Dissertations / Theses on the topic 'Antigen-specific immune responses'

Stem cell transplantati.on (SCT) represents the only curative treatment option for leukaemia. The bone marrow or peripheral blood stem cells transferred in the transplant procedure restore immune functions, allowing the targeting of infected cells or cells expressing tumour antigens. Cytotoxic T lymphocytes (CTL), key mediators of antigen specific killing, were investigated in the context of cytomegalovirus (CMV) infection or chronic myeloid leukaemia (CML). CMV infection after SCT in the absence of effective immunological control or antiviral therapy is a significant cause of morbidity and mortality. HLA-A*0201 tetramer reagents were prepared with three candidate peptides and used to test healthy donor and SCT patient samples. Only T cells binding to the tetramer made with the predominant pp65 epitope (AE42: 495-503) were found for HLA-A*0201 healthy individuals and patients after SCT and correlated with the estimation of the responding T cell population to this peptide as measured by interferon-Tproduction. Parallel tetramer and viral load monitoring of patients at risk of CMV infection after SCT highlighted an inverse correlation between the state of replication of the virus and the number of CMV specific CTL. Presentation of a CML tumour specific peptide epitope in the context of HLA- A*0301 was demonstrated both at the surface of tumour positive cells lines and patient cells. HLA/peptide tetramer reagents were prepared with this peptide and used to screen CML patient samples. Low frequencies of peptide specific CTL were detected in some patients and could be stimulated in vitro. While donor lymphocytes infusions would improve CTL responses after SCT, they also carry the risk of graft versus host disease and may not comprise an effective CTL population targeted to the antigen of interest. A better outcome may be obtained by infusion of enriched and or expanded specific T cell populations. The enrichment, stimulation and expansion of CTL with HLA/peptide tetramer or modified tetramer complexes was examined, using the HLA-A*0201/CMV CTL response as a model. The use of HLA/peptide tetramers to monitor recipients of SCT has implications for the improvement of the treatment of CMV after SCT and the definition of targets and criteria for effective adoptive transfer. Furthermore, the use of HLA/peptide tetramers could be investigated in the assessment of anti-tumour CTL responses, in enhancing existing responses and possibly in inducing primary immune responses to viral pathogens and to tumours.

A major advantage of DNA vaccination is the ability to induce both humoral and cellular immune responses. DNA vaccines are currently used in veterinary medicine, but their tendency to display low immunogenicity in humans has hindered their usage, despite excellent tolerability and safety profiles. Various approaches have been used to improve the immunogenicity of DNA vaccines. Recent human study data re-established the value of DNA vaccines, especially in priming high-level antigen-specific antibody responses. Data suggests that innate immune responses to the DNA vaccine plasmid itself contribute to the immunogenicity of DNA vaccines, however the underlying mechanisms responsible remain unclear. In this dissertation, we investigate the role of innate immunity in shaping antigen-specific adaptive immune responses following DNA vaccination. The current belief is that the cytosolic DNA sensing pathways govern DNA vaccine immunogenicity. To date, only the type I interferon inducing STING/TBK1 regulatory pathway has been identified as required for DNA vaccine immunogenicity. Surprisingly, neither the upstream receptor nor the downstream signaling molecules in this pathway have been characterized. I therefore investigated a candidate cytosolic DNA receptor, as well as the downstream transcription factors required for generation of antigen-specific immune responses. Additionally, the effects of pro-inflammatory signaling on DNA vaccine immunogenicity have yet to be comprehensively studied. Previous studies have only provided indirect evidence for the role of inflammatory v signaling in DNA vaccination. As such, I also investigated the role of the DNA sensing AIM2 inflammasome in DNA vaccination. My data indicates that AIM2 is a key modulator in DNA vaccination via a previously unrecognized connection to type I interferon. Importantly, this marks the first time a DNA vaccine sensor has been identified. Of note, this dissertation represents a departure from many published works in the field. Whereas previous studies have mostly utilized model antigens and only focused on the adaptive immune responses generated, I analyzed the effects on innate immunity as well. Using various innate gene knockout murine models, I quantified antigen-specific humoral and T cell responses, as well as serum cytokine and chemokines following immunization with a clinically relevant DNA vaccine. Overall, this data provides a basis for understanding the mechanisms of DNA vaccination, allowing for the design of more effective vaccines.

