Microbiology & Immunology - Theses
Permanent URI for this collection
Search Results
1 - 5 of 5
-
ItemDissecting the role of gd T cells in T cell priming for liver stage immunity( 2023-11)Liver resident memory T cells (TRM) are poised for protection against repeat infection and rapidly form a robust defence against tissue-specific insults such as liver stage malaria. A direct correlation between liver stage immunity and gd T cells has been observed both in mice (Zaidi et al. 2017) and in humans (Seder et al. 2013; Ishizuka et al. 2016), but the precise molecular mechanisms by which these gd T cells exert their protective effect are yet to be defined. In mice, intravenous injection with radiation-attenuated sporozoites (RAS) confers sterile protection against challenge with live sporozoites. This protection is mediated by responding antigen-specific CD8+ and CD4+ T cells that migrate to the liver and form resident-memory T cells (TRM). In the absence of gd T cells, protective CD8+ liver TRM are not generated, leaving mice susceptible to reinfection. Using Plasmodium-specific T cells as a readout for effective immunity, we determined that IL-4 is important for the accumulation of CD8+ and CD4+ T cells. By utilising complex in vivo systems including mixed-bone marrow chimeras and adoptive transfer of gd T cells, we revealed that gd T cell-derived IL-4 is crucial for the expansion of antigen-specific CD8+ T cells. In addition, in vivo neutralisation of IL-12 or IFN-g confirmed a partial dependency for these cytokines, despite their traditionally opposing function to IL-4. Given IL-4, IFN-g and IL-12 all have a clear role in CD8+ T cell priming following RAS vaccination, we hypothesised that IL-4 and IFN-g synergise to enhance cDC1 activity. These findings led to our development of a novel model to reconstitute cDC1-deficient mice using CRISPR-edited primary dendritic cells. This model enabled the investigation of the mechanism by which gd T cell derived IL-4 leads to DC activation and therefore effective CD8+ T cell expansion for memory development. Collectively, this project has shown a significant role for IL-4 in the priming of malaria-specific CD8+ T cells and demonstrates a novel pathway for collaboration between gd T cells, cDC1s, and CD8+ T cells, revealing the potential for harnessing gd T cells in vaccination strategies against malaria.
-
ItemFunctional and cellular heterogeneity of the myeloid cell system( 2019)Cells of the myeloid lineage form the innate part of the immune system and are characterized by a high level of functional plasticity, which is required to address the diverse set of functions of these mononuclear cells. Monocytes, Macrophages and dendritic cells (DC) are collectively categorized as the mononuclear phagocyte system (MPS), to highlight their functional equipment that specializes them to the phagocytosis of pathogens as a starting point to elicit an immune response. Besides this role, cells of the MPS are also involved in a wide variety of homeostatic functions including early development and regulation of physiological processes. However, the multitude of mechanisms required to acquire this functional plasticity remains poorly understood. The work that has been performed in the scope of this dissertation aimed to advance current knowledge of the causes and consequences of functional and cellular plasticity of the myeloid immune system. High-dimensional characterization of the effects of a Western diet on myeloid immune cell progenitor cells revealed a long-term transcriptional and epigenetic reprogramming of the myeloid cell compartment. The formation of an innate immune memory in myeloid progenitor cells leads to lasting inflammatory priming of monocytes, which may directly contribute to the progression of myeloid cell-associated diseases. In addition, single-cell RNA-seq elucidated unreported cellular heterogeneity of the monocyte and dendritic cell compartment in human peripheral blood. A combination of phenotypic and transcriptional analyzes resulted in a precise categorization of the human DC compartment consisting of pDCs, cDC1, two cDC2 subsets, and a deeply characterized preDC subset. Furthermore, a universal strategy for the integration of cellular atlases was conceptualized and applied to establish a consensus map of the human DC and monocyte cell space. This thesis provides mechanistic insights into the cellular composition of myeloid cells and their functional plasticity, which will form the foundation for further investigations into the dynamic changes of the immune cell compartment during diseases and will be critically relevant for designing effective treatments for a wide variety of pathologies linked to myeloid cells.
