| 1,3-dichloro-2-propanol(1,3-DCP)as one of the chloropropanols which are foodborne contaminants,is produced in the processing of food products.It was reported that 1,3-DCP has a lot of biological hazard and has been shown to induce hepatocyte lipid accumulation,so cause Non-alcoholic fatty liver disease(NAFLD)in mice.Lipid metabolism disorder is closely associated with obesity,diabetes,metabolic syndrome,cardiovascular disease,cancer and so on.Therefore,it is necessary to protect against 1,3-DCP-induced lipid metabolism disorder.Drinking green tea has lipid-lowering effect in our daily life,and one of the main active ingredients in green tea is Epigallocatechin-3-gallate(EGCG).EGCG can improve the metabolism,and have anti-oxidation,immune regulation,anti-tumor,anti-inflammatory and other kinds of biological activities.EGCG can effectively modulate cholesterol metabolism and toxicity induced by inhibition of dietary and alleviate liver injury.However,it remains unclear whether EGCG influences the 1,3-DCP-induced lipid metabolism abnormal and the deeper mechanism.The present study was therefore to investigate the intervention of EGCG on 1,3-DCP-induced metabolic abnormalities in vitro and in vivo.The experiment explained the mechanism from the protein and mRNA level combining with the in vitro and in vivo experiment.My experiments have important theoretical and practical significance to establish the new method of controlling 1,3-DCP and create a role of EGCG in dietary.My experiment is the first time to explore the protective effect and mechanism of EGCG on 1,3-DCP induced lipid metablism disorder in vitro and in vivo.The experiments are divided into two parts: In vitro: I established a model of lipid accumulated HepG2 cells with 1,3-DCP and observed the lipid accumulation when HepG2 cells were treated with different concentrations of EGCG.In the present study,I identified whether the EGCG protect against 1,3-DCP via Adenosine 5’-monophosphate(AMP)-activated protein kinase(AMPK)or protein kinase A(PKA)single pathway in vitro;In vivo: I established a model of lipid metabolism disorder mice with 1,3-DCP and observed the lipid accumulation in liver when C57BL/6J mice were administrated with different concentrations of EGCG.In the present study,I identified whether the EGCG protect against 1,3-DCP via AMPK or PKA single pathway in vivo.In vitro,cell viabilities of HepG2 cells treated with 1,3-DCP or EGCG were analyzed by MTT assay.0-20 μM EGCG and 0-0.4 mM 1,3-DCP were selected for further study.The results of Oil red O staining,TG and TC showed that the lipid accumulation does of 1,3-DCP is 80 μM and the protective doeses of EGCG are 5,10,20 μM.Besides,EGCG(5,10 and 20 μM)could significantly inhibit 1,3-DCP(80μM)-induced intracellular lipid accumulation in a dose dependent manner.The Western Blot data showed that EGCG significantly increased 1,3-DCP induced cAMP level,the protein expression of P-PKA,Phospho-cAMP-response element binding protein(P-CREB),PPARγ coavtivator-1α(PGC-1α)and Silent information regulator T1(SIRT1)in HepG2 cells.It was demonstrated that EGCG prevented 1,3-DCP induced lipid accumulation via PKA signaling pathway in HepG2 cells.Furthermore,EGCG increased 1,3-DCP induced protein expression of Liver kinase B1(LKB1)and P-AMPK,Phospho-Acetyl-CoA carboxylase(P-ACC)and Carnitine palmitoy transferase 1(CPT1).On the contrary,EGCG decreased 1,3-DCP induced protein expression of Glycerol-3-phosphate acyltransferase(GPAT),and Fatty acid translocase(FAT/CD36),3-hydroxy-3-methylglutaryl-CoA reductase(HMGCR),Sterol regulatory element-binding protein-1c(SREBP1c),stearoyl CoA desaturase 1(SCD1)and Fatty acid synthase(FAS).It was concluded that EGCG was effective in reducing 1,3-DCP-induced lipid accumulation via AMPK signaling pathway.In vivo,I examined the changes of serum TG,TC,Low density lipoprotein cholesterol(LDL-C),High density lipoprotein cholesterol(HDL-C),Glutamic-pyruvic transaminase(GPT/ALT)and Glutamic oxalacetic transaminase(GOT/AST).It proved that EGCG could alleviate the 1,3-DCP induced hyperlipidemia and hepatic cell damage in mice.Furthermore,the TC,TG level and HE,Oil Red O staining of liver indicated that EGCG could protect against 1,3-DCP-induced steatosis in liver.The Western Blot and qRT-PCR results showed that EGCG significantly increased 1,3-DCP induced protein and mRNA expressions of cAMP,P-PKA,P-CREB,LKB1,P-AMPK,P-ACC and CPT1 in liver,while decreased 1,3-DCP induced GPAT,CD36,HMGCR,SREBP1 c and Sterol regulatory element-binding protein-2(SREBP2).Thus,the experiment demonstrated that EGCG treatment produced improvements in lipid metabolic profiles and activated PKA and AMPK protein in 1,3-DCP induced mice.In conclusion,EGCG can protect against 1,3-DCP-induced lipid metabolism disorder via PKA and AMPK signaling pathways. |
