No description available.

No description available.

No description available.

by Zhi-Hui Xie. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1996. / Includes bibliographical references (leaves 214-244). / Abstract --- p.i / Acknowledgments --- p.iv / Publications --- p.v / Abbreviations --- p.vi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Neutrophils --- p.1 / Chapter 1.1.1 --- General --- p.1 / Chapter 1.1.2 --- HL-60 cells as model of neutrophils --- p.3 / Chapter 1.1.3 --- Physiological role --- p.6 / Chapter 1.1.4 --- Intracellular signalling --- p.12 / Chapter 1.1.4.1 --- General --- p.12 / Chapter 1.1.4.2 --- Intracellular signalling linked by FMLP receptors --- p.13 / Chapter 1.1.4.3 --- Intracellular signalling of neutrophil functions --- p.17 / Chapter 1.2 --- Neutrophil aggregation --- p.21 / Chapter 1.2.1 --- General --- p.21 / Chapter 1.2.2 --- Measurement of aggregation --- p.24 / Chapter 1.3 --- Prostaglandins/Prostanoids --- p.25 / Chapter 1.3.1 --- General --- p.25 / Chapter 1.3.2 --- Prostanoid receptors --- p.27 / Chapter 1.3.2.1 --- Agonists and antagonists --- p.28 / Chapter 1.3.2.2 --- EP1 receptors --- p.32 / Chapter 1.3.2.3 --- EP2 receptors --- p.33 / Chapter 1.3.2.4 --- EP3 receptors --- p.34 / Chapter 1.3.2.5 --- EP4 receptors --- p.37 / Chapter 1.3.2.6 --- DP receptors --- p.38 / Chapter 1.3.2.7 --- IP receptors --- p.39 / Chapter 1.3.2.8 --- TP receptors --- p.40 / Chapter 1.3.2.9 --- FP receptors --- p.40 / Chapter 1.4 --- Regulation of neutrophil functions by prostanoids --- p.42 / Chapter 1.4.1 --- Inhibition --- p.42 / Chapter 1.4.2 --- Activation --- p.46 / Chapter 1.4.3 --- Role of cyclic AMP --- p.48 / Chapter 1.5 --- Aim of this study --- p.51 / Chapter Chapter 2 --- Materials and methods --- p.53 / Chapter 2.1 --- Materials and solutions --- p.53 / Chapter 2.1.1 --- Materials --- p.53 / Chapter 2.1.2 --- "Buffers, solutions and cell culture medium" --- p.62 / Chapter 2.2 --- Methods --- p.64 / Chapter 2.2.1 --- Culture of HL-60 cells --- p.64 / Chapter 2.2.2 --- Differentiation of HL-60 cells --- p.65 / Chapter 2.2.3 --- Preparation of neutrophils --- p.65 / Chapter 2.2.4 --- Measurement of neutrophil aggregation --- p.67 / Chapter 2.2.5 --- Measurement of [Ca2+]i --- p.69 / Chapter 2.2.6 --- Microscopic observation --- p.70 / Chapter 2.2.7 --- Measurement of [3H]-cyclic AMP accumulation --- p.72 / Chapter 2.2.8 --- Measurement of [3H]-PGE2 receptor binding --- p.73 / Chapter 2.3 --- Data analysis --- p.76 / Chapter Chapter 3 --- Differentiation of HL-60 cells --- p.77 / Chapter 3.1 --- Introduction --- p.77 / Chapter 3.2 --- Results and discussion --- p.78 / Chapter 3.2.1 --- The observation of cell proliferation and morphology --- p.78 / Chapter 3.2.2 --- The response of HL-60 cells to FMLP --- p.80 / Chapter 3.2.3 --- The response of HL-60 cells to ATP --- p.83 / Chapter 3.3 --- Conclusion --- p.84 / Chapter Chapter 4 --- Activation of dHL-60 cells by PGE2 --- p.90 / Chapter 4.1 --- Effect of PGE2 on cell aggregation and [Ca2+]i --- p.90 / Chapter 4.1.1 --- Introduction --- p.90 / Chapter 4.1.2 --- Results --- p.90 / Chapter 4.1.2.1 --- Effect of PGE2 on cell aggregation --- p.90 / Chapter 4.1.2.2 --- Effect of PGE2 on [Ca2+]i --- p.91 / Chapter 4.1.2.3 --- Effect of PGE2 on human neutrophils --- p.93 / Chapter 4.1.2.4 --- Do PGE2 and PGE1 act at the same receptor? --- p.94 / Chapter 4.1.3 --- Discussion --- p.95 / Chapter 4.1.3.1 --- Activation of dHL-60 cells by PGE2 --- p.95 / Chapter 4.1.3.2 --- Effect of PGE2 on human neutrophils --- p.99 / Chapter 4.2 --- What does the aggregation response mean? --- p.101 / Chapter 4.2.1 --- Introduction --- p.101 / Chapter 4.2.2 --- Results --- p.101 / Chapter 4.2.2.1 --- Observation by light microscopy --- p.101 / Chapter 4.2.2.2 --- Observation by Scanning Electron Microscope --- p.102 / Chapter 4.2.3 --- Discussion --- p.103 / Chapter 4.3 --- Conclusion --- p.106 / Chapter Chapter 5 --- Characterisation of prostanoid receptors on dHL-60 cells --- p.121 / Chapter 5.1 --- Introduction --- p.121 / Chapter 5.2 --- Results --- p.121 / Chapter 5.2.1 --- Effect of prostanoid receptor agonists and antagonists on cell aggregation --- p.121 / Chapter 5.2.2 --- Effect of prostanoid receptor agonists on [Ca2+]i --- p.122 / Chapter 5.3 --- Discussion --- p.123 / Chapter 5.3.1 --- Involvement of prostanoid receptors --- p.123 / Chapter 5.3.2 --- Involvement of EP receptors --- p.126 / Chapter 5.4 --- Conclusion --- p.130 / Chapter Chapter 6 --- Binding studies of [3H]-PGE2 on rat neutrophils and dHL-60 cells --- p.137 / Chapter 6.1 --- Introduction --- p.137 / Chapter 6.2 --- Results --- p.138 / Chapter 6.3 --- Discussion --- p.140 / Chapter Chapter 7 --- The mechanism of action of PGE2 in activating dHL-60 cells --- p.146 / Chapter 7.1 --- Introduction --- p.146 / Chapter 7.2 --- Methods --- p.147 / Chapter 7.2.1 --- Pretreatment with enzyme inhibitors --- p.147 / Chapter 7.2.2 --- Pretreatment with toxins --- p.148 / Chapter 7.2.3 --- Pretreatment with PMA --- p.148 / Chapter 7.3 --- Results and discussion --- p.148 / Chapter 7.3.1 --- The role of Ca2+ --- p.149 / Chapter 7.3.2 --- The role of cyclic AMP --- p.152 / Chapter 7.3.3 --- The role of PI3-kinase --- p.159 / Chapter 7.3.4 --- The role of PLD --- p.165 / Chapter 7.3.5 --- The role of PLA2 --- p.169 / Chapter 7.3.6 --- The role of tyrosine kinase --- p.171 / Chapter 7.3.7 --- The role of PKC --- p.173 / Chapter 7.3.8 --- The role of G protein --- p.180 / Chapter 7.3.9 --- The role of cyclic GMP --- p.185 / Chapter 7.3.10 --- The role of KATP channels --- p.186 / Chapter 7.4 --- Conclusion --- p.187 / Chapter Chapter 8 --- General discussion and conclusion --- p.209 / References --- p.214

