Role Of LOX-1 Pathway In Polyunsaturated Fatty Acids Alleviate Steatohepatitis Injury

The concern about the non-alcoholic fatty liver disease(NAFLD)is increasingly popular since its incidence is increasing and younger.Epidemiological studies have demonstrated consumption of fish oil or perilla oil rich in ω-3 polyunsaturated fatty acids(PUFAs)protected against NAFLD progression and ameliorated hepatic damage and inflmmation,but the mechanisms remained unclear.As characterization of metabolic syndrome in the liver,NAFLD is closely related to diabetes and cardiovascular disease.A lot of studies have shown that oxidized LDL receptor-1(LOX-1)mediated atherosclerosis,the pathological process of diabetes,kidney inflammation and fibrosis,however,LOX-1 in the NAFLD have not yet been reported.Object:Establish the NAFLD animal model by feeding high fat and cholesterol and the damaged L02 liver cells by high saturated fatty acid(PA).To explore the mechanism of ω-3 PUFA in relieving the endoplasmic reticulum stress(ERS)and in improving NAFLD.To know better about the Role of LOX-1 in polyunsaturated fatty acids alleviate non-alcoholic fatty liver disease.Methods:192 male adult SD rats were divided into 4 groups randomly,each group have 48s:the control group(CON)was feeded normal diet,NAFLD model group(HFD)was feeded high-fat diet(20% fat content+2% cholesterol),the fish oil intervention group(F)was feeded high-fat+fish oil(30% of DHA and EPA)diet,the perilla oil intervention group(P)was feeded high-fat+perilla oil(60% of ALA)diet,each group was divided into 4 groups(4,8,12,16weeks)(n=12 in each subgroups).The rats were beheaded at the 4th,8th,12 th,16th week to collect blood,achieve body and liver weight,and detect the changes of rat serum(ALT,AST),blood lipid(TC,TG,HDL-C,LDL–C),SOD,GST and FFA.Rat liver were embedded in paraffin for HE and Masson staining.The expression of LOX-1 were detected by RT-PCR and Western Blot.The expression of marker genes(BIP,I/RE1,ATF6,ATF4,XBP-1)and marker protein in endoplasmic reticulum stress were detected by RT-PCR and Western Blot respectively.The expression of marker genes(IL-1β,TNFα)and marker protein in inflammatory reaction were detected by RT-PCR and Western Blot respectively.The expression of p-JNK、c-Jun、c-Fos were detected by Western Blot.Incubate the L02 cells with different PA concentrations and test its proliferative activity by MTT assayto getan optimal concentration.Set the normal,PA,PA + DHA,groups.The expression of LOX-1 were detected by RT-PCR and Western Blot.The expression of marker genes(BIP,IRE1,ATF6,ATF4,XBP-1)and marker protein in endoplasmic reticulum stress were detected by RT-PCR and Western Blot respectively.The expression of marker genes(IL-1β,TNFα)and marker protein in inflammatory reaction were detected by RT-PCR and Western Blot respectively.The expression of p-JNK、c-Jun、c-Fos were detected by Western Blot.Results:Compared with the CON group,the liver index and the level of serum ALT,TCH,LDL-C,FFA and GST increased significantly in HFD group,And the liver morphology changes,lipid accumulation and inflammatory infiltration could be easily observed on HFD liver cells,which means NASH had happened.Compared with the HFD group,liver index in FOH and POH groups decreased significantly,and serum lipid indexes and liver enzyme activity were significantly decreased.The syndrome of hepatocyte ballooning,steatosis,inflammatory cells infiltration and fibrosis had been improved.RT-PCR and Western Blot test results showed the LOX-1,BIP,IRE1,ATF6,ATF4,XBP-1,IL-1β,TNFα,p-JNK,c-Jun,c-Fos expression in the HFD group were higher than the normal group obviously,and Perilla oil and fish oil can reduce the gene and protein expression significantly.The best PA incubating concentration was 200μM.RT-PCR and Western Blot test results showed the LOX-1,BIP,IRE1,ATF6,ATF4,XBP-1,IL-1β,TNFα,p-JNK,c-Jun,c-Fos expression in the PA group were higher than the normal group obviously,and DHA can reduce the gene and protein expression significantly.Conclusion:The NAFLD rat model established by high fat and cholesterol is successfully established,fish oil and Perilla oil had an active effect in preventing and treating NAFLD,such as reducing blood lipid,improving liver function,reduce the relieving the endoplasmic reticulum stress,relieves the degree of inflammatory reaction.Our results indicate that activation of ER stress and inflammatory reaction is involved in PA-induced LOX-1 upregulation in L02,and that DHA inhibit LOX-1 induction through suppression of ER stress.