Canine Neutrophil Extracellular Traps Release Induced by the Apicomplexan Parasite Neospora caninum In Vitro.

Zhengkai Wei,Xuexiu He,Zhengtao Yang,Carlos Hermosilla,Xiaocen Wang,Anja Taubert,Pengtao Gong,Jianhua Li,Xichen Zhang

doi:10.3389/fimmu.2016.00436

Zhengkai Wei, Xuexiu He + Show 7 more

Open Access

https://doi.org/10.3389/fimmu.2016.00436

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Abstract

Neosporosis is considered as one of the main causes of abortion and severe economic losses in dairy industry. The Canis genus serving as one of the confirmed definitive hosts of the apicomplexan parasite Neospora caninum (N. caninum) plays a critical role in its life cycle. However, the effects of N. caninum on its definitive hosts of neutrophils extracellular traps (NETs) formation remain unclear. In the present study, N. caninum tachyzoite-induced canine NETs formation was observed by scanning electron microscopy (SEM). Visualization of DNA decorated with H3, neutrophil elastase (NE), and myeloperoxidase (MPO) within N. caninum tachyzoite-induced NETs were examined using fluorescence confocal microscopy analyses. Furthermore, the formation of canine NETs was quantified using Sytox Green staining, and the LDH levels in supernatants were examined by an LDH Cytotoxicity Assay® kit. The results clearly showed that NETs-like structures were induced by N. caninum tachyzoites, and the major components within these structures induced by N. caninum tachyzoite were further confirmed by fluorescence confocal microscopy visualization. These results suggest that N. caninum tachyzoites strongly induced NETs formation in canine polymorphonuclear neutrophils (PMN). In functional inhibition assays, the blockings of NADPH oxidase, NE, MPO, SOCE, ERK 1/2, and p38 MAPK signaling pathways significantly inhibited N. caninum tachyzoite-induced NETs formation. To our knowledge, this study is the first to report the formation of NETs in canine PMN against N. caninum infection.

Highlights

Neosporosis is caused by the apicomplexan protozoa Neospora caninum and is considered as one of the main diseases causing abortion, reproduction disorders, and severe economic losses in dairy industry worldwide [1, 2]
N. caninum tachyzoite-induced neutrophils extracellular traps (NETs) in canine polymorphonuclear neutrophils (PMN) were demonstrated by fluorescence confocal microscopy analyses (Figures 2B,E,H)
Fluorescence confocal microscopy analyses revealed that NETs formation can be induced by N. caninum tachyzoites, and the nature of these structures was mainly composed by DNA (Figures 2B,E,H)

Summary

Introduction

Neosporosis is caused by the apicomplexan protozoa Neospora caninum and is considered as one of the main diseases causing abortion, reproduction disorders, and severe economic losses in dairy industry worldwide [1, 2]. N. caninum is an apicomplexan parasite closely related to Toxoplasma gondii, which infects a wide intermediate host range [3, 4]. Its confirmed definitive hosts of N. caninum is the genus Canis, including domestic dogs (Canis familiaris) [5], dingoes (Canis lupus dingo) [6], and gray wolves (Canis lupus) [7]. Neosporosis in canines principally results in a neuromuscular disease, i.e., polyradiculoneuritis-myositis, but a variety of less common lesions due to focal necrosis can occurs in other organs [8, 9].

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