Our previous findings showed that TLR2 and TLR4 are expressed in cerebral cortical neurons, where their levels and downstream signaling via JNK are increased in response to energy deprivation in cell culture and ischemia in vivo (Tang et al., 2007
). In the present study we show that TLR4 expression increases during exposures to Aβ1-42 and the lipid peroxidation product HNE. JNK and caspase-3 activity levels were increased in neurons exposed to Aβ and HNE, and selective elimination of TLR4 function significantly suppressed the abilities of Aβ and HNE to induce activation of JNK and caspase-3. These findings suggest that neurons expressing TLR4 are vulnerable to degeneration in AD. Consistent with the latter possibility, we found that levels of TLR4 are decreased in AD brain tissue samples compared to samples from control subjects. The latter result suggests that neurons expressing TLR4 are vulnerable to degeneration in AD because TLR4 is expressed primarily in neurons (Tang et al., 2007
). Moreover, TLR4 expression is increased in glial cells in response to activation (inflammation); because glial cell activation is increased in AD brain (Mrak and Griffin, 2005
), it is unlikely that the decrease in TLR4 in AD is due to a decrease in glial cells. Collectively, our findings from cell culture studies and analysis of AD and control subjects suggest a possible contribution of TLR4-mediated signaling in the neuronal degeneration that may be triggered by Aβ and membrane lipid peroxidation in AD.
TLRs are widely conserved in organisms ranging from sponges to humans, and are believed to have evolved as a mechanism to protect against invading pathogens including bacteria, viruses and parasites (Beutler, 2005
; Barton, 2007
; Wiens et al., 2007
). However, TLRs are also activated in immune cells (T cells, dendritic cells and macrophages) in response to tissue injury, including trauma, hemorrhagic shock and ischemia (Mollen et al., 2006
; Frink et al., 2007
). We found that TLR2 and TLR4 are up-regulated and activated in neurons in response to an ischemic stroke (Tang et al., 2007
). Previous studies have shown that other molecules involved in innate immunity, including components of the complement cascade, are activated in neurons and glial cells in AD (Nataf et al., 1999
; Mack et al., 2006
; Shen and Meri., 2003
; Zanjani et al., 2005
). Our findings suggest that TLR4 signaling may also play a role in AD pathogenesis, possibly being activated by Aβ and membrane-associated oxidative stress. While activation of TLR4 in neurons might contribute to their death, the activation of TLR2 in microglia has been reported to enhance their phagocytic uptake of Aβ, a potentially beneficial process (Chen et al., 2006
). Another recent study by Tahara and colleagues demonstrated that activation of a microglial cell line (BV-2 cells) with TLR2, TLR4 or TLR9 ligands enhances their uptake of Aβ (Tahara et al., 2006
). Therefore, the roles of TLRs in AD are likely to be complex with activation of different TLRs resulting in different types of responses in neurons and glial cells.
Molecules produced by pathogens have been shown to act as TLR ligands (Trinchieri and Sher., 2007
). However, an unresolved issue in the field of TLRs concerns the endogenous ligands that activate them during tissue injury and inflammatory disease states. It has been proposed that heat-shock protein 70 (HSP70) is a ligand for TLR2 and TLR4 (Asea et al., 2002
; Vabulas et al., 2002
). Levels of HSP70 are increased in AD (Yoo et al., 1999
) and we have observed that levels of HSP70 are increased in association with TLR2 and 4 activation in neurons subjected to energy deprivation (Tang et al., 2007
) and in neurons exposed to HNE (unpublished data). Another candidate for the endogenous ligand that activates TLR4 in neurons in response to Aβ or HNE mediated toxic conditions is hyaluronan fragments. Levels of hyaluronate were reported to be increased in the temporal cortex of AD patients compared to age-matched control subjects (Jenkins and Bachelard, 1988
). Hyaluronases are activated in injured neural tissue resulting in the generation of hyaluronan fragments (Al'Qteishat et al., 2006
) that have been reported capable of activating TLR2 and TLR4 in immune cells (Termeer et al., 2002
; Jiang et al., 2005
). It is therefore possible that hyaluronan fragments contribute to the activation of TLRs in neurons and glial cells in AD.
Activation of one or more TLRs in neurons and glial cells may occur in response to injury to the nervous system. Our findings suggest that membrane lipid peroxidation is one injury-associated factor that can result in TLR4 activation in neurons. Lipid peroxidation and HNE are implicated in the degeneration of neurons that occurs in ischemic stroke (Keller et al., 1998
) and several neurodegenerative disorders including AD (Keller and Mattson, 1998
), ALS (Pedersen et al., 1998
) and Parkinson's disease (Yoritaka et al., 1996
). It will therefore be of considerable interest to elucidate the relationships between oxidative stress, TLR activation and the pathogenesis of these different disorders. Our findings therefore suggest that agents that target TLR expression, endogenous TLR ligands or TLR signal transduction molecules might be of therapeutic value for such disorders.