This project will examine the functional consequences of Toll-like receptor-3 (TLR-3) expression as it relates to features of Alzheimer's disease. Toll-like receptors, which are a family of 10 related pattern recognition receptors, have multiple functions in coordinating host defenses to microbial agents. We identified TLR-3 as being prominently expressed by microglia in AD brains, particularly those plaque-associated, while other significant TLR were not associated with glia. As TLR-3 signaling can have different consequences, we consider it timely to investigate in AD and related relevant human model systems. As TLR-3 is primarily an endosomal localized receptor, its function and the consequence of its activation to AD requires investigation. We have developed a central hypothesis based on preliminary results that stimulation of microglia and astrocytes with ligands for TLR-3 will induce anti-inflammatory and protective responses, and also promote A? removal. As stimulation of related endosomal TLR-9 has been suggested as a therapeutic option for removing amyloid from brain, we are positing that TLR-3 activation may have similar outcome. The goal of this project is to identify the nature of microglia and astrocyte responses to TLR-3 ligands in the presence of A?. This will be investigated in vitro in microglia and astrocyte cultures derived from postmortem brains, and in ex-vivo human brain slice cultures. We will also determine if chronic TLR-3 ligand administration to plaque-developing transgenic mice affects A? load and related neuropathology. We will investigate these features in three related aims.
Specific Aim 1; We will investigate what are the functional consequences of TLR-3 activation of human microglia and astrocytes on production of proinflammatory and anti-inflammatory cytokines and neurotoxic factors. Cultures from AD and normal brains will be stimulated with a TLR-3 ligand in the presence and absence of A?. We will use antibody arrays to measure cellular responses, and also in vitro neurotoxicity and neuroprotective assays to determine consequences of TLR-3 stimulation.
Specific Aim 2; We will investigate what are the functional consequences of TLR-3 activation of human microglia and astrocytes on uptake and degradation of amyloid beta peptide.
Both aims will be carried out in vitro using the TLR3- ligand poly IC and/or A? stimulated human microglia and astrocytes described in Aim 1.
Specific Aim 3; We will use two complex systems, where interactions of microglia, astrocytes and neurons occur, to investigate the same features as in aims 1 and 2. We will use ex vivo slice cultures of human brains and also amyloid plaque developing transgenic mice to determine whether TLR-3 activation promotes or reduces inflammation and neuronal survival, and increases or decreases glial uptake and degradation of A?.

Public Health Relevance

The relevance of this research to public health is that it could identify new features of inflammation in Alzheimer's disease. We are positing that activation of Toll-like receptor-3 in the human brain could be a target for treatment. At present, we do not know if this signaling needs to be enhanced or inhibited in Alzheimer's disease, there is evidence for both features;however either could be significant approaches to changing the path of inflammatory responses in neurodegeneration. These findings will not only be relevant for Alzheimer's but other neurodegenerative diseases.

National Institute of Health (NIH)
National Institute on Aging (NIA)
Exploratory/Developmental Grants (R21)
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Special Emphasis Panel (ZRG1-BDCN-W (02))
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Refolo, Lorenzo
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Banner Health
United States
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