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A. phagocytophilum is a tick-transmitted obligate intracellular Gram-negative bacterium, which replicates within its host’s neutrophil granulocytes. The human and animal granulocytic anaplasmosis often manifests as a nonspecific febrile illness. The ability of A. phagocytophilum to use particularly neutrophil granulocytes as suitable hosts is made unique by the multitude of their antimicrobial defense mechanisms. It is therefore of considerable interest to more closely investigate the immunological control of A. phagocytophilum by neutrophil granulocytes. The chosen cell culture model for the analysis was murine Hoxb8-granulocytes, generated by retroviral transduction with the Hoxb8-oncogene. These generated cells exhibit functional similarity to primary cells. This technique enables a notably higher level of purity and allowed unlimited production in comparison with ex vivo produced murine neutrophil granulocytes.
Cytokines and chemokines are presumably involved in the pathogenesis of a A. phagocytophilum infection. The extent up to which the neutrophil granulocytes act as their producer during a A. phagocytophilum infection was until now unknown. This study shows that murine Hoxb8-granulocytes produce regulated on activation, normal T cell expressed and secreted (RANTES), macrophage inflammatory protein-1α (MIP-1α) and tumor necrosis factor (TNF), after stimulation with A. phagocytophilum. In vivo studies at the Institute of Microbiology and Hygiene had already showed that the effector mechanisms of neutrophil granulocytes, such as nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, inducible nitric oxide synthase (iNOS) and myeloperoxidase (MPO), were not required for pathogen elimination in vivo. This in vitro study demonstrates that the absence of these effector mechanisms has no influence on the growth of A. phagocytophilum in Hoxb8-granulocytes. This suggests that the pathogen has developed strategies to evade these defense mechanisms.
Interferon-γ (IFN-γ) is an important modulator of the neutrophil granulocyte function. The results of this study demonstrate that a direct IFN-γ-stimulation of murine Hoxb8-granulocytes leads to a direct growth inhibition of A. phagocytophilum. Despite a noticeable induction of the iNOS mRNA-expression, due to the infection and IFN-γ-stimulation, was this effect however iNOS-independent. In previous studies IFN-γ-deficient mice exhibited an increased bacterial load during the early phase of the infection, whereas the final pathogen elimination remained unimpaired. The conclusion of this study is that a direct IFN-γ-effect on neutrophil granulocytes is essentially involved in the in vivo observed protective effect.