Characterisation of IL-33/ST2 signaling and crosstalk in mast cells and their modulation by ES-62

Ball, Dimity (2013) Characterisation of IL-33/ST2 signaling and crosstalk in mast cells and their modulation by ES-62. PhD thesis, University of Glasgow.

Abstract

In addition to their role in fighting infection, mast cells have long been implicated in the pathogenesis of allergic and autoimmune inflammatory diseases and cancers. Increasingly, however, there is recognition that these cells may also play a part in protecting against development of pathologies. Indeed, there is increasing evidence that mast cells comprise heterogeneous phenotypes that exhibit functional plasticity to allow them to play both pro- and anti-inflammatory roles during an immune response. This plasticity appears to reflect that immature mast cells are tailored by their particular microenvironment not only to trigger protective inflammatory responses but also to limit pathology by resolving inflammation and promoting wound healing and tissue repair. Mast cells can be activated by a range of stimuli including (pathogen-derived) antigen/allergen-mediated crosslinking of antibody-bound to Fc receptors, most notably FcεRI, pathogen-associated molecular patterns (PAMP) such as bacterial lipopolysaccharide (LPS) acting on TLR4, inflammatory cytokines such as IL-33 (via the IL-1R/TLR-like receptor ST2) and tissue-derived signals such as SCF (via cKIT). During infection these stimuli provoke a response optimised for pathogen clearance however in autoimmune or allergic disease such responses can initiate and exacerbate host pathology. Thus the challenge for therapeutic targeting of mast cells in inflammatory or malignant disease is to limit mast cells with pathogenic phenotypes whilst preserving those contributing to protective homeostatic and anti-pathogen responses.

Thus, as a first step, it was a core aim of these studies to generate in vitro mast cell models representing the phenotypic and maturational heterogeneity of mast cells in vivo, as these are difficult to isolate and purify due to their limited numbers in tissue. Distinct murine mast cell phenotypes, namely mature serosal peritoneal-derived mast cells (PDMC), connective tissue-like mast cells (CTMC) and mucosal-like mast cells (BMMC), the latter two subtypes both derived from bone marrow progenitors, were found to differentially respond, in terms of cytokine production and degranulation, to important immunoregulatory receptors in health and disease, namely FcεRI and TLR4. Consistent with their distinct functional responses, these mast cell subtypes were also found to display differential calcium signalling profiles in response to FcεRI and TLR4 signalling, further highlighting the importance in investigating phenotypically relevant and microenvironment-specific (serosal versus mucosal) mast cells in drug discovery programmes.

Recently there has been great interest in IL-33, a pro-inflammatory cytokine increasingly recognised as playing an important role in a variety of mast cell responses associated with allergic inflammatory disorders and tumour pathogenesis. Consistent with this, whilst IL-33 can stimulate mast cell cytokine production, but not degranulation, via the IL-1R/TLR-like receptor ST2, responses to this cytokine are amplified following IgE sensitization and/or exposure to SCF or serum. Such augmented responses reflect increased calcium mobilization, PLD, SphK, ERK and NF-κB signalling and mTOR activation and can be suppressed by existing therapeutics targeting the costimulatory signal, for example, Imatinib or Dasatinib for SCF/cKIT and potentially Omalizumab for IgE/FcεR1. Moreover, IL-33/ST2 signalling can modulate mast cell responses resulting from antigen-mediated crosslinking of FcεRI and LPS-TLR4 signalling. Indeed, ST2 signalling can differentially modulate LPS/TLR4 responses depending on the presence (enhances) or absence (inhibits) of IL-33, as in the latter case ST2 acts to limit LPS cytokine production, potentially by sequestering MyD88. This receptor crosstalk is likely to occur under pathological conditions, thus targeting of such cooperative signalling may allow the downregulation of hyper-inflammatory responses, whilst leaving protective and homeostatic mast cell responses intact.

ES-62 is an immunomodulator produced by filarial nematodes to dampen immune responses in order to promote parasite survival and prevent tissue damage without immunocompromising the host to infection. As a serendipitous side effect of its anti-inflammatory actions, ES-62 exhibits therapeutic potential in both allergic and autoimmune inflammatory disease and thus to further explore the potential for safe, targeted downregulation of pathogenic mast cell responses, the parasite product was exploited in order to identify signals regulating mast cell activation. ES-62 was found to be able to induce hypo-responsiveness of all three mast cell phenotypes in terms of degranulation and cytokine production in response to stimulation of FcεR1-, TLR4- or IL-33/ST2, either alone or in combination. ES-62 mediated these effects, at least in part, by mechanisms involving downregulation of PKCα (and in BMMC, MyD88) expression and calcium mobilisation and, in PDMC, potentially by subverting the negative feedback interactions of ST2 on TLR4. The precise mechanism of modulation varies both with receptor usage and mast cell phenotype as ES-62 exhibits differential effects in PDMC and BMMC. Nevertheless, collectively these data support the role of calcium-, PKCα- and MyD88- as key regulatory intersection sites in the functional crosstalk amongst these important immunoregulatory receptors and importantly, suggest they are potential targets for therapeutic intervention in pathogenic mast cell responses.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Keywords: Mast cell IL-33 ST2 ES-62 cell signaling
• Subjects: Q Science > QH Natural history > QH301 Biology; Q Science > QH Natural history > QH345 Biochemistry
• Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Infection & Immunity
• Supervisor's Name: Harnett, Professor Margaret
• Date of Award: 2013
• Depositing User: Miss Dimity Ball
• Unique ID: glathesis:2013-4750
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 08 Jan 2014 15:10
• Last Modified: 09 Jan 2014 11:48
• URI: https://theses.gla.ac.uk/id/eprint/4750