Jenkins, Stephen

Forbes, Stuart

Lynch, Ruairi

2019-10-14T09:54:31Z

2019-10-14T09:54:31Z

2019-11-25

BACKGROUND: Acute liver failure has significant mortality for which the only reliable treatment is transplantation. Murine models of acute liver injury are characterised by dramatic changes in the hepatic macrophage compartment, with an initial accumulation of pro-inflammatory Ly6Chi monocytes and loss of Kupffer cells, followed by the dominance of Ly6Clo macrophages that promote liver repair. The canonical type 2 immune cytokine Interleukin 4 (IL- 4) prevents damaging inflammation and accelerates tissue repair following helminth infection and physical wounding, partly through effects on myeloid cells including macrophages. Furthermore, exogenous IL-4 administered prior to injury with the hepatotoxin carbon tetrachloride (CCl4) has been shown to be hepatoprotective by directly promoting hepatocyte proliferation. For this reason, in my thesis I aimed to assess the effects of therapeutic IL-4 administered following acute liver injury and examine the underlying pathways with an emphasis on the effects of IL-4 on hepatic myeloid cells. METHODS: Male C57Bl/6 mice were given CCl4 intraperitoneally to induce an acute liver injury. IL-4 in the form of IL-4 complex (IL-4c) was administered subcutaneously to manipulate the repair and regeneration process. To investigate the role of IL-4Ra signalling in bone marrow derived cells, whole-body and tissue-protected chimeras were generated with wild type or IL-4Ra– /– donor bone marrow. An in vitro culture system was developed to validate the observed effect of IL-4 on Ly6Chi monocytes. Models of acetaminophen (APAP) induced acute liver injury as well as dextran sulfate sodium (DSS) colitis were used to investigate the translational potential of IL-4c therapy. The effect of IL-4c in experiments was assessed through immunohistochemical analysis of tissue sections, analysis of serum biochemistry and flow cytometric analysis of leukocyte populations. RESULTS: Therapeutic administration of IL-4c following CCl4 induced liver injury reduced markers of hepatic injury (ALT and necrotic area) and enhanced hepatic regeneration as measured by hepatocyte proliferation. This was paralleled by profound alterations to the monocyte/macrophage pool, with increases in the number of hepatic Ly6Clo macrophages but also a large reduction in the number of hepatic Ly6Chi monocytes. Using chimeras, I have shown that cell-intrinsic, IL-4Ra-dependent proliferation contributes to the observed accumulation of Ly6Clo macrophages. Importantly, the loss of hepatic Ly6Chi monocytes following administration of IL-4c was associated with a systemic IL-4Ra dependent loss of Ly6Chi monocytes. Analysis of tissues and blood revealed an increased detection of dead and apoptotic monocytes and culture of monocytes with IL-4 in vitro directly induced the monocyte death, a feature that could be overcome by high levels of the macrophage colony-stimulating factor (CSF-1). Hence, the systemic reduction in Ly6Chi monocytes in response to therapeutic delivery of IL-4c is due to IL-4Ra-dependent death of circulating monocytes rather than decreased output from the bone marrow. Experiments using APAP and DSS suggested that the beneficial effect of IL-4c may be dependent on the timing of delivery following injury. CONCLUSION: I have demonstrated a therapeutic effect for IL-4c following CCl4 mediated acute liver injury and I have also identified a novel role for IL-4 in promoting the death of Ly6Chi monocytes. This effect of IL-4 may offer insights and potential therapeutic targets in other inflammatory pathologies characterised by monocytosis. However, preliminary assessments in the APAP and DSS models would suggest that this may need careful optimisation.

http://hdl.handle.net/1842/36218

en

The University of Edinburgh

Lynch RW, Hawley CA, Pellicoro A, Bain CC, Iredale JP, Jenkins SJ. An efficient method to isolate Kupffer cells eliminating endothelial cell contamination and selective bias. J LeukocBiol. 2018;104:579–586. https://doi.org/ 10.1002/JLB.1TA0517-169R

liver injury

chemical messenger treatment

pro-inflammatory Ly6Chi monocytes

therapeutic IL-4

hepatic myeloid cells

mouse model

Role of exogenous Interleukin 4 in modulating the mononuclear phagocyte response to acute liver injury.

Thesis or Dissertation

Doctoral

PhD Doctor of Philosophy