The mechanisms that mediate innate recognition of infections in the immunospecialised
environment of the central nervous system (CNS) have not been characterised. This thesis
explores the capability of the CNS to detect infections and activate immune responses. The
majority of CNS neurones are post-mitotic and cannot be replaced if lost or damaged.
Consequently, the resting CNS is devoid of most immune processes, although substantial
inflammatory responses can be initiated by specialised glial cells. The innate immune system
recognises conserved molecular patterns on microorganisms by a set of pattern recognition
receptors which include the Toll-like Receptors (TLR). A growing consensus suggests they
are key to the initiation of innate immune responses. Cellular expression of TLR imparts the
ability to detect infection and determine pathogen type. A multitude of TLRs have now been
cloned, although their function and expression patterns have not been described in the CNS.
This thesis aims to explore whether cells of the CNS express TLRs and whether they are
capable of responding to different stimuli. To explore gene expression a novel custom
microarray was designed, developed and validated to assay the expression of selected gene
transcripts involved in innate immune responses. These included a multitude of pattern
precognition receptors, in addition to transcripts associated with stress responses and a
variety of cytokine, chemokine and interferon (IFN) transcripts. In addition, a highly
sensitive quantitative PCR technique was developed. Utilising both techniques this thesis
reports the first systematic analysis of TLR gene expression in the CNS.Gene transcript levels were first studied in glial cells at rest. Cells were stimulated with
bacterial lipopolysaccharide, or by infection with the neuroinvasive Semliki Forest virus
(SFV). Both microglia and astrocytes in culture expressed a multitude of TLRs that were
differentially modulated in a specific manner depending on the nature of the stimulus. The
expression of TLR suggests glial cells are capable of recognising a vast array of microbialassociated
molecules. Such a strategy may be an essential requirement for an organ mostly
devoid of recognisable immune processes.In vivo, the resting CNS exhibited extensive TLR expression with TLR 3 expressed at
exceptionally high levels, comparable to that of lymphoid tissue, but varying with mouse
strain. The data reported here show for the first time that TLRs in the brain are upregulated
during viral encephalitis. Furthermore, this response was appropriate to the pathogen, with
selective up-regulation of TLRs that sense viral infection. Intracerebral inoculation with
either SFV or rabies virus initiated substantial upregulation of TLR 2, 3 and 9. Type-I IFN
independent mechanisms mediated the up-regulation of TLR 2 following SFV infection,
whilst for the two TLRs that mediate recognition of viral nucleic acids, TLR 3 and TLR 9,
upregulation of gene expression was dependent upon and proportional to the type-I IFN
response. It is likely that by up-regulating TLR 3 and 9, type-I IFN acts to increase the
sensitivity of cells in the vicinity of virally infected cells. In this hypothesis, basal levels of
TLR detect viral RNA and induce type-I IFN synthesis. This IFN acts in both an autocrine
and paracrine way to up-regulate a number of genes including TLR itself. In this way cells in
the vicinity of virally infected cells have their virus sensing mechanisms upregulated. This
parallels events with protein kinase R, another interferon inducible activator of innate
cellular defences.Transmissible spongiform encephalopathies are a group of diseases characterised by
chronic neurodegeneration and glial cell activation. This thesis demonstrates that the CNS
significantly upregulated several TLRs, and in the case of TLR 2, by 10-fold towards
terminal disease. This response further describes the apparent non-productive innate
immune activation of these cells during these diseases. In summary, the finding that the
brain has the ability via TLR expression to detect infection and discern its type provides an
important contribution to understanding pathological processes in this organ
