The increasing awareness that infection by helminths can prevent the development of allergy and autoimmunity partially due to the induction of a regulatory immune response has led to the investigation of parasite-derived molecules and their interactions. This study sought to explore the unique nematode specific fatty acid and retinol binding (FAR) proteins and their potentially important retinol binding ability. Therefore, the FAR protein of the immunoregulatory murine gastrointestinal nematode, Heligmosomoides bakeri was chosen. Recombinant Hb-FAR-l (rHb-FAR- 1) was successfully expressed in the yeast Pichia pastoris, and its retinol and fatty acid binding ability was confirmed. Despite the rare activation of toll-like receptors (TLRs) by parasite-derived molecules, this protein was found to trigger TLR2 on human embryonic kidney cells 293. Overall, rHb-FAR-l failed to induce the maturation of bone marrow derived dendritic cells (DCs) or macrophages (M<l>s), although DC-derived IL-6 was increased. In contrast to many helminth antigens, rHb-FAR-l treated DCs or M<l>s were not found to be hyporesponsive to simultaneous TLR activation, but DCs displayed elevated cytokine levels (IL-6, TNF-a) and increased expression of surface activation markers (MHC-II, CD86) in response to two TLR2 ligands, Pam3CSK4 or FSL-l, in comparison to exposure to the TLR2 agonists alone. However, rHb-FAR-l failed to modify the response of DCs to LPS (TLR4), zymosan (TLR2) or ODN1826 (TLR9). Fluorescent labelling of rHb- FAR-l revealed a specific internalization of this antigen into vesicular compartments of both DCs and M<l>s, within 45 min. The immunoregulatory potential of rHb-FAR-l was tested in the non-obese diabetic mouse. Despite the lack of T cell polarization after intraperitoneal (ip) injection of rHb-FAR-l, the protein was able to induce increased levels of IL-lO by splenocytes in response to rHb-FAR-l ex vivo. Subsequent to these ip injections an expanded number of peritoneal exudate cells (PECs) was found in the peritoneal cavity, with a higher percentage of Bl B cells but a lower frequency of peritoneal M<l>s. The latter cell type displayed up-regulated expression of FIZZl and arginase-I, both markers for alternative activation of M<l>s. In addition to in vivo effects, rHb-FAR-l directly stimulated the expression of IL-6, IL-lO and TNF-a from naive PECs in vitro but was not able to interfere with the proliferation of splenic Band T cells. Furthermore, rHb-FAR-l failed to induce Foxp3 expressing regulatory T cells but polarized Th17 cells in an IL-6 dependent manner, suggesting a potential importance in the early transient Th17 response to H. bakeri, which prolongs parasite survival.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:580307 |
| Date | January 2011 |
| Creators | Siebeke, Ivonne |
| Publisher | University of Nottingham |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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