Molecular regulation of neutrophil responses

  • Drinalda Cela

Student thesis: Doctoral ThesisDoctor of Philosophy (PhD)

Abstract

Neutrophils are the most abundant immune cells in circulation, and they are crucial in antimicrobial defence. They rapidly migrate towards wounds or infection sites and limit microbial proliferation by phagocytosis, reactive oxygen species (ROS) production, degranulation and release of neutrophil extracellular traps (NETs). Dysregulated neutrophil responses, however, can be destructive and contribute to pathology in many inflammatory and autoimmune diseases. Mechanisms that fine-tune neutrophil responses are essential, yet our understanding of these mechanisms remains limited, as does our ability to modulate them in diseases. The work presented herein describes two molecular mechanisms orchestrated by CHK1 and PAD4 protein.
First, we identified that CHK1 protein regulates the formation of NETs. By using chemical inhibitors and inducible shRNA approach, we showed that CHK1 regulates key steps during the process of NET formation, including ROS production, translocation of NE from the cytoplasm to the nucleus and cleavage of Histone H3. In addition, by performing phosphoproteomic analysis of NET-stimulated neutrophils and PLB-985 cells, we identified the involvement of multiple proteins, including regulators of DNA damage, cell cycle and microtubule dynamics.
Secondly, we characterised the function of PAD4, an enzyme known to regulate neutrophil responses. To study the function of PAD4 during inflammation, we used the P. chabaudi mouse malaria model as well as in vitro culture of human macrophages and neutrophils. Using PAD4 deficient mice and chemical inhibitors, we demonstrate that PAD4 can regulate macrophage CXCL1 production and thus control neutrophil trafficking into the diseased organ. By affecting neutrophil migration, PAD4 can alter the severity of malaria in P. chabaudi infected mice.
Our study shed light on molecular regulation of neutrophil responses and how we can fine-tune neutrophil responses and potentially alter the outcome of inflammatory disease.
Date of Award27 Sept 2022
Original languageEnglish
Awarding Institution
  • University of Bristol
SupervisorBorko Amulic (Supervisor) & Christoph Wuelfing (Supervisor)

Keywords

  • neutrophils
  • NETs
  • Malaria
  • CHK1
  • PAD4

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