Cellular immune responses are orchestrated by an intricate balance of activating and inhibitory signals transmitted by cell surface receptors. Perturbations in this balance by overamplified or dysregulated signalling underlie many severe immunopathologies such as sepsis and cancer. In this work I describe the identification and characterisation of a novel, evolutionarily conserved immunoreceptor encoded within the human leukocyte receptor complex and syntenic region of mouse chromosome 7, named T cell-interacting, activating receptor on myeloid cells-1 (TARM1). The transmembrane region of TARM1 contained a conserved arginine residue, consistent with association with a signalling adaptor. Co-immunoprecipitation experiments confirmed that TARM1 associated with the ITAM adaptor FcR-gamma but not with DAP10 or DAP12. Flow cytometric screening of cells and tissues from pathogen-free mice showed that the TARM1 protein was constitutively expressed on the cell surface of mature and immature CD11b+Gr+ neutrophils isolated from bone marrow but not at peripheral sites. Following ip LPS treatment or systemic bacterial challenge, TARM1 protein expression was upregulated by myelocytes, mature neutrophils and inflammatory monocytes and TARM1+ cells were rapidly recruited to sites of inflammation. TARM1 expression was also upregulated by bone marrow-derived macrophages and dendritic cells following stimulation with TLR agonists in vitro. Ligation of the TARM1 receptor with specific antibody in the presence of TLR ligands, such as LPS, enhanced the secretion of proinflammatory cytokines by bone marrow-derived macrophages and primary mouse neutrophils, whereas TARM1 stimulation alone had no effect. Finally, an immobilised TARM1 Fc fusion protein suppressed CD4+ T cell activation and proliferation in vitro. These results suggest that a putative T cell ligand can interact with TARM1 receptor, resulting in bidirectional signalling and raising the T cell activation threshold while costimulating the release of proinflammatory cytokines by macrophages and neutrophils.

Some mounted ill. Bibliography: leaves 197-226.

Thesis (Ph. D.)--University of Rhode Island, 2006. / Typescript. Includes bibliographical references (leaves 162-177).

No description available.

No description available.

Neutrophils, key cells of the innate immune system, are responsible for preventing bacterial infections. They are rapidly recruited to sites of infection where they eliminate bacteria through killing methods that require reactive oxygen dependent processes. It has recently been established that neutrophils are capable of rapid and complex changes in gene expression during inflammatory responses. The concept that neutrophils only directly kill bacteria has been replaced by the concept that activated neutrophils can influence the immune response through the secretion of a variety of cytokines and by acting as antigen-presenting cell (APC) expressing MHC Class II, allowing for activation of T cells. Recent advances in neutrophil biology demonstrated that neutrophils also have an active regulatory role in angiogenesis and tumoral fate. It has been noted that a number of diseases including arthritis, periodontitis and acute respiratory distress syndrome (ARDS) are associated with neutrophil hyperactivity that results in significant tissue damage. Our group has previously shown that for some periodontal diseases, neutrophil hyperactivity is a key determinant of disease progression and severity. However, it remains unclear what factors are responsible for a patient developing a hyperactive neutrophil mediated disease. I hypothesize that local gene expression changes in neutrophils are responsible for the hyperactive behaviour of these cells during an inflammatory response. In order to assess this, I characterized the neutrophil gene expression profile in various compartments (bone marrow, blood and peritoneum in mice and blood and oral cavity in humans) and then characterized this genetic and phenotypic profile during an inflammatory response. I hypothesize that the neutrophil has a characteristic set of genes that are normally activated when it enters a site of inflammation from the circulation and that neutrophils can be polarized into a different functional subset under certain conditions that result in inflammation mediated diseases. To identify changes in neutrophil gene expression in the circulation and inflamed tissue I used recent advances in neutrophil isolation, RNA amplification, and microarray technologies to characterize the specific transcriptome associated with neutrophil site-specific responses.