapter 10.3. --- Discussion --- p.196 / Chapter Chapter 11: --- The Clinical Trial: Analysis of Serological Markers --- p.200 / Chapter 11.1. --- Introduction --- p.200 / Chapter 11.2. --- Results --- p.202 / Chapter 11.2.1. --- Demographic Data and Drop-out Cases --- p.202 / Chapter 11.2.2. --- Lung Function Test --- p.202 / Chapter 11.2.3. --- Steroid Dosage --- p.202 / Chapter 11.2.4. --- Serological Markers --- p.205 / Chapter 11.3. --- Discussion --- p.215 / Chapter Part VI - --- Conclusion / Chapter Chapter 12: --- Concluding Remarks and Future Perspectives --- p.217 / Chapter Part VII- --- Appendix / Parent Information Sheet --- p.222 / 家長資訊 --- p.223 / Consent Form --- p.224 / Licence to Conduct Animal Experiments --- p.225 / Bibliography --- p.227

Ventilatory drive is mediated by respiratory central pattern generators that are located in the brainstem, which are continuously modulated by specialised peripheral and central chemoreceptors to adjust ventilatory patterns according to changes in arterial PO2. These specialised oxygen-sensing chemoreceptors are activated in response to acute reductions in arterial PO2 and ultimately trigger a respiratory response that acts to restore oxygen-levels. However, the molecular mechanism by which mammals are able to regulate their breathing pattern in such a manner during hypoxia remains controversial. Therefore, the studies performed in this thesis aimed to investigate the possibility that this process may be mediated by the liver kinase B 1 (LKB1)/ AMP-activated protein kinase (AMPK) signalling pathway, which is central to cellular adaptations to metabolic stress. This first involved the development of transgenic mice in which Lkb1 or AMPK were deleted. Global knockout of Lkb1 (Sakamoto, 2006) or AMPK activity (Viollet et al., 2009) are embryonic lethal. Thus, the Cre/loxP system was used to develop transgenic mice that had either Lkb1 or both isoforms of the AMPK catalytic α- subunit (α1 and α2) conditionally knocked out in catecholaminergic cells (including therein hypoxia-activated cells of the brainstem and carotid body) by driving Cre expression through a tyrosine-hydroxylase-specific promoter region. The consequent effects on the ventilatory response to hypoxia were then examined using unrestrained whole-body plethysmography. This demonstrated that, in contrast to the hyperventilation evoked in controls, increased ventilation was virtually abolished in the Lkb1 and AMPK α1 and α2 double knockouts during hypoxia. Both knockout mice also exhibited periods of hypoventilation with frequent apnoeas during hypoxia. Additionally, studies on single AMPK α1 and AMPK α2 knockouts identified that the ventilatory dysfunction in AMPK α1 and α2 double knockouts was primarily caused by AMPK α1 deletion. In contrast, the severe ventilatory abnormalities exhibited during hypoxia following the deletion of Lkb1 and AMPK in catecholaminergic cells were mostly reversed upon exposure of mice to hypoxia with hypercapnia. Also, the ventilatory response to hypercapnia alone was without any major effect as a result of Lkb1 deletion or the dual-deletion of AMPK α1 and α2 catalytic subunits in catecholaminergic cells. This thesis therefore demonstrates, for the first time, that the LKB1-AMPK signalling pathway is key to respiratory adaptations during hypoxia, by regulating catecholaminergic oxygen-sensing cells, thus protecting against hypoventilation and apnoeas. The LKB1-AMPK signaling pathway can thereby determine oxygen and energy supply at both a cellular and whole-body level.

Meagre and the greater amberjack are promising emerging species in aquaculture. This is due to their fast growth rate, large size, high processing yield and high quality fillet. However, little is known about the immune system of these fish and several potential pathogenic threats to sustainable culture of these fish have been identified. Therefore, this thesis has begun to characterise the immune response of these species in order to better combat pathogenic threats. The focus of this thesis is the identification, modulation and monitoring of key innate and adaptive immune genes. This is achieved by identifying conserved regions of target gene sequence and designing consensus primers to these in order to generate a partial sequence, which could then be used as a base from which RACE PCR could be performed to obtain full sequence. The identified sequence was then used to produce qPCR primers, anti-meagre IgM and anti-meagre IgT monoclonal antibodies and a functional synthetic greater amberjack piscidin active peptide. Using the developed qPCR primers the constitutive expression of target genes was determined in the gills, gut, head kidney and spleen. Changes in transcript expression were then monitored in these tissues in response to PAMP stimulation in vivo and in vitro in order to better understand how these genes are modulated by viral, bacterial and fungal stimuli. Furthermore, the meagre development of the immune system was monitored from 1 day post hatch (dph) until 120 dph, providing insights into when immune maturity is achieved. Monoclonal antibodies were also produced and their viability tested in a variety of assays including Western blot, Immunohistochemistry and ELISA. Finally, A synthetic greater amberjack piscidin was produced and the bacteriostatic capabilities of this peptide were tested and showed promising results against known bacterial fish pathogens, indicating a potential for future uses within and outwith aquaculture.

