Date Published: March 10, 2015
Publisher: Public Library of Science
Author(s): Sarah A. C. Falcão, Tiffany Weinkopff, Benjamin P. Hurrell, Fabiana S. Celes, Rebecca P. Curvelo, Deboraci B. Prates, Aldina Barral, Valeria M. Borges, Fabienne Tacchini-Cottier, Camila I. de Oliveira, Shaden Kamhawi. http://doi.org/10.1371/journal.pntd.0003601
Abstract: BackgroundNeutrophils are the first line of defense against invading pathogens and are rapidly recruited to the sites of Leishmania inoculation. During Leishmania braziliensis infection, depletion of inflammatory cells significantly increases the parasite load whereas co-inoculation of neutrophils plus L. braziliensis had an opposite effect. Moreover, the co-culture of infected macrophages and neutrophils also induced parasite killing leading us to ask how neutrophils alone respond to an L. braziliensis exposure. Herein we focused on understanding the interaction between neutrophils and L. braziliensis, exploring cell activation and apoptotic fate.Methods and FindingsInoculation of serum-opsonized L. braziliensis promastigotes in mice induced neutrophil accumulation in vivo, peaking at 24 h. In vitro, exposure of thyoglycollate-elicited inflammatory or bone marrow neutrophils to L. braziliensis modulated the expression of surface molecules such as CD18 and CD62L, and induced the oxidative burst. Using mCherry-expressing L. braziliensis, we determined that such effects were mainly observed in infected and not in bystander cells. Neutrophil activation following contact with L. braziliensis was also confirmed by the release of TNF-α and neutrophil elastase. Lastly, neutrophils infected with L. braziliensis but not with L. major displayed markers of early apoptosis.ConclusionsWe show that L. braziliensis induces neutrophil recruitment in vivo and that neutrophils exposed to the parasite in vitro respond through activation and release of inflammatory mediators. This outcome may impact on parasite elimination, particularly at the early stages of infection.
Partial Text: Neutrophils are essential components of the early inflammatory response, acting as the first line of defense against invading pathogens (rev. in ). Neutrophil recruitment to the infection site occurs in response to various stimuli and is followed by cell rolling and adhesion to the vasculature, processes mediated by interactions between selectins and integrins . Pathogen phagocytosis subsequently elicits the production of superoxide, which is quickly dismutated into hydrogen peroxide and other secondary Reactive Oxygen Species (ROS), which are highly toxic to the invading pathogen . Phagocytosis stimulates the secretion of additional antimicrobial molecules such as neutrophil elastase, into the phagosome further contributing with pathogen killing . Resolution of inflammation requires efficient removal of apoptotic neutrophils by professional phagocytes such as resident macrophages . Phagocytosis of apoptotic neutrophils prevents the release of potentially toxic molecules and, in parallel, regulates the inflammatory response .
Numerous studies have demonstrated that neutrophils play a crucial role in immunity against bacterial, fungal  and intracellular pathogens . Earlier on, we demonstrated that L. braziliensis inoculation into the ear dermis of BALB/c mice leads to the development of a cutaneous ulcer, which heals spontaneously after ten weeks of infection . Additionally, co-inoculation of L. braziliensis and neutrophils decreased lesion size whereas depletion of neutrophils and monocytes had an opposing effect, significantly increasing parasite load and lesion size . Given that neutrophils are among the first cells to encounter the parasite at the site of the sand fly bite  and, thus, will readily encounter Leishmania parasites, the purpose of the current study was to investigate how neutrophils respond to L. braziliensis exposure, evaluating neutrophil activation and downstream events such as apoptosis.