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Visualizing Macrophage Extracellular Traps Using Confocal Microscopy
Published on: October 19, 2017
Pentatrichomonas hominis induces extracellular traps formation of macrophages via the TLR2/NADPH/PAD4 pathway
Qi-An Zhai1, Xi-Chen Zhang1, Hong-Bo Zhang1
1State Key Laboratory for Diagnosis and Treatment of Severe Zoonotic Infectious Diseases, Key Laboratory for Zoonosis Research of the Ministry of Education, Institute of Zoonosis, and College of Veterinary Medicine, Jilin University, Changchun, 130062, People's Republic of China.
Background:
Pentatrichomonas hominis (P. hominis) is a newly identified pathogenic zoonotic protozoan belonging to the Trichomonadidae family. P. hominis mainly parasitizes the cecum and colon of humans and other mammals, and it can cause diarrhea. Recently, macrophage extracellular traps (METs) have been shown to play an important role in resistance to parasitic infections. However, it remains unclear whether the release of METs by macrophages contributes to P. hominis resistance, and the underlying mechanism of this association has yet to be elucidated.
Methods:
Scanning electron microscopy (SEM) and immunofluorescence staining were used to determine whether P. hominis induced the formation of METs in mouse peritoneal macrophages to capture and immobilize the parasite as well as the components of METs, including the DNA backbone, myeloperoxidase (MPO), and histone H3. Reactive oxygen species (ROS) and signaling pathway inhibitor assays revealed that the mechanism of P. hominis-induced MET formation was dependent on nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) oxidase activation, store-operated calcium entry (SOCE),, and peptidyl arginine deiminase 4 (PAD4) activation. The toll-like receptors 2 (TLR2), extracellular regulated protein kinase 1/2 (ERK1/2) and p38 MAPK signaling pathway were also involved in this process. Trypan blue staining revealed a gradual decrease in the survival rate of P. hominis as the coculture time increased. Trypan blue staining also revealed an increase in the proportion of macrophages.
Results:
The results of this study indicate that P. hominis can induce the release of METs via the TLR2/NADPH/PAD4 pathways and that METs have a trapping and killing effect on P. hominis.
Conclusions:
This was the first study to reveal that PAD4 and TLR2 were found to be involved in the development of parasite-induced METs, thus providing guidance for further research on the mechanisms of host innate immunity against parasitic infection.
Insights
Pentatrichomonas hominis infection triggers macrophage extracellular traps (METs) via PAD4 and TLR2 pathways. These METs effectively trap and kill the parasite, offering insights into host innate immunity against protozoan infections.
Area of Science:
- Immunology
- Parasitology
- Cell Biology
Background:
- Pentatrichomonas hominis is a pathogenic zoonotic protozoan causing diarrhea in mammals.
- Macrophage extracellular traps (METs) are crucial in combating parasitic infections.
- The role of METs in P. hominis resistance and the underlying mechanisms were previously unknown.
Purpose of the Study:
- To investigate whether macrophages release METs in response to P. hominis.
- To elucidate the molecular mechanisms driving P. hominis-induced MET formation.
- To assess the efficacy of METs in trapping and killing P. hominis.
Main Methods:
- Scanning electron microscopy and immunofluorescence staining were used to visualize MET formation and components.
- Inhibitor assays assessed the roles of NADPH oxidase, SOCE, PAD4, TLR2, ERK1/2, and p38 MAPK signaling.
- Trypan blue staining evaluated parasite viability and macrophage survival.
Main Results:
- P. hominis induced MET release in mouse peritoneal macrophages.
- MET formation was dependent on the TLR2/NADPH/PAD4 signaling pathway.
- METs demonstrated a significant trapping and killing effect on P. hominis, reducing parasite survival.
Conclusions:
- This study demonstrates that P. hominis induces METs through PAD4 and TLR2 activation.
- METs play a critical role in the innate immune response against P. hominis.
- Findings provide a foundation for understanding host immunity against parasitic infections.
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