Signal Inhibitory Receptor on Leukocytes-1 Limits the Formation of Neutrophil Extracellular Traps, but Preserves Intracellular Bacterial Killing

Abstract

In response to microbial invasion, neutrophils release neutrophil extracellular traps (NETs) to trap and kill extracellular microbes. Alternatively, NET formation can result in tissue damage in inflammatory conditions and may perpetuate autoimmune disease. Intervention strategies that are aimed at modifying pathogenic NET formation should ideally preserve other neutrophil antimicrobial functions. We now show that signal inhibitory receptor on leukocytes-1 (SIRL-1) attenuates NET release by human neutrophils in response to distinct triggers, including opsonized Staphylococcus aureus and inflammatory danger signals. NET release has different kinetics depending on the stimulus, and rapid NET formation is independent of NADPH oxidase activity. In line with this, we show that NET release and reactive oxygen species production upon challenge with opsonized S. aureus require different signaling events. Importantly, engagement of SIRL-1 does not affect bacterially induced production of reactive oxygen species, and intracellular bacterial killing by neutrophils remains intact. Thus, our studies define SIRL-1 as an intervention point of benefit to suppress NET formation in disease while preserving intracellular antimicrobial defense.

Differential Signalling and Kinetics of Neutrophil Extracellular Trap Release Revealed by Quantitative Live Imaging

“…In addition, neutrophils sense microbe size and release NETs in response to large pathogens 42 , arguing for an active process that occurs directly after neutrophil contact with the microbe. In contrast with fluorescence plate reader data from our previous study 19 , we here show that only MSU triggers rapid NET release while NET formation in response to opsonized S. aureus , PMA, and ICs occurs at a later phase. Others have also shown that PMA 18 and ICs 22 induce late NET release.…”

“…Consistent with this study, we previously have shown that, when using a fluorescence plate reader assay, Sytox Green fluorescence appears within 30 min when neutrophils were exposed to viable S. aureus 16, 19 . We concluded at the time that neutrophils are able to rapidly release NETs in response to these stimuli.…”

“…This demonstrates that some stimuli require NADPH-induced ROS to release NETs. On the contrary, we and others showed that DPI does not affect NET release when neutrophils are stimulated with MSU 19 , ICs 48 , uric acid 49 , Leishmania 21 , ionomycin 50 and opsonized S. aureus 16, 19 . In addition, it was shown that CGD neutrophils are still able to release NETs when exposed to uric acid 49 , which suggests the presence of a NADPH-oxidase independent NET signalling pathway.…”

“…Rapid NET release has been shown in response to Staphylococcus aureus ( S. aureus ) 16 , Fc receptor (FcR) activation 17, 18 , MSU 19 , Candida albicans ( C. albicans ) hyphea 20 , and Leishmania 21 , while PMA 18 , ICs 22 and C. albicans yeast 20 trigger NET formation at later time points. Many of these studies were performed with fluorescence plate reader experiments using DNA binding dye’s (i.e.…”

In vitro induction of NETosis: Comprehensive live imaging comparison and systematic review

“…aureus has consistently been described to be a potent inducer of NETosis [12,14,34,40,71,75,104,131–133]. In literature search a variety of other bacteria were reported, that also induced NETosis, although most species are weak NETosis inducers compared to S .…”