Celecoxib Regulates Autophagy In Nonalcoholic Fatty Liver Disease

OBJECTIVE: Nonalcoholic fatty liver disease(NAFLD)is one of the leading causes of chronic liver diseases around the world,and is charactered by an excessively high accumulation of fat deposits in the liver resulting from causes other than chronic alcohol abuse and drugs.It is widely acknowledged that insulin resistance,dysfunctional lipid metabolism,oxidative stress,inflammation,impaired autophagic flux,apoptosis/necrosis may all contribute to NAFLD.Celecoxib is a selective COX-2 inhibitor which has the effects of antipyretic,analgesia,and anti-inflammatory.For that reason,it has been used for releasing pain and curing rheumatoid arthritis in clinic.Recent studies have suggested that celecoxib has the effects of ameliorating NAFLD,but the underlying mechanisms remain unknown.Here,we discuss the possible mechanisms of celecoxib regulating autophagy on ameliorating NAFLD.METHODS: Hepatic cells L02 were treated with different concentrations of palmitate(200μM,400μM)with or without celecoxib(20μM)in vitro for certain time(12h,24h).Red oil O and triacylglyceride(TG)kit were used to test the level of steatosis of hepatic L02 cells.The changes in protein expression of COX-2,LC3 II/I,p62 were detected by western blot.Autophagic flux was evaluated by western blot and RFP-GFP double fluorescence system in the above treated cells.L02 steatosis cells were treated with rapamycin or chloroquine to induce or to inhibit autophagy respectively,then steatosis level and autophagic flux wasevaluated as above.COX-2 siRNA and celecoxib were used to inhibit COX-2.SD rats were feeded with high-fat diet(HFD)for 4 weeks,then were intragastricly administrated with celecoxib(20mg/kg/day)or normal saline for another 4 weeks along with HFD.Hematoxylin and eosin(HE)staining was used to indicate lipid deposits.Serum was collected to detect biochemical parameters(ALT、AST、TG、TC).RESULTS: Lipids were accumulated in L02 cells when treated with palmitate in a time and dose dependent manner.Protein expression of LC3 II/I was higher and p62 was lower on the early stage of steatosis in L02 cells,however,on the late stage both of them were higher,indicating that autophagic flux was activiated on the early stage of steatosis,but blocked on the late stage.At the same time COX-2 was induced in above cells.Rapamycin(autophagy agonist)alleviated steatosis with activating autophagic flux while chloroquine(autophagy inhibitor)aggravated steatosis with inhibiting autophagic flux.When Celecoxib was added to L02 cells treated with palmitate,steatosis was alleviated with decreasing COX-2 as well as treated with COX-2 siRNA.Western blot and RFP-GFP double fluorescence system indicated that autophagic flux was partially restored in steatosis L02 cells by treatment of Celecoxib or COX-2 siRNA.In animal models,the level of liver steatosis in high-fat diet rats was even worse than that of control with higher body weight and higher level of blood ALT,AST,TG,TC.Furthermore,celecoxib could ameliorate liver steatosis and the above biochemical parameters.CONCLUSIONS: Autophagy protects NAFLD by alleviating steatosis.Autophagic flux is activiated on the early stage of steatosis and blocked on the late stage.Celecoxib partially restores autophagic flux via downregulation of COX-2 and alleviate steatosis in hepatic cells.Celecoxib could ameliorate liver steatosis in high-fat diet rats.