SACM - United Kingdom
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Item Restricted B cells selection in Germinal center(University of Birmingham, 2024-02) Alzhrani, Khaled Rashed; Toellner, Kai-MichealVaccination triggers the long-term production of high-affinity antibodies. Germinal centers (GCs) are sites within lymphoid tissues where B cells undergo clonal expansion and affinity maturation during T cell-dependent antibody responses. However, the regulation of affinity-matured B cell selection within GCs remains poorly understood. In this study, we examined the roles of antibody secretion and feedback in GC regulation using various mouse models: those incapable of secreting IgM or undergoing class-switching (IgMi), those unable to undergo affinity maturation or class-switching (AIDKO), and those restricted to producing and secreting IgG1 antibodies (IgG1M). Following immunisation with a T cell-dependent (TD) antigen, IgMi and AIDKO mice exhibited an increased percentage of GC B cells. In contrast, the percentage of GC B cells in IgG1M mice was reduced compared to wild-type mice. Introducing antibody feedback by injecting antigen-specific IgM into IgMi or AIDKO mice slightly reduced the proportion of GC B cells. It was shown that more stringent antibody feedback strongly inhibits B-Tfh cell interactions. The Nr4a3-Tocky reporter system is unstable fluorescent protein that changes fluorescence from blue to red and can record the timing of TCR signalling. It was utilized to track interactions between GC B cells and Tfh cells in mice with altered B cell receptors (IgMg1 and IgG1M mice) or absence of affinity maturation (AIDKO mice). IgMg1 enhances interaction with and activation of Tfh cells, also reflected by higher Tfh frequencies. Additionally, testing expression of Nr4a3-blue and red forms of the reporter reveals an increase in newly activated Tfh cells in IgMg1 and IgG1M versus wildtype mice after immunisation. There was no change in Nr4a3-reporter expression in Tfh cells of AID-deficient mice, indicating unaffected Tfh cell activation in AID-deficient GCs. This work gives insight, by using mouse models with altered antibody feedback, into how IgM and IgG1 antibody secretion shapes GC developments and how altering BCR isotypes and signalling regulates Tfh cell activity through antigen presentation within the GC.14 0Item Restricted Host parasite interactions in Trichuris infections(The University of Manchester, 2024-06-27) Othman, Abdulrazzag Abdulaziz A; Else, Kathryn J; Hepworth, MatthewInfection with T. trichiura affects around 500 million people worldwide. Currently, no effective vaccines against worm infection are available to prevent infection of humans. The murine species of Trichuris, Trichuris muris, serves as an excellent model of human trichuriasis, and is used widely to study the host immune response against Trichuris. It is well documented that resistance to T. muris infection in mice requires a T helper 2 (Th2) type of immune response with T helper 1 (Th1) cells dominating in susceptible individuals. Understanding how Th2 immune responses are generated, is an unmet need which will facilitate vaccine development. It was previously thought that B cells are not essential for worm expulsion. However, our lab has recently shown that B cells play an important role in the generation of protective immunity. Thus B cells not only important in antibody production but also play a regulatory role in the generation of protective Th2 responses to Trichuris muris. This PhD thesis explores the cellular sources of interleukin-10 (IL-10) during T. muris infection, focusing on subsets of T cells, macrophages, and B cells using the IL-10 reporter (Vert-X) mice. Follicular B cells and marginal zone B cells were found to be the main source of IL-10 in the mesenteric lymph nodes at day 21 post-infection. This pattern was shifted at day 35 post-infection, when B1a, B1b, and marginal zone B cells became the main IL-10 producers. These data suggest the importance of B cell-derived IL-10 in controlling immunopathology and facilitating long term immune regulation. In large intestine, follicular B cells were identified as the primary IL-10 producers in the lamina propria leukocytes at day 21 post-infection. Building on these findings, the thesis goes on to investigate the role of B cell-derived IL-10 in immune responses and pathology in a low dose Trichuris muris infection using a transgenic CD19creIL10flfl mouse model that lacks IL-10 production in B cells. Notably, the absence of B cell-derived IL-10 made mice less able to support a chronic infection, without altering gut pathology. Interestingly, the absence of B cell-derived IL-10 was associated with a reduction in IL-6 mRNA and in some cases reduced IFN-γ levels in the gut, alongside an increased in FoxP3+ Treg cells. The study also extended to exploring the importance of B cell-derived IL-10 in the context of a high dose T. muris infection. No significant differences were identified between mice where B cells could and could not make IL-10 in terms of gut pathology and the kinetics of worm expulsion. However interestingly, in the absence of B cell derived IL-10 mice had high numbers of M2 macrophages in the gut tissue which correlated with non-significantly reduced levels of IFN-γ mRNA. This research has informed our understanding of the contribution of B cell-derived IL-10 to the host immunity to T. muris. It highlights the necessity for further investigation to understand the mechanisms by which the B cell-specific IL-10 deficient mouse presents with elevated intestinal numbers of Foxp3+ Treg cells and elevated M2 macrophages in the context of low and high dose T. muris infection respectively17 0