Interleukin-17A (IL-17A)is an important pro-inflammatory cytokine that plays a critical role in host defenses against diverse pathogens. Studies have shown that IL-17Aplays protective role against sub-lethal H1 and H3 subtypes influenza infections, but it is unclear about the role of IL-17A in the highly pathogenic H5N1 and lethal H1N1 influenza virus infection. B cell is an important effector cell type in anti-influenza immunity. Although roles of B cell in influenza infection have been extensively investigated, it is unclear whether and how IL-17AregulatesB cell response during influenza infection. I examined the role of IL-17A against influenza infection by challengingIL-17A knockout (KO) and wild-type (WT) mice with highly pathogenic H5N1 and lethal H1N1 influenza viruses. Following challenge, IL-17AKO mice exhibited significantly lower survival rate, profoundly reduced body weight, more severe tissue damage and higher viral burden in the lung tissues. These evidences suggest that IL-17Aplays a protective role in lethal influenza infection. To study whether IL-17Amodulates B cell response against influenza, I found that both B-1a and B-2 cells were detected in the lung tissue and pulmonary draining lymph node, Mediastinal lymph node (MedLN),as early as 2days post-infection. Meanwhile, B-1a cells predominantly contributed to the early virus-specific IgM in the respiratory tract. However, virus-specific IgM markedly reduced in IL-17A KO mice when compared with WT controls. Adoptive transfer of B-1a cells or B-1a cell-derived antibodies conferred protection in IL-17A KO mice. These results demonstrate that IL-17A plays a critical role in modulating early antibody production of B-1a cells against lethal influenza infection. To further elucidate how IL-17A regulates B-1a cell response, I observed that B-1a cells migrated into MedLN and lung tissues during infection and underwent plasmacytic differentiation with increased antibody production in airways. IL-17A deficiency impaired these processes of B-1a cells, while intra-nasally instillation of IL-17A restored B-1a cell response by promoting both B-1a cell migration and plasmacytic differentiation. By inducing blimp-1 expression in B-1a cells in an NF-κB dependent pathway, IL-17A directly promoted plasmacytic differentiation of B-1a cells both in vivo and in vitro. Furthermore, chromatin immuno-precipitation analysis confirmed that NF-κB directly bound to the promoter of blimp-1 gene and promoted blimp-1 expression in B-1a cells following IL-17A stimulation. To determine the functional significance of IL-17A signaling in modulating B cell response against influenza infection, I first uncovered markedly reduced B cell response, predominantly B-1a cell response in IL-17A KO mice, showing reduced local migration and impaired plasmacytic differentiation in the early stage of infection. Next, intra-nasal administration of IL-17A into IL-17A KO mice significantly restored this B-1a cell response. Moreover, I detected expression of IL-17A receptor in B-1a cells. IL-17A treatment could promote antibody production from B-1a cells by inducing blimp-1 expression in an NF-κB dependent pathway. Taken together, these findings identify a novel role of IL-17A in actingas an immune modulator of B cell response against influenza infection, which will contribute to a fuller understanding of B cell biology and anti-viral response in host defense. / published_or_final_version / Pathology / Doctoral / Doctor of Philosophy

