Dendritic cells (DCs) are regulators of the immune response and are thought to be critical in maintaining tolerance towards the intestinal microbiota. Recent data have identified distinct subsets of DCs with specific functional properties. The objective of this thesis was to further define CD103⁺ and CX3CR1⁺ DCs in the intestine and to determine how DCs and regulatory T (Treg) cell responses are influenced by the microbiota. Using multicolour flow cytometry, we identified two CD103⁺ DC subsets with differential aldehyde dehydgrogenase (ALDH) activity and two populations of CX3CR1⁺ cells. In the mesenteric lymph node CD103⁺ALDH⁺ DCs were highly mature (CD86<sup>hi</sup>, MHCII<sup>hi</sup>), likely migratory (CCR7⁺) and enhanced Treg cell induction compared with ALDH⁻ DCs. CX3CR1<sup>int</sup> cells accumulated during bacterially-induced colitis suggesting a pro-inflammatory role whereas CX3CR1<sup>hi</sup> cells were associated with the production of the anti-inflammatory cytokine IL-10 during homeostasis. We also assessed the generation of CD103⁺ DCs from bone marrow progenitors. Although only small proportions of CD103⁺ DCs were detected in culture with FLT3L or GM-CSF alone, the combination of FLT3L and GM-CSF induced CD103⁺ DCs with a phenotype similar to those found in the small intestine. Using this system we showed that TLR ligands and retinoic acid induce ALDH enzyme activity in vitro. In order to assess how DCs and Treg cells respond to changes in the microbiota we employed broad-spectrum antibiotic treatment to deplete endogenous bacteria and also analyzed the impact of colonization with the model organism Helicobacter hepaticus. Interestingly, we did not detect alterations in the proportions of different DC subsets following antibiotic treatment or H. hepaticus infection. However, using a novel FoxP3<sup>huCD2</sup>-IL-10<sup>GFP</sup> reporter mouse, we found that IL-10 production by Treg cells was ablated following antibiotic treatment and significantly elevated following H. hepaticus infection. Preliminary investigation of the mechanism underlying this effect suggests a role for IL-27. In summary, this thesis provides further detail on the phenotype of intestinal DCs and shows that Treg cell IL-10 production is sensitive to the composition of the microbiota.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:655006 |
| Date | January 2011 |
| Creators | Johnson, Andrew M. F. |
| Contributors | Powrie, Fiona |
| Publisher | University of Oxford |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://ora.ox.ac.uk/objects/uuid:14284c3c-1aa4-4125-ad31-e74ded4e75bc |
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