FK866 Attenuates Acute Hepatic Failure Through C-jun-N-terminal Kinase(JNK)-dependent Autophagy

Background and ObjectivesFK866,as the highly specific small molecule noncompetitive inhibitor of nicotinamide phosphoribosyltransferase(NAMPT),exhibits a protective effect on acute liver failure(ALF),but the mechanism by which FK866 affords this benefit has not yet been elucidated.Autophagy has a protective effect on acute liver injury.However,the contribution of autophagy to FK866-conferred hepatoprotection is still unclear.C-jun N-terminal kinase(JNK),has been reported to play a critical role in determining hepatocyte damage during liver failure and be a potent negative regulator of autophagic response.This study aimed to investigate whether FK866 could attenuate D-galactosamine(GaIN)/lipopolysaccharide(LPS)and concanavalin A(ConA)-induced ALF through JNK-dependent autophagy.MethodsIn the current study,Eight-week-old male C57BL/6 mice and primary hepatocytes were as experimental objects.First,in vivo,ALF mice models were established using GaIN/LPS and ConA.Mice were pretreated with FK866 at 24,12,and 0.5 h before treatment with GaIN/LPS and ConA.The serum aminotransferase levels were detected,and the liver damage was determined using ematoxylin and eosin(HE),and the mRNA expression levels of proinflammatory cytokines in mice livers were measured by quantitative polymerase chain reaction(PCR),which used to explore the function of FK866 on mice ALF.The protein expression levels of autophagy indicators were measured using western blot method and the effect of FK866 on autuphagy was investigated.Suppression or induction of autophagy was conducted by injection of 3-methyladenine(3MA)or rapamycin,and the contribution of autophagy to FK866-conferred benefit in ALF was determined.JNK activity was inhibited by SP600125 and the role of JNK in the FK866-induced autophagy was investigated.Second,in vivo,primary hepatocytes of C57BL/6 mice were isolated and cultured,and primary hepatocytes were stimulated using GaIN/LPS and treated the above drugs in animal experiments.In addition,autophagy-related protein 7(Atg7)and Jnk specific small interfering RNAs were used to knockdown Atg7 and Jnk genes respectively.The formation of autophagosomes and autolysosomes was observed by infection with adenovirus-encoding mRFP-GFP-LC3B,and the autophagic flux was futher analyzed.The hepatocyte toxicity was evaluated using lactate dehydrogenase-cytotoxicity colorimetric assay kit.ResultsBoth in GaIN/LPS and ConA-induced ALF models in mice and GaIN/LPS-induced hepatotoxicity in cultured hepatocytes,FK866 pretreatment significantly attenuated hepatocellular injury and promoted autophagy and inhibited JNK activation.Induction of autophagy by rapamycin could mimic FK866-afforded hepatoprotective effect.Suppression of autophagy using 3MA or Atg7 siRNA impaired FK866-conferred hepatoprotection.Inactivation of JNK increased autophagy and exhibited clearly hepatoprotective function.ConclusionsFK866 could ameliorate GaIN/LPS and ConA-induced ALF through induction of autophagy while suppression of JNK.These findings suggest that FK866 acts as a simple and applicable preconditioning intervention to protect against ALF;autophagy and JNK may also provide therapeutic targets for ALF treatment.