Studies of the recognition and activation of natural killer cells

Abstract

Natural killer (NK) cells mediate part of the innate immune response that impedes the spread of invading microorganisms and shapes the downstream adaptive immune response. NK cells spontaneously kill tumor cells and other cells upon contact. Killing requires engagement of germline encoded receptors that mediate positive and negative signals. In addition to their constitutive cytotoxic capacity, NK cells may be activated under conditions created by infection or by exposure to tumour cells. Activated NK cells become more cytotoxic and produce innate cytokines, primarily interferon (IFN)-[gamma] and tumor necrosis factor (TNF)-[alpha]. Activation is regulated by other innate cytokines but also involve contact with target cells. Many viral, bacterial and protozoan infections are accompanied by activation of NK cells, and the presence of NK cells has been shown to enhance host survival in several experimental infections. The nature of these protective effects is not well understood and varies depending on the type of infection.

The general aim of this thesis was to study the prerequisites for activation of NK cells. Activation of NK cells implicate several features; increased cytotoxicity, cytokine production, chernotaxis, blastogenesis and proliferation. The study focused on activation of NK cell-mediated cytotoxicity and IFN-[gamma] production in response to different stimuli. For this purpose, the mechanisms causing activation of NK cells was studied in vivo after exposure to protozoan parasites as well as in vivo and in vitro on exposure to tumour cells.

Many proteins are anchored to the cell membrane by glycosyl-phosphatidylinositol (G-PI) moieties. Phospholipase C (PLC) cleaves G-PI-anchored molecules from the cell surface. Treatment with PLC reduced the susceptibility of some cells to NK cell killing. This suggests that a cell surface molecule, anchored by a G-PI moiety, mediates NK killing. A further analysis showed that PLC treatment interfered at the level of triggering the NK cells. It is thus likely that PLC acts by preventing a signal that activates the lytic machinery of NK cells upon binding.

In order to study prerequisites for activation of NK cells in response to the protozoan parasite Trypanosoma cruzi, we established a model which allowed us to follow NK cytotoxicity and expression of innate cytokine production during the first week of infection. T. cruzi induced increased NK cell cytotoxicity, which was dependent on IFN-[alpha]/[beta], as well as initial expression of NK cell dependent IFN-[gamma] in the spleen. Later production of IFN-[gamma] was dependent on CD4+ cells and IL-12. NK cell cytotoxicity and production of NK cell IFN-[gamma] were differentially regulated. Furthermore, endogenous IL-12, but not IFN-[alpha]/[beta], was necessary for survival. Thus, neither endogenous IFN-[alpha]/[beta], nor NK cell cytotoxicity are protective against T. cruzi.

Infection of BALB/c mice with Leishmania amazonensis lead to activation of NK cytotoxicity and IFN-[gamma] induction in draining lymph nodes. The expression of IFN-[gamma] was, however, independent of NK cells. Furthermore, depletion of NK cells in the initial phase of infection did not result in any significant exacerbation of the disease.

The MHC class I phenotype of NK target cells can determine their susceptibility to lysis. In order to find out if MHC class I expression can also influence the general activation state of NK cells, we inoculated mice intraperitoneally with turnout cells lacking MHC class 1. These cells induced both cytotoxic activation and NK cell-dependent IFN-[gamma] transcription in intraperitoneal NK cells. The responses were inhibited by restoration of MHC class I expression in the tumour cells. Thus, the MHC class I phenotype of NK target cells can regulate the activation of NK cells.

The pathways by which NK cells are activated and the importance of NK cells in protection are different in response to different challenges. This study provides some examples of mechanisms regulating NK cell functions.