Early and late events in NET formation

Neutrophils are immune cells that are central to host defense against infection and injury. Among other antimicrobial responses, neutrophils are known to produce NETs (for neutrophil extracellular traps) – networks of extruded decondensed chromatin adorned with antimicrobial molecules, which primarily serve to immobilize and inactivate pathogens (bacteria, viruses, yeasts, and fungi). In addition, it has been widely reported that NETs play a role in the progression of various pathologies such as autoimmune, inflammatory and cardiovascular diseases, as well as cancer. Hence the importance of understanding the molecular mechanisms underlying NET generation. In humans, this response spans over about four hours ex vivo, with distinct early (≤ 30 min) and late (90-180 min) signaling events. However, our understanding of the various downstream cellular processes remains incomplete. In this work, we report that histone citrullination (as determined by immunoblot) is an immediate-early event in NET generation (<5 min), that is under the control of the p38, MEK, and TAK1 signaling pathways, which govern the early phase of NET generation. Conversely, chromatin decondensation is a late event (occurring at 3-3.5 h as assessed by confocal microscopy) that is controlled by all signaling pathways known to affect NET formation (p38, MEK, TAK1, Syk, and PI3K), whether they act early or belatedly. Furthermore, we demonstrate (using epifluorescence microscopy and a pharmacological antagonist) that endogenous mediators released by NETing neutrophils feed back on the cells by engaging a common receptor, RAGE, thereby driving NET induction. Other endogenous RAGE ligands similarly participate in the conditioning of adherent neutrophils to quickly form NETs (i.e. within 1h, instead of 4h). Finally, we show that regarding the late-acting PI3K signaling pathway, different PI3K isoforms are mobilized depending on the class of physiological stimuli (e.g. inflammatory cytokines and chemoattractants) employed to trigger NET generation. Collectively, our results shed light on various cellular processes driving the NET response. In particular, the key role of endogenous mediators is a novel finding that offers new therapeutic opportunities for the various pathological conditions in which NETs are involved.

Name thesis:degree_name

M. Sc.

Level thesis:degree_level

Maîtrise

Discipline thesis:degree_discipline

Immunologie

Grantor dc:publisher

Université de Sherbrooke

Year dc:date.issued

2022

Author dc:creator

• De Carvalho Oliveira, Vanessa

Advisor dc:contributor.advisor

• Amrani, Abdelaziz

• Neutrophil extracellular traps
• Endogenous mediators
• RAGE
• PI3K isoforms
• NETs
• Médiateurs endogènes
• Isoformes de PI3K

Language dc:language.iso

en

Handle dc:identifier.uri

http://hdl.handle.net/11143/21759

OAI identifier oai:identifier

oai:usherbrooke.scholaris.ca:11143/21759