title: Novel mechanisms of immune regulation during parasitic infections
Creators: name: Ng, Susanna
Publisher: name: Griffith University
Year: 2018
Subjects: subject: Parasitic infections
subject: Immune regulation
subject: Cytokines
subject: Vaccine development
subject: CD4+ T cell

description: The host immune response elicited during parasitic infections involves the release of potent pro-inflammatory cytokines. While essential for mediating parasite clearance, these pro-inflammatory cytokines cause tissue damage that are responsible for many disease symptoms associated with parasitic infections. The negative effects of pro-inflammatory cytokines are controlled by antiinflammatory cytokines. The balance between pro- and anti-inflammatory immune responses are regulated by different mechanisms that restrict the activation of effector cell populations, such as the expression of co-inhibitory receptors, the expansion of immunoregulatory cell populations and/or reprogramming of cells to produce anti-inflammatory cytokines. In some cases, these regulatory mechanisms are activated prior to control of infection and can suppress the development of host anti-parasitic immunity, thus preventing the establishment of long lasting immunity and promoting persistence of infection. These events, in turn, can have major consequences for disease outcome. In the case of Plasmodium infection, immunoregulatory mechanisms that are established in the host at a young age have been attributed to the diminished efficacy of malaria vaccines when tested in endemic settings. With Leishmania infection, immunoregulatory pathways have been proposed to cause visceral leishmaniasis (VL) and increase the likelihood of developing a complication of chronic L. donovani infection called post-kala-azar dermal leishmaniasis (PKDL). As such, a better understanding about the immunoregulatory mechanisms that develop during parasitic infections could have positive implications in the area of vaccine development and the use of immunotherapy to enhance host antiparasitic immunity. Given the importance of the innate immune system in priming, influencing and regulating adaptive immune responses, we first examined the role of group 1 innate lymphoid cells (ILCs) during Plasmodium infection. Group 1 ILCs have been termed the innate counterparts of T helper (Th)1 cells. The indispensable role of Th1 cells during Plasmodium infection led us to hypothesize that IFNγ- producing group 1 ILCs play an important role in the host immune response against Plasmodium infection. In this study, we showed that group 1 ILCs were rapidly lost during the early stages of Plasmodium infection in humans and mice. Additionally, our results indicated that ILCs are redundant in the anti-parasitic immune response against Plasmodium parasites. These findings are consistent with a recent report proposing the redundancy of ILCs in human adults. Next, we changed our focus to CD4+ T cells. CD4+ T cells isolated from the peripheral blood of VL patients expressed distinct molecular signatures, compared with 30 days after anti-parasitic drug treatment. A mouse model of L. donovani infection allowed us to investigate differences in CD4+ T cells in a state where parasites were cleared effectively (in the liver), and in instances when dysregulated CD4+ T cell responses caused parasite persistence (in the spleen). Our results showed characteristic molecular signatures of splenic and hepatic CD4+ T cells that distinguished these cells, even in naïve mice. We defined three molecular signatures of CD4+ T cells: a core, chronic and resolving signature, and interrogated the key molecules and pathways associated with each signature. Finally, we identified natural killer cell granule protein 7 (NKG7) as a molecule within the chronic signature for further characterisation and testing as a potential therapeutic target. Changes in the expression of NKG7 at the transcript level have been observed in a number of studies. However, to our knowledge, its role in immune cell responses has not been investigated. Using a membrane reporter mouse, we showed that the expression of Nkg7 transcripts in naïve mice was restricted to NK cells. However, in vitro polarisation of CD4+ T cells with IL-27 induced Nkg7 expression in a dose dependent manner. Conversely, we found that transforming growth factor (TGF)β suppressed IL-27-mediated expression of Nkg7. While expression of Nkg7 has been associated with many cytotoxic molecules, we found no evidence of Nkg7 being a marker of any specific CD4+ T cell subset, including recently described cytotoxic CD4+ T cells. Characterisation of Nkg7 expression in immune cell subsets indicated that CD4+ T cells comprised a significant proportion of Nkg7-expressing cells during L. donovani infection. Confocal immunofluorescence (IF) microscopy showed the localisation of CD4 and Nkg7 co-expressing cells in hepatic granulomas, highlighting their close proximity to infected Kupffer cells. We also found that Nkg7-deficient mice displayed increased parasite burdens and lower serum IFNγ, TNF, and MCP-1 levels, compared to wild-type control animals. This identified a novel protective role for NKG7 during L. donovani infection. Finally, we identified Mincle (encoded by CLEC4E) as a potential binding partner for NKG7 using computational methods. Overall, the results reported in this thesis identify and characterise novel mechanisms of immune regulation during parasitic infections. A better understanding of the functions of these regulatory molecules and pathways will contribute to our knowledge of anti-parasitic immunity and identify new molecules that can be manipulated to improve CD4+ T cell responses. These findings may help to develop better immunotherapies and vaccines to treat and protect against parasitic infections, respectively, as well as have broader applications beyond the context of parasitic infections.