Date Published: February 14, 2013
Publisher: Public Library of Science
Author(s): Elías Barquero-Calvo, Anna Martirosyan, Diana Ordoñez-Rueda, Vilma Arce-Gorvel, Alejandro Alfaro-Alarcón, Hubert Lepidi, Bernard Malissen, Marie Malissen, Jean-Pierre Gorvel, Edgardo Moreno, Renée M. Tsolis.
Polymorphonuclear neutrophils (PMNs) are the first line of defense against microbial pathogens. In addition to their role in innate immunity, PMNs may also regulate events related to adaptive immunity. To investigate the influence of PMNs in the immune response during chronic bacterial infections, we explored the course of brucellosis in antibody PMN-depleted C57BL/6 mice and in neutropenic mutant Genista mouse model. We demonstrate that at later times of infection, Brucella abortus is killed more efficiently in the absence of PMNs than in their presence. The higher bacterial removal was concomitant to the: i) comparatively reduced spleen swelling; ii) augmented infiltration of epithelioid histiocytes corresponding to macrophages/dendritic cells (DCs); iii) higher recruitment of monocytes and monocyte/DCs phenotype; iv) significant activation of B and T lymphocytes, and v) increased levels of INF-γ and negligible levels of IL4 indicating a balance of Th1 over Th2 response. These results reveal that PMNs have an unexpected influence in dampening the immune response against intracellular Brucella infection and strengthen the notion that PMNs actively participate in regulatory circuits shaping both innate and adaptive immunity.
Neutrophils are the first line of defense against microbial pathogens. Upon bacterial infection, these polymorphonuclear leukocytes (PMNs) become activated and are rapidly recruited to the infection site where they can efficiently constrain and kill microbes via phagocytosis, extracellular release of granule contents, cytokine secretion, and the formation of neutrophil extracellular traps . In addition to playing a primary role during the course of innate immunity against acute bacterial infections, PMNs may also influence adaptive immunity –. For instance, PMNs seem to compete with dendritic cells (DCs) and macrophages (Møs) for the availability of antigen , shed large membrane vesicles that inhibit the maturation of monocyte-derived DCs and monocyte-derived Møs ,  and are capable of negatively influencing B and CD4+ T cell responses activity . The role of PMNs as key regulators of NK cell functions has also been confirmed in patients with severe congenital neutropenia and autoimmune neutropenia . Therefore, in addition to their direct antimicrobial activity, mature neutrophils seem to be endowed with unsuspected immunoregulatory functions that seem to be conserved across species .
In order to determine the influence of murine PMNs in the immune response during B. abortus infection, a time course protocol was designed to include: neutrophil depletion, bacterial inoculation, spleen bacterial counts, histopathological examination and analysis of immune cells (Figure S1). As expected, at the second week of infection mice depleted of PMNs with anti-RB6 (PMN-depleted) already mounted an efficient response against the foreign monoclonal antibody, neutralizing the PMN depletion effect (Figure S1). In spite of this, experiments in PMN-depleted mice were carried out until 15 days to follow the outcome of the infection after the initial PMN depletion.
PMNs have been viewed as the most efficient effectors cells involved in acute inflammatory responses against bacterial infections. This perspective has been reinforced by the use of antibody-neutropenic mouse models in which bacteria such as Streptococcus pneumoniae, Acinetobacter baumannii, Salmonella enterica, Listeria monocytogenes, Yersinia enterocolitica, Staphylococcus aureus, Ochrobactrum anthrophi and Legionella pneumophila, among others were used as infecting agents, mostly with fatal consequences –, , . In addition, lethality due to increased proinflammatory responses mediated by PMNs has also been resolved in antibody-neutropenic models. For instance, it has been demonstrated that in the absence of PMNs, mice infected by the intranasal route with S. pneumoniae serotype 8 survived for longer periods than uninfected mice  indicating that local proinflammatory responses induced by PMNs may be also lethal. However, due to the fact that PMN antibody depletion cannot be maintained beyond one week, the assessment of the role of PMNs in adaptive immunity in chronic bacterial infections such as brucellosis or tuberculosis has been precluded. Therefore, Genista, a viable neutropenic mouse emerged as an important model to study the influence of PMNs in the development of adaptive immunity .