A major advantage of DNA vaccination is the ability to induce both humoral and cellular immune responses. DNA vaccines are currently used in veterinary medicine, but their tendency to display low immunogenicity in humans has hindered their usage, despite excellent tolerability and safety profiles. Various approaches have been used to improve the immunogenicity of DNA vaccines. Recent human study data re-established the value of DNA vaccines, especially in priming high-level antigen-specific antibody responses. Data suggests that innate immune responses to the DNA vaccine plasmid itself contribute to the immunogenicity of DNA vaccines, however the underlying mechanisms responsible remain unclear. In this dissertation, we investigate the role of innate immunity in shaping antigen-specific adaptive immune responses following DNA vaccination. The current belief is that the cytosolic DNA sensing pathways govern DNA vaccine immunogenicity. To date, only the type I interferon inducing STING/TBK1 regulatory pathway has been identified as required for DNA vaccine immunogenicity. Surprisingly, neither the upstream receptor nor the downstream signaling molecules in this pathway have been characterized. I therefore investigated a candidate cytosolic DNA receptor, as well as the downstream transcription factors required for generation of antigen-specific immune responses. Additionally, the effects of pro-inflammatory signaling on DNA vaccine immunogenicity have yet to be comprehensively studied. Previous studies have only provided indirect evidence for the role of inflammatory v signaling in DNA vaccination. As such, I also investigated the role of the DNA sensing AIM2 inflammasome in DNA vaccination. My data indicates that AIM2 is a key modulator in DNA vaccination via a previously unrecognized connection to type I interferon. Importantly, this marks the first time a DNA vaccine sensor has been identified. Of note, this dissertation represents a departure from many published works in the field. Whereas previous studies have mostly utilized model antigens and only focused on the adaptive immune responses generated, I analyzed the effects on innate immunity as well. Using various innate gene knockout murine models, I quantified antigen-specific humoral and T cell responses, as well as serum cytokine and chemokines following immunization with a clinically relevant DNA vaccine. Overall, this data provides a basis for understanding the mechanisms of DNA vaccination, allowing for the design of more effective vaccines.

Gene directed enzyme prodrug therapy using E.coli the enzyme nitroreductase (NR) to activate the prodrug CB1954, is being developed as an attractive targeted chemotherapy for eradication of localized tumours. In addition to direct killing of NR-expressing tumour cells and potentially also their immediate neighbours via local spread of the activated prodrug, the consequent release of tumour antigens from dying tumour cells has the potential to induce antitumour immune responses. The present study investigates the capacity of NR/CB1954-mediated tumour cell death to activate CD8\(^+\) T cell responses using ovalbumin (OVA), as a model tumour antigen. The transgenic adenocarcinoma mouse prostate tumour cell line (Tramp-C1) was modified to stably express the therapeutic NR gene together OVA. These modified tumour cells were used to seed tumours in mice and OVA-specific T cell responses to gene therapy were investigated. Treatment of mice bearing NR-expressing tumours with CB1954 enhanced expansion of endogenous OVA-specific CD8\(^+\) T cells and marginally enhanced OVA-specific cytotoxic T lymphocyte (CTL) activity, however long-term CD8\(^+\) T cell dependent immunity was insignificant. The possibility of enhancing NR/CB1954-mediated long-term antitumour immune responses by combining with other immunogene therapies namely, 4-1BB costimulatory ligand (4-1BBL) or granulocyte macrophage colony stimulation factor (GM-CSF) was further explored. These combined therapies notably increased the frequency of memory OVA-specific CD8\(^+\) T cell and CTL response in some lymphoid tissue relative to NR/CB1954 monotherapy. One of the obstacles to cancer immunotherapy is the development of T cell anergy early in the course of tumour progression, therefore it was of interest to investigate the potential of NR/CB1954 and 4-1BBL combined tumour therapy to reverse CD8\(^+\) T cells anergy in vivo. This study describes preliminary results showing the effect of this combined therapy on the proliferative and functional responsiveness of anergic CD8\(^+\) T cells. In conclusion, these findings indicate that NR/CB1954-mediated tumour cell death is a weakly immunogenic process that facilitates short-term antitumour CD8\(^+\) T cell responses. Combining NR/CB1954 with intratumoural GM-CSF or 4-1BBL immunotherapy can enhance the frequency and effector function of memory tumour antigen-specific CD8\(^+\) T cells; and thus has the potential to provide long-term antitumour immunity.

