Key components of the innate immune response to infections are the Toll-like receptors
(TLRs), which are able to detect invading pathogens and subsequently generate inflammatory
responses. Many details of the signalling pathways of TLRs have emerged from gene targeted
mice or inhibition studies in transformed cell lines. However, the signalling pathways activated
in primary human cells and disease tissues are less well understood. Previous studies identified
differences in TLR signalling between human cells of myeloid- and non-myeloid origin. While
over-expression of a dominant negative construct of the TLR adaptor molecule MyD88
inhibited TLR4 signalling in HUVECs it had no effect on TLR4 signalling in macrophages.
Based on this observation, this thesis examined the function of MyD88 in primary human
monocyte derived dendritic cells (DCs). Unexpectedly, over-expression of MyD88 dn resulted
in the activation of DCs. Subsequent experiments, provided evidence for a DC specific
inhibitory mechanism, which depends on endogenous MyD88 and is disrupted by TIR domain
over-expression. To further investigate the mechanism, the function of SIGIRR in human DCs
was examined. SIGIRR is a member of the TIR domain containing receptor family that has
been shown to be expressed in murine and human DCs but not macrophages. However, results
in this thesis show that SIGIRR is expressed by human DCs as well as macrophages. While
SIGIRR has been studied in murine models, nothing is known about its function in primary
human cells. Therefore, an adenoviral construct encoding wild-type SIGIRR was generated and
over-expressed in DCs and macrophages, which impaired TLR2, TLR3, TLR4, TLR5, TLR7/8
and IL-1R but not TNFα signalling. In accordance with these results, siRNA knock down of
SIGIRR in macrophages led to an increase of TLR3, TLR4, TLR7/8 and IL-1R but not TNFα
induced cytokine production. Therefore, SIGIRR seems to be an inhibitor of TIR domain
dependent signalling, affecting the MyD88 dependent as well as the TRIF dependent signalling
pathway. Furthermore, immunoprecipitation studies of SIGIRR with MyD88 suggest, that
SIGIRR interacts with MyD88 constitutively through TIR domain interaction, indicating that
SIGIRR inhibits TLR/IL-1R signalling by sequestering MyD88. Given the potency of SIGIRR
to inhibit TLR/IL-1R signalling in human DCs and macrophages, this thesis further
investigated its role in rheumatoid arthritis disease models. Over-expression of SIGIRR wt in
human RA synovial membrane cultures inhibited the spontaneous secretion of cytokines by
those cells. In contrast, SIGIRR deficient mice were resistant to collagen induced arthritis
(CIA). SIGIRR null mice immunised with CIA showed a loss in IgG2a anti-collagen antibody
production as well as reduced Th1 and Th17 immune responses but increased Th2 immunity.
Subsequent results indicated that SIGIRR is able to regulate CD4+ T cell development through
the inhibition of ST2/IL-33 signalling. Therefore, while SIGIRR inhibits pro-inflammatory
cytokine release during the progression stages of RA, it may also regulate the development of
Th2 development, thereby reducing CIA incidence. These studies highlight the importance of
investigating signalling pathways in physiologically relevant cells in order to fully understand
the roles TLRs and specific signalling molecules play in the human immune system and human
disease process.