Probiotics are living microorganisms, which when administered in adequate amounts confer a health benefit on the host. They have been reported to relieve acute diarrhoea, atopic dermatitis and irritable bowel syndrome in disease-specific animal studies and in human intervention trials. However, probiotics are regularly consumed by general healthy population with limited knowledge in the immunomodulation of probiotics of local and systemic immune responses in healthy experimental models. Serving as the first line of defense against microbial infections and the largest immunological organ in animal host, the epithelium lining the small and large intestine is supposed to be the first organ to encounter probiotics as probiotics are always orally taken. It is believed that probiotics regulate the local immunities in the gut, which acts as the pivot in modulating the systemic immune responses. Accordingly, it was hypothesized that probiotic bacteria can modulate both local and systemic immune responses in healthy population; and the immunomodulation of combination of probiotics is different from that of individual strains. Wildtype healthy C57BL/6 mice were fed with different probiotic strains − Lactobacillus rhamnosus GG (LGG), Lactobacillus rhamnosus LC705 (LC705), Bifidobacterium breve Bb99 (Bb99), Propionibacterium freudenreichii ssp. shermanii JS (PJS) or Escherichia coli Nissle 1917 (EcN), or mixture of probiotics − GGmix (LGG, LC705, Bb99 and PJS) and ECPJSmix (PJS and EcN), for three weeks. After that, intestine, liver, spleen and blood were investigated. Probiotics suppressed intestinal T helper (Th)17 immune response but enhanced systemic (hepatic and splenic) Th17 immune response, suggesting that immune homeostasis was maintained in healthy individuals. Mechanism of action of LGG was further studied in this project as LGG is the widely studied probiotics. It was hypothesized that LGG exerts immunomodulatory effects by bacteria cells and/or its derived soluble factors such as lactic acid. Immunomodulatory effects of LGG cells and their soluble factors on dendritic cells (DCs), macrophages and monocytes from healthy blood donors were investigated as antigen-presenting cells (APCs) are pivots of bridging innate and adaptive immunities. Cytokine secretion profile, expressions of toll-like receptors (TLRs) and activation-related receptors of the APCs were examined. Both LGG cells and their soluble factors promoted type 1-responsiveness while soluble factors promoted type 17-responsiveness as well. Yet, lactic acid seemed not to be the one which enhanced type 1 and type 17 immune responses in soluble factors. With better understanding on the immunomodulation of probiotics in healthy models, prophylactic efficacy of probiotics in preventing infections and diseases can be availed. / published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

Graduate

published_or_final_version / Medicine / Master / Doctor of Medicine

No description available.

No description available.

In this thesis, mechanisms of immunoregulation by CD4$ sp+$ T cells during blood-stage P. chabaudi AS infection in C57BL/6 mice were studied. The kinetics of in vitro production of the Th1-derived cytokine, IFN-$ gamma$, versus the Th2-derived cyokines, IL-4, IL-5 and IL-10, by spleen cells as well as of polyclonal and malaria-specific antibodies in the sera were examined during infection using enzyme-linked immunosorbent assays. Upon antigenic stimulation, spleen cells were found to produce high levels of IFN-$ gamma$ several days prior to peak parasitemia, while high levels of IL-10 production occurred at the time of peak parasitemia followed by IL-4 and IL-5 later in infection. The levels of polyclonal IgG2a isotype were found to be increased during both the acute and chronic phases of infection, whereas the levels of polyclonal IgM, IgG1 and IgG2b isotypes were found to be increased only during the chronic phase of infection. High titers of malaria-specific IgG2a and IgG1 were detected during the primary as well as secondary infections. Investigation of in vitro proliferation of spleen cells to mitogens and malaria specific antigen revealed that the responses of splenic lymphocytes from infected mice to Con A, PHA and LPS were suppressed, with the most severe suppression occurring during the first 14 days post infection. Evidence is provided demonstrating that nitric oxide (NO) and prostaglandins (PG), products of activated macrophages, mediated suppression of lymphocyte proliferation in response to Con A and PHA, whereas only PG were found to suppress LPS-stimulated proliferation. In addition, NO was found to mediate suppression of proliferation of spleen cells from infected mice in response to parasite antigen. Taken together, results from these studies suggest that immune activation and immunosuppression occur simultaneously during blood-stage malaria with P. chabaudi AS infection in C57BL/6 mice. (Abstract shortened by UMI.)

