Program in Human Neutrophils Fails to Induce an Apoptosis Differentiation Anaplasma phagocytophilum Mechanisms: Insights into Pathogen Immune Evasion

Polymorphonuclear leukocytes (PMNs or neutrophils) are essential to human innate host defense. However, some bacterial pathogens circumvent destruction by PMNs and thereby cause disease. Anaplasma phagocytophilum, the agent of human granulocytic anaplasmosis, survives within PMNs in part by altering normal host cell processes, such as production of reactive oxygen species (ROS) and apoptosis. To investigate the molecular basis of A. phagocytophilum survival within neutrophils, we used Affymetrix microarrays to measure global changes in human PMN gene expression following infection with A. phagocytophilum. Notably, A. phagocytophilum uptake induced fewer perturbations in host cell gene regulation compared with phagocytosis of Staphylococcus aureus. Although ingestion of A. phagocytophilum did not elicit significant PMN ROS, proinflammatory genes were gradually up-regulated, indicating delayed PMN activation rather than loss of proinflammatory capacity normally observed during phago-cytosis-induced apoptosis. Importantly, ingestion of A. phagocytophilum failed to trigger the neutrophil apoptosis differentiation program that typically follows phagocytosis and ROS production. Heat-killed A. phagocytophilum caused some similar initial alterations in neutrophil gene expression and function, which included delaying normal PMN apoptosis and blocking Fas-induced programmed cell death. However, at 24 h, down-regulation of PMN gene transcription may be more reliant on active infection. Taken together, these findings suggest two separate antiapoptotic processes may work concomitantly to promote bacterial survival: 1) uptake of A. phagocytophilum fails to trigger the apoptosis differentiation program usually induced by bacteria, and 2) a protein or molecule on the pathogen surface can mediate an early delay in spontaneous neutrophil apoptosis. P olymorphonuclear leukocytes (PMNs 3 or neutrophils) are a first line of defense in the human innate immune response to bacterial pathogens. Most ingested bacteria are killed by the combined effects of PMN reactive oxygen species (ROS) and cytotoxic granule components (1). However, some pathogens have evolved means to circumvent killing by PMNs and cause disease. The mechanisms used by pathogens to evade destruction by the innate immune system are incompletely characterized. Anaplasma phagocytophilum, the agent of human granulocytic anaplasmosis, is an obligate intracellular bacterium known to survive within PMNs (2, 3). A. phagocytophilum enters neutrophils primarily through a receptor-mediated endocytic pathway (4), and thus may ultimately reside in a modified endosomal compartment (5). Studies evaluating functional alterations in A. phagocytophi-lum-infected PMNs suggest that the pathogen delays PMN apo-ptosis (6, 7), minimizes proinflammatory cytokine release (8, 9), and inhibits and/or fails to activate ROS production (10 –13). Mechanisms underlying the ability of A. phagocytophilum to inhibit …