Effects Of Aerobic Exercise On Hepatic Ceramide Metabolism And Endoplasmic Reticulum Stress In NAFLD Mice

Objective:Aerobic exercise is an effective therapeutic strategy for NAFLD,while the underlying mechanism remains unclear.dyslipidemias and endoplasmic reticulum stress are important pathological mechanisms of NAFLD.The Large amount of lipid deposition in liver is the main feature of NAFLD,which also drives the development of NAFLD.Previous studies on liver lipids in NAFLD mainly focused on triglycerides,diglycerides and cholesterol.Subsequent studies found that ceramide,as an important lipid regulating cellular stress and death,was elevated in liver of NAFLD.The effects of aerobic exercise on hepatic ceramide metabolism and the role of ceramide in the improvement of NAFLD by aerobic exercise remain unclear.The purpose of this study was to investigate whether aerobic exercise could regulate hepatic ceramide metabolism and inhibit ER stress,thereby improving NAFLD.The research mainly focuses on the following two aspects: the effect of aerobic exercise on liver ceramide metabolism,the effect of aerobic exercise and inhibition of ceramide synthesis on liver endoplasmic reticulum stress and NAFLD.Methods:C57BL/6J male mice were randomly divided into five groups after one week of adaptation: low-fat diet group（LFD）,high-fat diet group（HFD）,high-fat diet plus exercise group（HE）,high-fat diet plus injection phosphate buffer group（HP）,high-fat diet plus injection myriocin group（ceramide de novo synthesis inhibitor）（HM）.After the intervention,serum triglyceride（TG）,total cholesterol（TC）,high density lipoprotein cholesterol（HDL-C）,low density lipoprotein cholesterol（LDL-C）and liver enzyme alanine aminotransferase（ALT）were detected the level of aspartate aminotransferase（AST）was measured,the pathological changes of liver were observed,and the contents of TG and TC were measured.The contents of hepatic ceramide C14:0,C16:0,C18:0,C18:1,C20:0,C22:0,C24:0 and C24:1 were detected by LC-MS/MS,The m RNA levels of liver ceramide related metabolic enzymes（SPTLC1,SPTLC2,Cer S1,Cer S2,Cer S4,Cer S5 and Cer S6）were detected by q PCR,and the liver endoplasmic reticulum stress related proteins expression levels were detected by Western blots: glucose regulated protein 78（GRP78）,C/EBP homologous protein（CHOP）,Phospho-Protein kinase-like endoplasmic reticulum kinase（p-PERK）,Protein kinase-like endoplasmic reticulum kinase（PERK）,Phospho-eukaryotic translation-initiation factor 2α（p-e IF2α）and eukaryotic translation-initiation factor2α（e IF2α）.Results:1.NAFLD is alleviated by aerobic exercise and ceramide synthesis inhibitors:Compared with LFD group,the contents of serum TG,TC,HDL-C,LDL-C and the levels of AST and ALT in HFD group were significantly increased.The results of liver pathological staining showed that liver lipid droplets increased significantly,there was obvious steatosis,and the contents of TG and TC in liver increased significantly;Aerobic exercise intervention can effectively reduce the levels of serum TG,TC,LDLC,AST and ALT,and significantly reduce the contents of liver TG and TC.The results of liver pathological staining showed that liver lipid droplets were significantly reduced;Inhibition of ceramide synthesis by myriocin significantly reduced the levels of serum TC,HDL-C,ALT,AST,and the levels of liver TG and TC.The results of liver pathological staining showed that liver lipid droplets were significantly reduced.2.Aerobic exercise and ceramide synthesis inhibitors can improve the hepatic ceramide metabolism disorder: compared with LFD group,the level of total ceramide in liver of HFD group was significantly increased,and the changes of ceramide content were as follows: high fat diet significantly increased the content of C14:0,C16:0,C18:0,C18:1,C20:0 and C22:0 ceramide,and decreased the content of C24:0 ceramide;Aerobic exercise can effectively reduce the level of liver total ceramide in mice,and the changes of ceramide content are as follows: aerobic exercise can significantly reduce the content of C14:0,C18:0,C18:1 and C20:0 ceramide and increase C24:0ceramide;myriocin injection could significantly reduce the level of liver total ceramide in mice.Except that the content of C24:1 and C18:1 ceramide had no significant change,the content of other ceramide decreased significantly.3.Aerobic exercise can regulate the liver ceramide metabolism: compared with LFD group,the changes of ceramide related metabolizing enzyme m RNA in HFD group were as follows: the levels of SPTLC1,Cer S1,Cer S4,Cer S5 and Cer S6 m RNA increased significantly,and the level of Cer S2 m RNA decreased significantly;Aerobic exercise can significantly reduce the levels of SPTLC1,Cer S1 and Cer S6 m RNA in liver and increase the level of Cer S2 m RNA.4.Aerobic exercise and inhibition of hepatic ceramide synthesis can improve hepatic endoplasmic reticulum stress: Compared with LFD group,the expression levels of ER stress-related proteins p-PERK,PERK,p-e IF2α,e IF2α,CHOP and GRP78 were significantly increased in HFD group.Aerobic exercise significantly reduced the protein expression levels of p-PERK,p-e IF2α,e IF2α,CHOP,GRP78 and p-PERK/PERK ratio in liver.The expression levels of p-PERK,PERK,p-e IF2α,e IF2α and CHOP protein in the liver of mice were significantly decreased by myriocin injection Conclusions:（1）Both aerobic exercise and inhibition of ceramide synthesis can reduce ER stress,change ceramide ratio and relieve NAFLD.（2）The improvement of NAFLD by aerobic exercise was related to the improvement of hepatic ceramide metabolism disorder and the reduction of endoplasmic reticulum stress.