The role of natural killer cells in adaptive immune responses

Human natural killer (NK) cells have been shown to respond to numerous pathogenic stimuli, producing IFN-y as well as cytolytic effector molecules such as perforin and granzymes. Previous work on NK cell responses to Plasmodium fa/ciparum-infected erythrocytes (Pf-RBC) has shown that these responses are dependent on contact with accessory cells, such as macrophages and myeloid-lineage dendritic cells (mDCs), as well as on soluble mediators such as IL-2, IL-12, IL-15, IL-18 and IFN-a. It has also been observed that these responses are heterogeneous between donors; part of this heterogeneity seems to be genetically encoded, depending for example on killer immunoglobulin-like receptor (KIR) genotype, but heterogeneity in accessory cell stimuli has also been observed. The work described in this thesis further dissects the NK cell response to Pf-RBC in malaria na·ive donors, with a focus on describing the role of T cells in NK cell responses to pathogens. My data demonstrate that while NK cells are dependent upon accessory cell signals, they are also highly dependent upon signals emanating from antigen-specific CD4+ T cells; T cell help for NK cell responses is MHC class II-dependent, IL-2-dependent and contact-independent. I have observed significant production of IL-2 from CD4+ T cells in the very early hours of co-incubation of peripheral blood mononuclear cells (PBMC) and Pf-RBC, preceding NK cell-derived IFN-y. Having documented T cell-dependent NK cell activation by Pf-RBC in malaria na"ive donors, I next explored whether similar T cell-dependent NK cell responses were observed in African children undergoing vaccination with the candidate preerythrocytic malaria vaccine, RTS,S. I characterized expression of CD69 and production of IFN-y in NK cells and T cells and expression of CD25 in T cells. My data demonstrate that robust recall NK cell and T cell responses are mounted during in vitro re-stimulation with the RTS,S vaccine antigen, Hepatitis B surface antigen (HBs) peptides and while circumsporozoite (CS)-induced IFN-y responses were not as strong, expression of CD25 in T cells and CD69 in NK cells and in T cells were significantly higher in RTS,S vaccinated children than in rabies vaccinated controls. Nearly half of the IFN-y was derived from NK cells. I also measured secreted levels of IFN-y, IL-2 and IL-10 in culture supernatants. I observed high levels of IFN-y in culture supernatants of RTS,S vaccinated PBMC only in response to HBs peptides, however, there was strong vaccine antigen-specific IL-2 production to both HBs and CS peptides, which was significantly more robust in RTS,S vaccinees than in the rabies vaccinated controls. Finally, my data demonstrate that the IL-2 secretion in response to HBs and CS peptides was highly correlated with the early activation of NK cells (expression of CD69). Finally, to formally test the hypothesis that antigen-specific CD4+ T cells can enhance NK cell responses to pathogens, I carried out a study of the T cell and NK cell response to heat-killed rabies virus in individuals undergoing rabies vaccination. The results of this study demonstrate that vaccine antigen-specific CD4+ T cells induced by vaccination can recruit NK cells to secrete IFN-y, to degranulate, to release perforin and to proliferate. The post-vaccination NK cell response is detected within 6-12 hours after re-exposure to rabies virus and, somewhat unexpectedly, is sustained for at least 7 days, well after the T cell response is underway. Importantly, in the first 24 hrs after re-exposure to virus, NK cells represent more than 70% of the IFN-y secreting effector cells, indicating a potentially very important role for NK cells in the early phase of the post-vaccination effector response. Ever since the immune system was dichotomized into innate and adaptive arms, vaccine-induced immunity has been explained solely in terms of priming of effector and memory Band T lymphocytes. The potential for innate immune celis to contribute to enhanced cytotoxicity or cytokine production post-vaccination has been almost completely overlooked. My data suggest that IL-2 secreting CD4+ T celis and NK cell activation markers, such as IFN-y production, expression of CD69 and CD25, upregulation of the lysosome-associated membrane protein (LAMP)-1 and release of perforin, may prove to be more reliable indicators of vaccine efficacy than simply counting the numbers of IFN-y-secreting PBMCs and that these parameters need to be considered for inclusion of future protocols for evaluating vaccine immunogenicity and efficacy in clinical trials.