The thesis deals with the Efficient Consumer Response system, which is based on partnership between retailer and his supplier to prepare value added for customer. In the teoretical part of the thesis are mentioned 2 main strategies: Efficient Replenishment and Category management. Practical part of the thesis dedicates usage of the system in practice.

Plants have evolved a plethora of effective mechanisms to protect themselves from biotic stresses. Jasmonates (JAs) are employed as vital defence signals against both insect and pathogen attack. Jasmonic acid (JA) signalling plays a central role in plant defence and development. S-nitrosylation, a redox-based post-translational modification plays an important role in plant disease resistance. S-nitrosoglutathione (GSNO) is formed by the reaction of antioxidant glutathione (GSH) and nitric oxide (NO) and acts as a mobile reservoir of NO bioactivity. The Arabidopsis thaliana S-NITROSOGLUTATHIONE REDUCTASE (AtGSNOR1) controls multiple modes of disease resistance via S-nitrosylation. In this context, the Arabidopsis lossof- function mutant atgsnor1-3 exhibits higher susceptibility to Botrytis cinerea a necrotrophic pathogens and Pieris rapae insect attack. Accumulation of JA was reduced in atgsnor1-3 after mechanical wounding. JA marker genes were also downregulated in atgsnor1-3 compared to Col-0 after Methyl Jasmonate (Me-JA) treatment. The relative gene expression of Vegetative Storage Protein (VSP) was reduced in atgsnor1-3 compared to wild type. Further, protein-protein interaction experiments in yeast two hybrid assays revealed an inhibition of Coronatine-insensitive 1 (COI1) and Jasmonate ZIM domain (JAZ1) interactions upon NO donor application. Interestingly it was also shown that Nitric oxide donor may inhibited the degradation of JAZ1-β-glucoronidase (GUS) fusion protein driven by a CaMV35s:: JAZ1-GUS transgene in GUS histochemical analysis but not in flurometric assay. A biotin switch assay of recombinant JAZ1-Maltose-binding protein (MBP) has shown that JAZ1-MBP was S-nitrosylated and mass spectrometry suggested Cysteine229 (Cys229) was the site of this modification. Further, CaMV35S::JAZ1-Flag transgene expressed in either a wild-type or atgsnor1-3 genetic background, suggested that JAZ1 was S-nitrosylated in vivo. Collectively, our data imply that JA-signalling engaged in response to either insect predation or attempted B. cinerea infection is under redox control as high SNO in atgsnor1-3 has disrupted the JA signalling pathway. Furthermore, our data suggest that S-nitrosylation of Cys-229 of JAZ1 may control JA-mediated signalling by blocking the interaction of this protein with COI1, thus reducing the turnover of JAZ1 by the 26S proteasome and consequently enabling continued JAZ1-mediated repression of JA-dependent gene expression in the presence of Me-JA. Thus our findings highlight the importance of NO and associated S-nitrosylation in JA signalling during plant immune response.

Constitutive activation of the ATM dependent DNA damage response and repair pathways have been reported in pre-malignant and malignant human tissues and may undermine the efficacy of genotoxic cancer therapies. Therefore, ATM inhibitors may overcome resistance to current cytotoxics and potentiate the effects of radiotherapy. A colorectal cancer model was investigated to develop a framework for the rational use of ATM inhibitors. HCT116 p21-/- cells display constitutive activation of the ATM DNA damage response but display a defect in the ionising radiation induced S-phase checkpoint, termed radioresistant DNA synthesis. This radioresistant phenotype is associated with increased basal levels of Cdc25A protein, deficient DNA damage-induced degradation of Cdc25A and Chk2 mis-localisation. HCT116 p21-/- and SW620 cells, which exhibit basal Chk2 threonine-68 phosphorylation, were unable to abrogate the S-phase checkpoint when treated with an ATM inhibitor, suggesting that the ATM– Chk2 arm is non-functional in these cells: inhibition of ATM did not potentiate the efficacy of ionising irradiation. To assess activation of the pathway a tumour microarray was created using 179 treatment naïve sporadic colorectal cancers; 152 were of the microsatellite stable phenotype. Phosphorylated Chk2 threonine-68 was present in 22 % of microsatellite-stable colorectal tumours and 33 % of tumours with the microsatellite instability phenotype. In a colorectal cancer cell line model constitutive activation of the ATM DDR pathway reflected an attenuated ATM-Chk2 axis and inhibition of ATM in these circumstances was unable to potentiate the efficacy of ionising irradiation. Basal Chk2 threonine-68 phosphorylation may reflect a deregulated ATM DNA damage response pathway and/or checkpoint adaption and therefore use of an ATM inhibitor in this background may have limited efficacy. A predictive model is proposed that integrates functionality of the ATM-Chk2 axis, p53 mutation status and defects in DNA repair pathways when considering ATM inhibitor therapy. Ultimately, molecular phenotyping and functional analysis of processes deregulated in cancer will permit individualisation of current treatment modalities, improving their efficacy and limiting patient toxicity.