Both HIV-1 and HIV-2 can cause the Acquired Immunodeficiency Syndrome (AIDS), yet the majority of patients infected with HIV-2 are clinically well and most never experience detrimental effects from the infection. Lessons learned from role of the immune system in the long-term non-progression characteristic of HIV-2 infection could offer insight into how a balanced immune response can protect from the immune system destruction associated with HIV-1 infection. The studies presented in this thesis were carried out in a community-based cohort of HIV-2 long-term non-progressors (LTNPs) in rural Guinea Bissau. The initial work (Chapter 3) outlines the first comprehensive analysis of HIV-2-specific immune responses. The data identified Gag as the most immunogenic HIV-2 protein. The magnitude of interferon gamma (IFN-y) generated against Gag was inversely related to virus load (VL). The most frequently recognised HIV-2 peptides clustered within a defined region of Gag with responses to a si~gle peptide associated with low VL. Given this information, the delineation of HIV-2-specific immune responses was focused on this immunodominant region of the HIV-2 proteome. Included in the detailed analysis was the identification of novel HIV-2 epitopes and ex vivo characterisation of HIV-2-specific T cell function (Chapter 4), in vitro characterisation of HIV-2-specific COB T cell clones (Chapter 5), as well as ex vivo phenotypic analysis of HIV-2-specific COB T cells (Chapter 6). Systemic immune activation is a hallmark of disease progression in both HIV-1 and HIV-2 infection. The final chapter (Chapter 7) examines the contribution of HIV-2 plasma antigenaemia to immune system activation which to date was incompletely defined. Cellular and plasma markers of immune activation were related to clinical (VL and CD4 T cell counts), immunological (HIV-2-specific IFN-y secretion) and virus sequence (p26 phenotype) correlates of protection from progression to AIDS. The aim of this work was to understand better what aspects of the host immune response in chronic HIV-2 infection contribute to the relative maintenance of the immune system during the course of this infection.

Les communications cellulaires sont indispensables au bon fonctionnement des organismes multicellulaires, notamment pour s’adapter à un environnement changeant en permanence. Les cellules du système immunitaire n’échappent pas à cette règle mais les interactions entre cellules immunitaires restent peu connues et compliquée à étudier. La récente apparition des technologies de séquençage dites ‘cellules uniques’ représente une opportunité unique pour étudier ces communications. Dans cette thèse, différentes approches expérimentales et analytiques ont été développées pour étudier ces communications à une échelle de cellules uniques. Ces stratégies ont ensuite été appliquées à différents contextes pathologiques, incluant le COVID-19, la maladie d’Alzheimer ou une immunisation par des pathogènes inactivés, et ont permis d’identifier des voies de communications cellulaires jusqu’ici inconnues ou mal comprises. Néanmoins, l’efficacité de ces approches est limitée par l’absence d’informations sur la localisation des cellules et des travaux supplémentaires intégrant ce genre de données est essentiel pour aller plus loin dans la dissection des communications entre cellules immunitaires
Cellular communications are essential to the proper functioning of multi-cellular organisms, particularly in order to adapt to a constantly changing environment. The cells of the immune system are no exception to this rule, but the interactions between immune cells remain little known and complicated to study. The recent emergence of 'single cell' sequencing technologies represents a unique opportunity to study these communications. In this thesis, different experimental and analytical approaches have been developed to study these communications on a single cell scale. These strategies were then applied to different disease contexts, including COVID-19, Alzheimer's disease or immunisation with inactivated pathogens, and identified previously unknown or poorly understood cellular communication pathways. However, the effectiveness of these approaches is limited by the lack of information on cell location and further work integrating such data will be essential to go further in the dissection of immune cell communications

