Effects ofAnaplasma phagocytophilaon NADPH Oxidase Components in Human Neutrophils and HL-60 Cells

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Abstract
The human granulocytic ehrlichiosis agent,Anaplasma phagocytophila, resides and multiplies exclusively in cytoplasmic vacuoles of granulocytes.A. phagocytophilarapidly inhibits the superoxide anion (O2−) generation by human neutrophils in response to various stimuli. To determine the inhibitory mechanism, the influence ofA. phagocytophilaon protein levels and localization of components of the NADPH oxidase were examined.A. phagocytophiladecreased levels of p22phox, but not gp91phox, p47phox, p67phox, or P40phoxreactive with each component-specific antibody in human peripheral blood neutrophils and HL-60 cells. Double immunofluorescence labeling revealed that p47phox, p67phox, Rac2, and p22phoxdid not colocalize withA. phagocytophilainclusions in neutrophils or HL-60 cells, and p22phoxlevels were also reduced.A. phagocytophiladid not prevent either membrane translocation of cytoplasmic p47phoxand p67phoxor phosphorylation of p47phoxupon stimulation by phorbol myristate acetate. The inhibitory signals for O2−generation was independent of several signals required forA. phagocytophilainternalization. These results suggest that rapid alteration in p22phoxinduced by binding ofA. phagocytophilato neutrophils is involved in the inhibition of O2−generation. Absence of colocalization of NADPH oxidase components with the inclusion further protectsA. phagocytophilafrom oxidative damage.