The role of programmed cell death in the acute respiratory distress syndrome
File(s)
Author(s)
Handslip, Rhodri
Type
Thesis
Abstract
The acute respiratory distress syndrome (ARDS) remains associated with a high mortality rate. Despite advances in critical care and dedicated research, treatment options are limited. Pulmonary necrosis is a key histological feature of diffuse alveolar damage which defines ARDS. Necroptosis is the archetypal form of programmed necrosis which is canonically activated by tumour necrosis factor (TNF)-α.
TNF signalling is also highly implicated in the pathobiology of ARDS. Necroptosis is executed through receptor interacting serine/threonine-protein kinase (RIPK)-1 and 3 driven phosphorylation of mixed lineage kinase like domain (MLKL). Phosphorylated MLKL (pMLKL) oligomerises and forms plasma membrane pores, leading to necroptotic cell lysis consequently releasing pro-inflammatory cytokines and DAMPs extracellularly.
Our objectives were: firstly, to develop an in vitro pulmonary endothelial model of different forms of TNFR-1 mediated programmed cell death, principally to examine the necroinflammatory response. Secondly, to characterise the necroinflammatory response in clinical ARDS to investigate whether necroptosis is activated and may contribute to the pathobiology of ARDS.
The TNFR-1 cell death model stimulated TNFR-1 complex I, II and successfully activated necroptosis. Necroptosis caused upregulation of cell death, endothelial activation, and cytokine production, which were attenuated by necroptosis inhibitors, confirming necroptosis activation. Intracellular RIPK3-pMLKL complexes were formed and released following necroptosis activation. Paradoxically, RIPK3-pMLKL complexes were also extruded following MLKL inhibition, in the absence of cell death, potentially revealing a protective mechanism.
Secondly, raised RIPK3 and necrosis marker Cytokeratin-18 were detectable in ARDS blood and BAL fluid samples. RIPK3 levels were persistently increased in the non-resolving ‘hyperinflammatory’ ARDS subphenotype by comparison to the resolving ‘hypoinflammatory’ subphenotype. RIPK3 was bound to pMLKL in complexes in the circulation of general, Influenza and COVID ARDS patients, implicating necroptosis activation in multiple aetiologies of ARDS. The data potentially provides insight into the role of necroptosis in the pathobiology of ARDS.
TNF signalling is also highly implicated in the pathobiology of ARDS. Necroptosis is executed through receptor interacting serine/threonine-protein kinase (RIPK)-1 and 3 driven phosphorylation of mixed lineage kinase like domain (MLKL). Phosphorylated MLKL (pMLKL) oligomerises and forms plasma membrane pores, leading to necroptotic cell lysis consequently releasing pro-inflammatory cytokines and DAMPs extracellularly.
Our objectives were: firstly, to develop an in vitro pulmonary endothelial model of different forms of TNFR-1 mediated programmed cell death, principally to examine the necroinflammatory response. Secondly, to characterise the necroinflammatory response in clinical ARDS to investigate whether necroptosis is activated and may contribute to the pathobiology of ARDS.
The TNFR-1 cell death model stimulated TNFR-1 complex I, II and successfully activated necroptosis. Necroptosis caused upregulation of cell death, endothelial activation, and cytokine production, which were attenuated by necroptosis inhibitors, confirming necroptosis activation. Intracellular RIPK3-pMLKL complexes were formed and released following necroptosis activation. Paradoxically, RIPK3-pMLKL complexes were also extruded following MLKL inhibition, in the absence of cell death, potentially revealing a protective mechanism.
Secondly, raised RIPK3 and necrosis marker Cytokeratin-18 were detectable in ARDS blood and BAL fluid samples. RIPK3 levels were persistently increased in the non-resolving ‘hyperinflammatory’ ARDS subphenotype by comparison to the resolving ‘hypoinflammatory’ subphenotype. RIPK3 was bound to pMLKL in complexes in the circulation of general, Influenza and COVID ARDS patients, implicating necroptosis activation in multiple aetiologies of ARDS. The data potentially provides insight into the role of necroptosis in the pathobiology of ARDS.
Version
Open Access
Date Issued
2022-11
Date Awarded
2023-06
Copyright Statement
Creative Commons Attribution NonCommercial Licence
License URL
Advisor
Patel, Brijesh
Takata, Masao
Sponsor
European Society of Intensive Care Medicine
Wellington Hospital
Imperial College London
Publisher Department
Department of Surgery & Cancer
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)