Study On The Role Of Liver-specific Konckout Of Sphingosine Kinase 2 And Polydatin In Non-alcoholic Fatty Liver Disease

Part 1Scope:Sphingosine kinases?SphKs?are the key enzymes that catalyze the transformation of sphingosine into the bioactive substance sphingosine-1-phosphate?S1P?in vivo.SphKs has two subtypes,SphK1and SphK2,to regulate the dynamic balance of S1P/ceramide which closely related to many diseases.However,the role of SphK2 in NAFLD is still unclear.The purpose of this study was to explore the role and molecular mechanism of SphK2 in NAFLD as to provide a experimental basis for the treatment of NAFLD and the screening of drug targets.Methods:1:C57BL/6 mice?WT?and SphK2 knockout mice(SphK2^Hep-/-)were fed with high-fat diet?HFD?for 20 weeks.2:Primary hepatocytes were induced by palmitic acid?PA?in vitro.3:Mice liver metabolomics was used to screen out the target metabolites related to SphK2 and explore the relationship between SphK2 and metabolites in hepatocytes.Results:The SphK2^Hep-/-HFD group showed more severe hepatic insulin resistance and triglyceride deposition than WT HFD group.Meanwhile,in palmitate-induced primary hepatocytes,SphK2^Hep-/-mice also showed more serious lipid toxicity,including hepatocytes apoptosis and lipid accumulation.PPAR?-CD36 signaling pathway was activated in SphK2^Hep-/-mice liver further aggravating lipid accumulation in the liver.Conclusion:Hepatocytes SphK2 is involved in hepatic lipid metabolism induced by HFD in mice and affects the pathological process of NAFLD.The mechanism is mainly about interposing the uptake of free fatty acids in hepatocytes by regulating the PPAR?-CD36 signaling pathway,thereby effecting the liver deposition of triglyceride and further affecting the development of liver insulin resistance and NAFLD.This study indicated that SphK2 is expected to be a new target for the research of NAFLD.Part 2Scope: Non-alcoholic steatohepatitis?NASH?is characterized by lipid accumulation in hepatocytes and inflammatory cell infiltration.In view of the anti-oxidative?inoxidizability and anti-inflammatory effects of polydatin,the current study aimed to investigate the pharmacological effects of polydatin on NASH and its related fibrosis.Methods: C57BL/6 mice were fed with methionine-choline deficient?MCD?diet to induce NASH and liver fibrosis,and treated with or without polydatin?5 mg/kg/day,i.p?for 4 weeks.Hep G2 cells induced by palmitic acid?PA?were treated with polydatin.Results: The elevations of serum alanine aminotransferase?ALT?and aspartate aminotransferase?AST?,active caspase-3,TUNEL-positive cells,and triglyceride content were decreased by polydatin treatment.In addition,administration of polydatin to MCD-fed mice reduced oxidative stress by down-regulating NOX4 enzymes.Furthermore,the reduction in inflammation and CD68 macrophage activation correlated with inhibition of Toll-like receptor?TLR?-4/NF-?B p65 signaling pathway by polydatin treatment.Polydatin also attenuated lipid accumulation,inflammation and apoptosis in Hep G2 cells challenged by palmitic acid?PA?combined with or without lipopolysaccharide?LPS?.Finally,the reduction of hepatic fibrosis by polydatin treatmentcorresponded to a reduction in hepatic gene expression of fibrosis markers.Conclusion: These results suggest that polydatin prevents NASH and fibrosis via inhibition of oxidative stress and inflammation,highlighting polydatin as a potential therapeutic agent for prevention and treatment of NASH.