Thesis (M.D.)--University of Hong Kong, 2001. / Includes bibliographical references (leaves 130-155).

Thesis (Ph. D.)--University of Queensland, 2002. / Includes bibliographical references.

Enterovirus 71 (EV71) belongs to the Enterovirus genus of the family Picornaviridae and is the major causative agent of hand, foot and mouth disease (HFMD). Although clinical manifestations of HFMD are usually mild and self-limiting, severe HFMD patients suffer from a diverse array of neurological diseases and sometimes these diseases are fatal. HFMD usually occurs in young children and gradually becomes a new threaten in Asia. Unfortunately, effective EV71 vaccine is not available to date and alternative treatments are still in debate. This is partially due to the lack of understanding of EV71 pathogenesis and host immune responses against EV71. Natural killer (NK) cells are key effector cells in host antiviral activities by directly killing viral-infected cells and producing cytokines and chemokines, especially in early phase of viral infection. After enteroviruses infection, NK cells were one of the most abundant cell types in the inflammatory infiltrate, and appeared to limit both enteroviruses replication and virus-induced disease in experimental mice model. However, role of human NK cells during EV71 infection, especially the direct interaction between EV71 and human NK cells, was not studied extensively. Clinical observation manifested that patients with severe EV71 infection have marked diminished NK cells in peripheral blood. Therefore we hypothesized that EV71 might directly target human NK cells as one of its immunoevasion strategies. Here, we demonstrated for the first time that fresh primary human NK cells were susceptible to EV71 infection. By flow cytometry and florescence microscope, EV71 capsid protein VP1 was able to be detected in viral-infected NK cells as soon as 6 hours after infection and peaked at 24 hour after infection. In the same time, EV71 viral RNA was detected by quantitative RT-PCR and the viral copies increased from 6 hour onwards to peak at 12 hours after infection. We further demonstrated the infectious entry of EV71 in human NK cells was depended on clathrin-mediated endocytosis. Next, we illustrated that EV71 infection could trigger NK cells apoptosis as evidenced by increased Annexin V+, PI+, and activated caspase 3+ cells in EV71-treated NK cells. We further proved that the cytotoxicity of NK cells was inhibited by EV71 infection and this inhibition might not be related with down-regulation of NKp46, but may be related to the increased apoptosis. In conclusion, our data suggested that EV71 might directly target and kill NK cells as a strategy to evade human innate immunity, which might facilitate virus replication, transmission and then contribute to viral-related pathogenesis. / published_or_final_version / Paediatrics and Adolescent Medicine / Master / Master of Philosophy

published_or_final_version / Medicine / Master / Doctor of Medicine

No description available.

Uterine Natural Killer cells are the major lymphocyte population in the pregnant uterus in early gestation; outnumbering both T and B cells. The numerical expansion of uterine Natural Killer cells is thought to result from the expansion of preexisting progenitor cells resident to the uterus, recruitment of Natural Killer cells from the circulation, or a combination of both pathways. Uterine Natural Killer cells are capable of cytotoxic killing and express receptors that can recognize foreign paternal antigens. Therefore, it has been argued that uterine Natural Killer cell activation can lead to killing of fetal cells and abortion. However, fetal rejection by uterine Natural Killer cells does not occur in normal pregnancies and other functions for uterine Natural Killer cells have been proposed. These include: the regulation of maternal blood supply responsible for providing oxygen to the fetus, regulation of maternal blood pressure and, in species with invasive placentation, regulation of decidualization, the process of endometrial cell expansion and transformation during the menstrual cycle and during pregnancy. These cell functions juxtapose the concept that uterine Natural Killer cell activation is harmful to the fetus and offer a new perspective that uterine Natural Killer cells regulate functions unrelated to traditional transplantation immunology. In this dissertation, work is presented showing that uterine Natural Killer cells express molecules which regulate blood pressure and decidualization. Also presented are data supporting the hypothesis that the numerical increase of uterine Natural Killer cells is due to the recruitment of Natural Killer cells from the blood. These results support roles for uterine Natural Killer cells other than cytotoxic killing and advance the understanding of uterine Natural Killer cells as dynamic players that support pregnancy-associated biological processes unrelated to traditional understandings of immune surveillance. / Thesis (Master, Microbiology & Immunology) -- Queen's University, 2009-12-15 11:33:37.11