Enhanced enteric delivery systems were examined in the cichlid Oreochromis mossambicus to evaluate the requirements for potential oral vaccination strategies in teleost fish. Human gamma globulin (HGG) was delivered enterically (orally or anally) to the intestine of O.mossambicus as a standard test antigen. The co-administration of the saponin Quillaja saponaria (Quil-A) was evaluated as a novel oral adjuvant, delivered with the antigen in a soluble form and also as a delivery vehicle in the form of micelles and immunestimulatory complexes (ISCOMS). HGG absorption across the intestine was monitored utilising sensitive immunocytochemical techniques which showed that the enterically delivered antigen was transcytosed in a temporally complex manner, underwent extensive interaction with the enterocytes and gut-associated lymphoid tissue (GALT) cells and was eventually transported to the vicinity of the intestinal circulatory system Co-administration of Quil-A resulted in an increase in the HGG absorbed, increased antigen localisation in the lamina propria and substantial interaction of the adjuvant with the enterocyte lumenal membranes. The levels of HGG absorbed into the plasma were directly quantified by enzyme-linked immunosorbent assay (ELISA) and showed that administration of Quil-A concurrently with HGG greatly increased absorption above levels observed without adjuvant in the delivery system. Western blotting and laser densitometry demonstrated that HGG was absorbed as both an intact macromolecular antigen and as fragmented epitopes of distinct molecular weights. The fragmentation of the enterically delivered HGG was modified by the delivery of Quil-A suggesting that manipulation of conformational aspects of the delivered antigen may be possible. The systemic and mucosal immune responses to HGG administration were monitored and enteric immunisation of antigen with Quil-A was found to be effective in increasing specific antibody levels in the plasma, bile and cutaneous mucus of immunised fish. Preliminary studies on the use of cholera toxin β-subunit, aluminium hydroxide and ammonium chloride showed that cholera toxin β-subunit acted to increase both level of absorbed antigen, after enteric delivery and the subsequent immune response to HGG whereas the other two adjuvants were unable to mediate any such responses.

Protective immunity of carp to ichthyophthiriasis has been confirmed, and demonstrated for the first time in juvenile carp, 10-12 weeks old. Standard immunisation procedures were developed here using the theront in preference to cysts. Immunisation included exposure of fish on 3 separate occasions of 14 day intervals to doses of approximately 2,000 theronts per fish, 80/ cm³. Procedures were controlled so that infections were not allowed to continue beyond the primary stage pH (7.0-7.2 ) and temperature (20±2° C) were maintained throughout experimental periods. Four weeks after third immunising dose, fish were exposed to a potentially lethal challenge, approximately 8,000 theronts per fish, 320/cm³. Following immunisation , fish showed total protection up to 1 month and decreasing protection up to 3 months during which period, mortalities were recorded on challenge. Humoral antibody was monitored at specific stages of experimental infections, peak response 6-8 weeks following exposure with detectable levels of antibody persisting for at least 12 weeks. Immunosuppression was demonstrated following intraperitoneal administration of synthetic corticosteroid Triamcinolone acetonide, doses of 200 µg, 100 µg and 10 µg gˉ¹ body weight, and corticosteroid Hydrocortisone 21-hemisuccinate, doses of 100 µg and 10 µg gˉ¹ body weight, given 14 days after challenge. Immunosuppression was not associated with any significant fall in antibody titre. Studies in cross immunity between Tetrahymena pyriformis (CCAP 1630/W Claff, 1939 (w)) and Ichthyophthirius multifiliis showed no evidence of the former conferring protection to ichthyophthiriasis. Methods of administration of T. pyriformis to juvenile carp included intraperitoneal injection of freeze dried cilia and whole, live T. pyriformis. The kinetics of the humoral response were measured over 12 weeks, peak antibody titres occurring 6-8 weeks following antigen administration. Proliferative responses measured by autoradiography were recorded prior to peak antibody production. Overall results are discussed in relation to immunosuppression, mechanisms of immunity and control and treatment of the disease.

Bibliography: leaves 170-206. / vi, 206 leaves : ill. (some col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / In T-dependent antibody responses, costimulator molecules provide contact-medited signals during interactions between T cells and B cells that regulate lymphocyte activation. This study investigated the hypothesis that costimulator molecules are differentially expressed on lymphocytes from neonates and young children compared with adults, contributing to limitations of T-dependent antibody responses in early life. Results suggest that lymphocytes from young children should be able to deliver and respond to costimulatory signals. The differences in lymphocyte expression of these costimulator molecules in young children are unlikely to fully account for limitations in humoral immunity in early life, and may even represent a specialised adaptation for this stage of immune development. / Thesis (Ph.D.)--University of Adelaide, Dept. of Paediatrics, 2000