-
ItemCharacterization of roles of IRF8 in dendritic cell development and function( 2018)Dendritic cells (DC) are an essential component of immune system. Antigen presentation by DCs initiates T cell responses and builds up defense against pathogens. There are two major types of DC, conventional DCs (cDCs) and plasmacytoid DCs (pDCs), both of which derive from the same progenitor. The cDCs can be further classified by surface marker CD11b and CD8. While cDCs function as the major antigen presenting cells, pDCs are more efficient in type I IFN production. Regarding the functional difference, it is important to comprehend factors that control the production of these two types of DC. It has been reported that IRF8 mutation in human causes selective loss of DC subsets. Although IRF8 has been identified as key regulator in DC transcription network, study addressing the function of IRF8 in DC development and function remains incomplete. In this project, we show that the generation of pDCs and cDC1s depends largely but not entirely on IRF8. Upon germline deletion of IRF8, normal function of pDCs was disrupted while the antigen presentation of cDC1s was complemented by other mechanisms.
-
ItemUsing dendritic cell receptors to enhance immunity( 2017)Dendritic cells (DCs) are the most potent initiators of immune responses, being highly specialised for the uptake and presentation of antigens (Ag) to activate T cells. Their priming potential can be harnessed to generate stronger immune responses by targeting Ag to DCs via monoclonal antibodies (mAbs) specific for DC-expressed surface receptors. This thesis builds upon the concept of targeting DCs in two main ways: firstly, by investigating a novel method of targeting adjuvant to DCs, and secondly, by investigating how DC-targeting constructs can be used to prime and boost responses. It was considered whether not only Ag, but also adjuvants could be targeted to DCs to improve their efficacy. A recent finding that the DC receptor DEC-205 can bind to and mediate the immunostimulatory effects of CpG oligonucleotide (ODN) adjuvants led to the hypothesis that CpG ODNs could be targeted to DCs via DEC-205 in order to enhance their potency. The interaction between DEC-205 and CpG ODNs was further characterised to determine the molecular properties of ODNs required for binding. This information was then used to enhance the DEC-205 binding capacity of a particular CpG ODN that normally only weakly binds DEC-205. Enhanced DEC-205 binding was found to significantly improve the stimulatory capacity of this ODN, demonstrating that targeting adjuvant to DCs could be a viable method to improve adjuvant potency. Another receptor, CD14, has also been reported to bind CpG ODNs, so the potential for CD14 to act in synergy with DEC-205 was investigated. However, CD14 was not observed to mediate the uptake or stimulatory effects of CpG ODNs. The identification of natural ligands of DEC-205 is critical for understanding its physiological function. Although ODNs are synthetic molecules, their binding to DEC-205 may signify that DEC-205 is capable of binding other types of DNA that structurally resemble ODNs. A panel of biological DNA samples was screened for DEC-205 binding. While none of the DNA samples were observed to bind DEC-205, some DNA samples were found to bind another receptor, RAGE, suggesting a role for RAGE as a detector of both pathogenic and self-DNA. Most vaccines must be administered more than once, or “boosted”, to achieve optimal efficacy, and DC-targeted vaccines should be no exception. However, our data suggested that simply administering the same DC-targeting construct twice does not effectively boost the response. This was due to interference from the primary antibody response, which can cross-react with and neutralise a subsequently administered boosting construct. To overcome this issue, the efficacy of various heterologous prime-boost strategies designed to reduce the reactivity of the primary response against the boosting construct was assessed. Ultimately, a combination of anti-Clec9A and anti-XCR1 targeting constructs was found to induce the least cross-reactivity and strongest response after boosting. These findings contribute to the development of better adjuvants and immunisation strategies that optimise the efficacy of DC-targeted vaccines. More broadly, they also highlight the value of understanding the underlying biological mechanisms that drive immune responses, which can then be applied to the rational design of more effective vaccines.
-
Item