Tissue damage initiates the release of a complex, interacting collection of chemical signals. The coordinated function of these signals gives rise to an inflammatory event, whereby circulating immune cells are recruited to clear pathogens. Invertebrate models of tissue damage have revealed a key role for damage-associated signals, specifically hydrogen peroxide (H2O2), in attracting immune cells to sites of tissue damage. The Src family kinase (SFK), Lyn, is oxidised by H2O2 in zebrafish wound models, and subsequent activation of Lyn triggers directed cell motility. In mammalian systems, H2O2 is an important second messenger, however, its role as a damage signal and its association with SFK signalling remains unclear.
The aims of this thesis are to investigate how immune cell function and migration in response to damage-associated signals transfers into other models. To address this, we have used an in vivo Drosophila melanogaster embryonic wounding model and in vitro assays of human innate cell function and migration.
The migration of Drosophila hemocytes in response to a wound was impaired in embryos lacking functional Src42A or Shark kinase. However, hemocyte motility was impaired in embryos bathed in exogenous H2O2. In vitro, H2O2 inhibited human innate cell motility, chemotaxis, actin reorganisation and phagocytosis, but activated intracellular signalling pathways and did not affect receptor expression or cell viability. Exogenous ATP activated chemokinesis and rapid actin reorganisation. The in vitro effects of immune-related ligands were inhibited by pharmacological inhibition of SFK, Syk, and PI3K signalling. In particular, inhibition of class IA PI3K isoforms p110β and p110δ, but not p110α, disrupted monocyte MCP-1-mediated actin reorganisation and spreading. SFKs are required for Drosophila immune cell migration to a wound and for human innate cell migration to chemoattractants. While endogenous ROS production is important for immune cell function, exogenous H2O2 may negatively modulate downstream mediators of chemokine signalling. H2O2 and ATP are distinct in their abilities to activate immune cells and initiate chemokinesis. Intracellular kinases regulate basal and chemoattractant-mediated motility and are therefore attractive targets for therapeutic management of inflammatory disease.
|Date of Award
|20 Jan 2015
|Stephen Ward (Supervisor)