Gentiopicroside Ameliorates P2x7R-NLRP3 Inflammasome-mediated Alcoholic Hepatosteatosis

Scope:Gentiopicroside(GPS),which is attributed to the bitterness of gentian root extract,possesses multiple pharmacological activities.Since,regulating P2x7R-NLRP3inflammasome activation with small molecule inhibitors might be a potential therapeutic strategy to treat and prevent alcoholic hepatosteatosis,we investigated whether this process would be modulated by GPS.Methods:An in vivo model was established by intragastrically treating mice with ethanol ethanol(5 g/kg,body weight),and an in vitro model was created by treating HepG2 cells with 50 mM ethanol or treating RAW 264.7 macrophages and murine bone marrow-derived macrophages(BMDM)with LPS and subsequently stimulated with 3mM ATP.Results:In mice model of alcohol-induced liver steatosis,GPS significantly decreased serum aminotransferase and accumulation of triglyceride.GPS elevated LKB1/AMPK,which was associated with the downregulation of Srebp1 and ACC.Additionally the Inhibition of PPARa and of ACC phosphorylation were recovered by GPS.GPS administration effectively blocked IL-1β production and caspase-1 activation,as well as NLRP3 and P2x7R protein expression.In HepG2 cells,ethanol-induced lipid accumulation was associated with decreased LKB1/AMPK,phosphor-ACC and PPARa and increased srebp1 and ACC.These changes were reserves by GPS.In addition GPS treatment inhibited the increase of P2x7R protein level,IL-1β protein secretion and caspase-1 activity.P2x7R gene silencing enhanced phosphorylation of AMPK,and reduced srebpl expression in ethanol-treated hepatocytes.Moreover,GPS down-regulates P2x7R-mediated inflammatory response against extracellular ATP in LPS-primed RAW 264.7 macrophages and BMDM.P2x7R deficiency attenuated IL-1β cleavage in response to ATP in LPS-primed RAW 264.7 macrophages,and GPS could further suppress IL-1β cleavage.Conclusion:Activation of LKB1/AMPK signaling alcoholic steatohepatitis by GPS might be mediated P2x7R-NLRP3inflammasome,suggesting a therapeutic utility of P2x7R blockade in the treatment of ALD...