The acquisition of dendritic cell tolerance during malaria infection results in differential T-cell activation.

摘要

This dissertation is focused on several aspects of the host immune response to Plasmodium yoelii, a murine malaria parasite. The studies described herein suggest that in situations where host and parasite are well adapted, as in our model using P. yoelii and B6/B10D2 mice, malaria infection guides the immune system to preferentially produce anti-inflammatory cytokines as infection progresses. When anti-inflammatory cytokines are not produced, such as in IL-10 knockout (KO) mice, severe pathology is seen.;Initial studies, as described in chapters 2 and 3 of this dissertation, demonstrated that CD4 T cell responses to an exogenous antigen (Ova) were diminished in infected mice. Subsequent work linked this immune dysfunction to a suppressive/inhibitory population of splenic macrophages. We then showed that the capacity of purified splenic dendritic cells (DCs) to stimulate IL-2 production and T cell proliferation was equivalent to that of DCs isolated from spleens of uninfected mice, and that inhibition of IL-2 production was recapitulated when splenic macrophages from infected mice were added back to cultures of purified DCs. The specific mechanism by which macrophages inhibit IL-2 production has yet to be identified but appears to be independent of IL-10, TGF-beta, nitric oxide, PGE2 and tryptophan catabolism.;When looking further at T cell responses induced by DCs from naive versus infected mice, we found that while IL-2 levels are comparable, the expression and secretion of inflammatory cytokines varied dramatically. In chapter 4, we show that the cytokine profiles obtained from T cells cultured with DCs from infected mice also varied significantly with stage of infection. Specifically, T cells activated by DCs from mice 3 days post infection (p.i.) produced high levels of IFN-gamma, TNF-alpha and little IL-10, whereas T cells stimulated by DCs from day 17 p.i. mice predominantly produced IL-10 with little accompanying IFN-gamma and TNF-alpha.;We then determined how the phenotype of DCs shapes T cell responses as a function of time post infection. An extensive series of cell sorting experiments indicated that DCs isolated from mice during the acute phase of infection (day 3 p.i.) produced much larger quantities of IL-12p40 and TNF-alpha in response to innate immune stimuli such as LPS and CpG DNA. As the infection progressed to day 17 p.i. however, purified DCs produced statistically smaller amounts of IL-12p40 and TNF-alpha while secreting larger amounts of IL-10 as measured both by in vitro stimulation assays as well as ex vivo mRNA analysis. These data are consistent with the previously described T cell stimulation assays where DCs isolated early during infection activate IFN-gamma producing T cells whereas DCs isolated late during infection induce IL-10 producing T cells. Thus malaria infection results in down-regulation of pro-inflammatory cytokines and up-regulation of anti-inflammatory cytokines.;In chapter 5 we attempted to determine the mechanism of regulation. We found that IL-10 is necessary for IL-12 downregulation, but not TNF-alpha. Failure to down-regulate IL-12 in IL-10 KO mice was especially evident in vivo following administration of LPS to naive and infected mice. Again, lack of IL-10 had little effect on the down regulation of TNF-alpha with in vivo LPS stimulation. The disparate regulation of IL-12 and TNF-alpha is currently being explored and appears independent of NF-kB p50 as well as the MAP kinases p38 and ERK 1/2.;These changes in cytokine production a malaria infection progresses correlates to clinical severity of disease. Downregulation of IL-12 (and IFN-gamma indirectly) by IL-10 protects the host from inflammation-induced pathology. Mice lacking IL-10 develop significantly more hepatic necrosis and greater anemia despite a lower parasite burden compared with wild type mice. Our findings are consistent with epidemiological data from human malaria infections where increased pro- to anti-inflammatory cytokine ratios are correlated with an increased severity of malaria syndromes such as severe malarial anemia, cerebral malaria and placental malaria.;The observation that DCs play a dominant role in guiding the activation of IL-10 producing anti-inflammatory T cells and that IL-10 is responsible for mitigating pathology during murine malaria infection provides a basis for the development of potential therapeutics utilizing this phenomenon. One hypothetical use for such information would be the production of an "anti-disease" vaccine where IL-10 producing DCs would be used to specifically activate an anti-inflammatory adaptive immune response. Clearly, more research will be necessary before such trials become feasible as there is a tight balance between too much inflammation and too little inflammation in the context of malaria infection both in humans and animals, alike. (Abstract shortened by UMI.)