Abstract

Emerging evidence indicates that innate host response contributes to the therapeutic effect of antimicrobial medications. Recent studies have shown that Leishmania parasites derived by in vitro selection for resistance to pentavalent antimony (SbV) as meglumine antimoniate (MA) modulate the activation of neutrophils. However, whether modulation of neutrophil activation extends to natural resistance to this antileishmanial drug has not been established. We have evaluated the influence of clinical strains of L. (V.) panamensis having intrinsic tolerance/resistance to SbV, on the inflammatory response of neutrophils during ex vivo exposure to MA. Accordingly, neutrophils obtained from healthy donors were infected with clinical strains that are sensitive (n = 10) or intrinsically tolerant/resistant to SbV (n = 10) and exposed to a concentration approximating the maximal plasma concentration (Cmax) of SbV (32 µg/ml). The activation profile of neutrophils was evaluated as the expression of the surface membrane markers CD66b, CD18, and CD62L by flow cytometry, measurement of reactive oxygen species (ROS) by luminometry, and NET formation using Picogreen to measure dsDNA release and quantification of NETs by confocal microscopy. These parameters of activation were analyzed in relation with parasite susceptibility to SbV and exposure to MA. Here, we show that clinical strains presenting intrinsic tolerance/resistance to SbV induced significantly lower ROS production compared to drug-sensitive clinical strains, both in the presence and in the absence of MA. Likewise, analyses of surface membrane activation markers revealed significantly higher expression of CD62L on cells infected with intrinsically SbV tolerant/resistant L. (V.) panamensis than cells infected with drug-sensitive strains. Expression of other activation markers (CD18 and CD66b) and NET formation were similar for neutrophils infected with SbV sensitive and tolerant clinical strains under the conditions evaluated. Exposure to MA broadly impacted the activation of neutrophils, diminishing NET formation and the expression of CD62L, while augmenting ROS production and CD66b expression, independently of the parasite susceptibility phenotype. These results demonstrated that activation of human neutrophils ex vivo is differentially modulated by infection with clinical strains of L. (V.) panamensis having intrinsic tolerance/resistance to SbV compared to sensitive strains, and by exposure to antimonial drug.

Highlights

  • Cutaneous leishmaniasis (CL) is a neglected tropical disease of emerging or reemerging public health importance in 98 countries, with more than a million new cases reported annually (Alvar et al, 2012; Pace, 2014)

  • The modulation of radical oxygen species (ROS) production by meglumine antimoniate (MA) exposure of neutrophils infected with L. (V.) panamensis was confirmed by the dose-dependent response to four clinically relevant concentrations of SbV (Figure 2B)

  • The results of this study show that infection of neutrophils with phenotypically/genotypically distinguishable clinical strains of L. (V.) panamensis that are intrinsically sensitive or resistant to SbV differentially modulates neutrophil activation

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Summary

Introduction

Cutaneous leishmaniasis (CL) is a neglected tropical disease of emerging or reemerging public health importance in 98 countries, with more than a million new cases reported annually (Alvar et al, 2012; Pace, 2014). Growing evidence indicates that neutrophils, the first host cells recruited to the site of infection, may play a role in the pathogenesis of leishmaniasis depending on the infecting Leishmania spp (Afonso et al, 2008; Carlsen et al, 2013; Hurrell et al, 2015; Charmoy et al, 2016; Hurrell et al, 2016; Passelli et al, 2021). Despite their important role in cutaneous leishmaniasis, the involvement of neutrophils in the response to antileishmanial drugs has not been examined

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