Interaction between the immune and nervous systems is critical for repair, tumour surveillance and protection from infection. Toll-like receptors are evolutionarily ancient pattern-recognition receptors that are implicated in innate responses to infection. In work done in Canada we have shown that signals delivered through the Toll-like receptor-2 (TLR2) are critical for early microglial response to experimental brain injury. Microglia are macrophage-like cells in the brain and spinal cord that act as rapid sensors of loss of homeostasis of any kind. Activated microglia can recruit T cells and macrophages to the central nervous system via production of chemokines and cytokines.
Our findings suggest that endogenous ligands for TLR2 are produced within the brain independent of infection and these ligands direct microglial response. This represents a novel role for TLR's in brain function. In Denmark, we propose to identify TLR2 ligands that are produced in response to injury in the mouse brain. Localized axona l degenerative injury will be used to induce TLR2 ligands. These will be assayed for their ability to induce intercellular signals, and mRNA and protein for specific cytokines and chemokines in microglial cultures.
Criteria for functional TLR2 ligands will include purification with TLR2-Fc but not TLR4-Fc fusion proteins, and induction of responses in wild-type but not TLR2- or signalling-deficient microglia, that are blocked with TLR2- specific reagents. Ligands that meet these criteria will be characterized by proteomic analysis including 1- and 2-D gel electrophoresis, immunoblotting and mass spectrometry. Ligands are expected to be lipoproteins or peptidoglycans - our assays will detect both. Ligand-specific monoclonal antibodies will be generated and used to probe location and function. Long-term goals include gene manipulation in animal models.
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