Computational analysis of total amino acid sequences indicate that select combinations that occur less frequently are correlated to increased immunogenicity in humans. Much evidence has been gathered in silico, but little is known about in vivo experimental validation. This concept can be applied to adjuvant research where increased immunogenicity is desirable and can aid in the potency and efficacy of vaccines. A rare peptide called 5mer4 was found to adjuvant influenza vaccines by increasing survival, humoral and cellular immune responses with a speculated NK cell mediated mechanism. Therefore we hypothesize that rare peptides are able to stimulate an increased immune response in comparison to common peptides through a NK-mediated fashion. The first aim of this study is to determine whether rare sequences are able to stimulate an increased immune response collectively in comparison to commonly occurring peptides. Mice vaccinated with rare, semi-common and common peptides indicate a trend of heightened cellular immune response from rare peptides. However, select rare peptide sequences based on high IFNγ responses do not always correlate directly to increased vaccine efficacy against H5N1-H05 influenza virus, indicating that additional immune parameters need to be taken into consideration. When compared against other adjuvants, 5mer4 performed better in both humoral and survival studies. Previous findings suggest NK cell involvement warranted the second aim of this thesis which is to further delineate the role of NK cells as rare peptide immune modulators. Macrophages were evaluated to determine the effect of peptide, but no increase in stimulation could be observed. NK cells incubated with rare peptides show increased levels of early activation marker CD69 in comparison to common peptides. Microscopy data indicates that rare, but not common peptides are able to bind to NK cells. Depletion of NK abrogated adjuvant activity of 5mer4 peptide, suggesting the necessary role of NK cells for adjuvant effect. Taken together, rare peptides have shown the ability to modulate the immune response through NK cell activation verifying our hypothesis. These findings can be extrapolated towards multiple fields such as anti-tumor therapies and can lead to the development of immunomodulators with high efficacy at a lower cost.

In this thesis I take a multi-disciplinary approach to identify and characterise ‘Evolutionary Significant Units’ (ESUs) and ‘Management Units’ (MUs) of killer whale in the eastern North Atlantic. Several markers and traits including mitochondrial DNA control region, 15N stable isotope values, tooth wear, tooth count, total body length and pigmentation pattern indicated sympatric lineages could be categorized in to two ecologically & morphologically disparate types, which should be considered as distinct ESUs. One type (type 2) appears to be a specialist and may predate cetaceans. The other type (type 1) appears to be a generalist, although the total niche width of the population appears to be more dependent upon between-individual variation than within-individual variation in dietary composition. However, the other indicator of long-term diet, apical tooth wear, was found in all type 1 individuals, suggesting some overlap in the components of the diet. There are shallow genetic differences between the two types based on mtDNA control region, however analysis of bi-parentally inherited nuclear DNA markers are needed to see if the two types are reproductively isolated. The data above on diet is further supported by observational data of individuals moving between the Icelandic herring grounds to the seal pupping haul-outs around the Northern Isles, Scotland. However, large scale movement of individuals is mostly correlated with the movement of large predictable prey stocks such as the Icelandic and Norwegian stocks of Atlantic herring. Therefore there is the potential for prey choice to cause intrinsic isolation through temporal or spatial isolation even when the prey type is similar, e.g. mackerel, Icelandic herring or Norwegian herring. The microsatellite data are consistent with this pattern and taken together the genetic and mark-recapture data identify four demographically independent MUs of type 1 killer whales. We only identify one community or MU of type 2 killer whales, which is small (10 individuals) and appears to be suffering from demographic stochasticity. Our results suggest intrinsic isolation through temporal and spatial isolation as the most parsimonious mechanism for reducing gene flow between populations of the same type. Further work is needed to determine if there is gene flow between types or if adaptive divergence feeds back to reduce gene flow through mate choice.

Computational analysis of total amino acid sequences indicate that select combinations that occur less frequently are correlated to increased immunogenicity in humans. Much evidence has been gathered in silico, but little is known about in vivo experimental validation. This concept can be applied to adjuvant research where increased immunogenicity is desirable and can aid in the potency and efficacy of vaccines. A rare peptide called 5mer4 was found to adjuvant influenza vaccines by increasing survival, humoral and cellular immune responses with a speculated NK cell mediated mechanism. Therefore we hypothesize that rare peptides are able to stimulate an increased immune response in comparison to common peptides through a NK-mediated fashion. The first aim of this study is to determine whether rare sequences are able to stimulate an increased immune response collectively in comparison to commonly occurring peptides. Mice vaccinated with rare, semi-common and common peptides indicate a trend of heightened cellular immune response from rare peptides. However, select rare peptide sequences based on high IFNγ responses do not always correlate directly to increased vaccine efficacy against H5N1-H05 influenza virus, indicating that additional immune parameters need to be taken into consideration. When compared against other adjuvants, 5mer4 performed better in both humoral and survival studies. Previous findings suggest NK cell involvement warranted the second aim of this thesis which is to further delineate the role of NK cells as rare peptide immune modulators. Macrophages were evaluated to determine the effect of peptide, but no increase in stimulation could be observed. NK cells incubated with rare peptides show increased levels of early activation marker CD69 in comparison to common peptides. Microscopy data indicates that rare, but not common peptides are able to bind to NK cells. Depletion of NK abrogated adjuvant activity of 5mer4 peptide, suggesting the necessary role of NK cells for adjuvant effect. Taken together, rare peptides have shown the ability to modulate the immune response through NK cell activation verifying our hypothesis. These findings can be extrapolated towards multiple fields such as anti-tumor therapies and can lead to the development of immunomodulators with high efficacy at a lower cost.

1 v. (various paging) : photos, graphs, tables ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (Ph.D.)--University of Adelaide, Dept. of Oral Biology, 1980