Involvement of TLR4/Type I IL-1 Receptor Signaling in the Induction of Inflammatory Mediators and Cell Death Induced by Ethanol in Cultured Astrocytes

Abstract

Activated astroglial cells are implicated in neuropathogenesis of many infectious and inflammatory diseases of the brain. A number of inflammatory mediators and cytokines have been proposed to play a key role in glial cell-related brain damage. Cytokine production seems to be initiated by signaling through TLR4/type I IL-1R (IL-1RI) in response to their ligands, LPS and IL-1β, playing vital roles in innate host defense against infections, inflammation, injury, and stress. We have shown that glial cells are stimulated by ethanol, up-regulating cytokines and inflammatory mediators associated with TLR4 and IL-1RI signaling pathways in brain, suggesting that ethanol may contribute to brain damage via inflammation. We explore the possibility that ethanol, in the absence of LPS or IL-1β, triggers signaling pathways and inflammatory mediators through TLR4 and/or IL-1RI activation in astrocytes. We show in this study that ethanol, at physiologically relevant concentrations, is capable of inducing rapid phosphorylation within 10 min of IL-1R-associated kinase, ERK1/2, stress-activated protein kinase/JNK, and p38 MAPK in astrocytes. Then an activation of NF-κB and AP-1 occurs after 30 min of ethanol treatment along with an up-regulation of inducible NO synthase and cyclooxygenase-2 expression. Finally, we note an increase in cell death after 3 h of treatment. Furthermore, by using either anti-TLR4- or anti-IL-1RI-neutralizing Abs, before and during ethanol treatment, we inhibit ethanol-induced signaling events, including NF-κB and AP-1 activation, inducible NO synthase, and cyclooxygenase-2 up-regulation and astrocyte death. In summary, these findings indicate that both TLR4 and IL-1RI activation occur upon ethanol treatment, and suggest that signaling through these receptors mediates ethanol-induced inflammatory events in astrocytes and brain.

Critical Role of TLR4 Response in the Activation of Microglia Induced by Ethanol

“…Furthermore, cell treatment with either high concentrations of ethanol or lipid raft disrupting agents (streptolysin O or saponin) inhibits ethanol-induced activation of the TLR4 signaling pathway (24). The effects of ethanol on TLR4 appear to be specific, because other short-chain n-alcohols do not induce TLR4 recruitment, although they slightly activate MAPKs signaling (23), and because other TLRs seems not to appear to be activated by ethanol (our unpublished results). We demonstrate, however, that ethanol activates TLR4/IL-1RI receptors in astrocytes (42).…”

Drug addiction: targeting dynamic neuroimmune receptor interactions as a potential therapeutic strategy

“…primary microglial and astrocyte cultures. 13 Similarly, morphine increases the expression of CCL2, CCL5 and IFNγ-inducible protein. 14 In vivo, systemic injections of either opioids and alcohol increases the expression of inflammatory mediators in key neuroanatomical areas associated with addiction such as the prefrontal cortex and hippocampus but not the cortex within mice.…”

Opportunities for the Development of Neuroimmune Therapies in Addiction

“…The activation of TLR4 predominantly contributes to glial activation and the subsequent release of numerous proinflammatory cytokines (Mayfield et al, 2013). Importantly, these TLR4-related processes are involved in the behavioral and neuroinflammatory effects of drugs of abuse (Mayfield et al, 2013), as TLR4 activation has been shown to be integral to alcohol-induced glial activation and proinflammatory signaling (Alfonso-Loeches, Pascual-Lucas, Blanco, Sanchez-Vera, & Guerri, 2010;Blanco, Pascual, Valles, & Guerri, 2004;Blanco, Valles, Pascual, & Guerri, 2005;Fernandez-Lizarbe, Pascual, & Guerri, 2009), as well as alcohol's behavioral effects in rodents (Wu et al, 2012). Furthermore, in rodents, naltrexone attenuates proinflammatory TLR4-related signaling (Hutchinson et al, 2011) and blocks ethanolinduced glial activation and neuronal death (Qin & Crews, 2012), while (+) naloxone reduces acute alcohol-induced sedation and motor impairment (Wu et al, 2012).…”