Ethanol modulates astrocyte activation and neuroinflammation via miR-339/NLRP6 inflammasome signaling

Alcohol is the most abused substance among adolescents and has a profound impact on health, society, and the economy. Alcohol intoxication is linked to neuroinflammation and neuronal damage, which result in behavioral alterations such as motor dysfunction, neuronal injury, cognitive deficits, and inflammation. Alcohol-induced neuroinflammation is associated with the activation of central nervous system cells, including astrocytes, and the release of proinflammatory cytokines. In this study, we investigated the role of the NLRP6 inflammasome signaling pathway in inducing cellular activation and neuroinflammation in human primary astrocytes exposed to ethanol. Our results demonstrated that ethanol upregulates the expression of NLRP6 inflammasome signaling mediators, including NLRP6, caspase 1, and proinflammatory cytokines IL-1β and IL-18, in human primary astrocytes. Gene silencing studies using NLRP6 siRNA further validate ethanol-mediated activation of NLRP6, cleavage of caspase 1, IL-1β, and IL-18 in human primary astrocytes. miR array analysis of ethanol-exposed human primary astrocytes reveals decreased levels of miR-339, accompanied by an upregulation of NLRP6 inflammasome signaling and astrocyte activation. Through bioinformatics analyses, Argonaute immunoprecipitation assays, and miR-339 overexpression experiments, we identify NLRP6 as a novel 3'-UTR target of miR-339. Overall, our findings confirmed the involvement of miR-339 in NLRP6 inflammasome signaling and its association with cellular activation and neuroinflammation in human primary astrocytes exposed to ethanol and provide novel insights highlighting a previously unrecognized mechanism in alcohol-induced neuroinflammation.