The fetus and its placenta, collectively called the conceptus, are semi-allogeneic to the mother, as they express transplantation antigens of paternal origin. Foreign tissues generally experience immunological rejection by the host immune system; however in a normal healthy pregnancy the conceptus does not undergo immune attack. Emerging evidence indicates the conceptus avoids rejection through a number of mechanisms including the induction of active maternal immune tolerance specific for paternal antigens. However, the mechanisms responsible for establishing this tolerance remain undefined, including the timing of the first encounter with paternal antigen and the cellular processes by which paternal antigen is presented to the maternal immune system. Exposure to paternal transplantation antigens occurs in two waves: initially in the context of male seminal fluid at conception, and secondly after placental trophoblast invasion of maternal tissues in mid-gestation pregnancy. Therefore the aim of this research was to evaluate the female immune response to paternal antigens in seminal fluid and those associated with the conceptus. The mechanisms of antigen presentation, the impact of the cytokine environment and the consequences of T cell activation on pregnancy were also investigated. A transgenic system using ovalbumin (OVA) as the model paternal antigen was established. The transgenic Act-mOVA mouse expresses OVA constitutively and ubiquitously under a B-actin promoter and OVA was shown to be present in seminal fluid and in the fetal and placental tissue of sired progeny. The OVA-reactive CD8+ OT-I and CD4+ OT-II T cells were employed to gauge the relative amount of OVA antigen presented, with the strength of the maternal immune response quantified based upon the extent of T cell proliferation, as assessed by CFSE dye-dilution. Utilising bone marrow chimeric mice, it was demonstrated that upon insemination by an Act-mOVA male, seminal fluid-derived OVA was processed and indirectly presented by maternal bone marrow-derived antigen presenting cells to induce activation and proliferation of the CD8+ OT-I T cells within the uterinedraining para-aortic lymph nodes of the female. Likewise, OT-II T cells were responsive to MHC class II-restricted presentation of seminal fluid OVA. Post-implantation conceptus-derived OVA was detected within peripheral lymph nodes and the spleen where it was presented via the MHC class I and class II-restricted pathways to induce systemic proliferation of both OT-I and OT-II T cells. Furthermore, as gestation advanced the extent of OVA presentation and hence T cell proliferation intensified. Conceptus-derived OVA was still presented systemically until 20 days pp. The impact of the uterine cytokine environment was assessed to determine its influence on seminal OVA antigen processing and presentation. Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a key factor in regulating the leukocyte population of the female reproductive tract. GM-CSF-deficient female mice were unable to process and present seminal fluid OVA as effectively or efficiently as their wildtype counterparts, as assessed by their reduced capacity to drive OT-I and OT-II T cell proliferation following insemination by an Act-mOVA male. Finally, with highly-reactive OVA-specific T cells activated in response to seminal and conceptus OVA antigen, it was of interest to determine the effect of OT-I T cell activation on fetal survival and pregnancy success. It was found that OT-I T cells activated in vivo to paternal OVA antigen in the context of seminal fluid and pregnancy were not deleterious to pregnancy outcomes. However the transfer of cytotoxic OT-I T cells generated in vitro in the presence of an IL-2 into female mice carrying OVA-expressing conceptuses was detrimental to fetal survival. Collectively these experiments demonstrated that the initial exposure to paternal antigen, and hence the first opportunity to develop paternal antigen-specific tolerance, occurs at insemination. Paternal antigen is presented to the maternal T cell repertoire throughout gestation and may play a role in maintaining immune tolerance during pregnancy. The processing and presentation of paternal-derived antigen is chiefly performed by female bone marrow-derived antigen presenting cells. The cytokine environment of the mated female reproductive tract is critical in allowing optimal antigen processing and presentation, to generate an immune response consistent with maternal immune tolerance of the conceptus.
Thesis (Ph.D.) - University of Adelaide, School of Paediatrics and Reproductive Health, 2008

碩士
國立臺灣大學
公共衛生學研究所
83
Recently, asthma incidence rate is getting higher in different parts of the world, In addition, air pollutants are considered to be highly correlated with asthma incidence. This current investigation evaluated the influence of ozone on antigen-specific IgE, IgG1 and IgG2a antibody responses. The test mice, BALB/c and B6, were exposed to ozone 0.1, 0.5 and 1 ppm with aerosolized antigen OVA-TNP. The results indicated that 0.1ppm ozone enhanced the anti-OVA-TNP IgE and IgG1 antibody producitons of the BALB/c mice. Honever, 0.5 ppm and 1 ppm ozone was observed to inhibit, antibody responses in some cases. On the other hand, there was no significant difference of 0.5 ppm ozone in productions of antigen specific IgE, IgG1, or IgG2a antibody productions in intraperitoneallysensitised mice.