Effect Of Gegen Qinlian Decoction On ACC1,FASN,SCD1 And LPL In Adipose Tissue Of Rats With Insulin Resistance

ObjectiveThis study aims to explore the effects of Gegen Qinlian Decoction（GQD）,a traditional Chinese medicine compound,on the adipose tissue ACC1（Acetyl Co A carboxylase 1）,FASN（Fatty acid synthase）,SCD1（Stearoyl-Co A desaturase-1）,and LPL（lipoprotein lipase）of rats with insulin resistance induced by high-fat diet.Additionally,the possible mechanism of GQD will be explored from the perspectives of upstream regulatory factors and gene methylation.The research could provide a new view.for studies on the mechanism of Gegen Qinlian Decoction in the prevention and treatment of insulin resistance.MethodsMale SD rats with insulin resistance model were fed high-fat diets that provided 60%of energy（kcal）with fat.The blank control group（Control,C）rats were fed with normal diet for 12weeks.The rats with insulin resistance was screened out according to the insulin resistance index.The insulin resistant rats were randomly divided into Gegen Qinlian Decoction low-dose group（GQDL）,medium-dose group（GQDM）,high-dose group（GQDH）,positive drug rosiglitazone group（Rosiglitazone,RGN）,simvastatin group（Simvastatin,SVT）and insulin resistance model group（Insulin Resistance,IR）.Oral administration or water for 16 weeks.The dosage of each group is:GQDL is 1.65 g·kg^-1,GQDM is 4.95 g·kg^-1,GQDH is 14.85 g·kg^-1,RGN is 5 mg·kg^-1,and SVT is 10 mg·kg^-1.The C and IR groups were given water 10 m L·kg^-1.After 16 weeks of administration,epididymal adipose tissue and liver tissue were taken.The enzyme activity of adipose tissue ACC1,FASN,LPL were detected by using enzyme activity kit and SCD1,ANGPTL3,ANGPTL4,ANGPTL8 protein level were examined by Westorn Blot.The mRNA levels of Acc1,Fasn,Scd1,Srebp1,Chrebp,Lpl,Angptl4,Angptl8 in adipose tissue and Angptl3,Angptl4 gene in liver were determined by fluorescence quantitative PCR.In addition,the methylation levels of Acc1,Fasn,Scd1 gene fragments were detected by Bisulfite sequencing Polymerase Chain Reaction（BSP）.Finally,the experimental results were analyzed by one-way analysis of variance,independent sample t test and non-parametric test.Results1.Animal experiment results:Compared with rats fed with ordinary diet,the body mass,TG content and the index of insulin resistance in rats fed with high-fat diet were increased significantly（P<0.01）.,but high-density lipoprotein cholesterol（HDL-C）was significant reduced（P<0.01）.Compared with the IR group,the weight gain of rats in the GQDL,GQDM,and GQDH groups was significantly reduced（P<0.01）,the IR index of the rats in the GQDL,GQDM,and RGN groups was significantly reduced（P<0.05）,and the low-density lipoprotein cholesterol（LDL-C）of the rats in the GQDM and GQDH groups was decreased significantly（P<0.01）as well as HDL-C in the GQDH（P<0.01）.2.Results about the effect of GQD on de novo lipogenesis-related enzymes ACC1,FASN,SCD1 in adipose tissue of IR rats:Compared with group C,lots of parameters level were significant decreased in adipose tissue from IR group or GQD group.For example,ACC1 enzyme activities in IR group（P<0.05）,the mRNA expression of Acc1,Fasn,Scd1 Srebp-1,Chrebp gene in IR group（P<0.05）,SCD1 protein content in GQDH group（P<0.05）,and the methylation level of CpG sites of Scd1-1 gene of rats in GQDL,GQDM,and GQDH groups（P<0.05）.Compared with the IR group,levels of many parameters were increased in GQDs,RGN or SVT,such as,FASN enzyme activity in the GQDL,GQDM,and SVT groups（P<0.05）,the Scd1 gene mRNA expression in the RGN group（P<0.05）,the total CpG sites methylation level of Fasn in GQDL and the CpG sites at+506 and+508 methylation level in the GQDM（P<0.01）,and the CpG sites of Scd1 exon+333～+404 fragment in RGN group（P<0.05）.Nevertheless,compared to the IR group,protein of SCD1 in the GQDL,GQDH,and RGN groups was significantly reduced（P<0.05）,and the level of cytosine methylation at non-CpG sites of Acc1 exon of in GQDM group was significantly decreased（P<0.05）.3.Results about the effect of GQD on LPL enzyme activity and LPL inhibitory protein ANGPTL3,4,8 in liver or adipose tissue of IR rats:Compared with group C,levels of many were decreased significantly in IR,GQDs,RGN or SVT.For example,the Lpl mRNA in adipose tissue of IR,GQDL,GQDM,GQDH,RGN,SVT group were significantly decreased（P<0.05）,furthermore,Angptl3 mRNA of liver in the GQDL,GQDM,GQDH,RGN groups（P<0.05）and Angptl4 mRNA of liver in the GQDL,GQDM,RGN,and SVT groups（P<0.05）were reduced obviously.However,the expression of ANGPTL8 protein in liver tissue of RGN group increased significantly（P<0.05）.Compared with the IR group,the expression of Angptl3 in the liver of the GQDL,GQDM,GQDH,and RGN groups decreased significantly（P<0.05）,and the expression of Angptl4 in the liver of the RGN group decreased significantly（P<0.05）.Conclusion1.GQD can effectively reverse overweight and insulin resistance caused by high-fat diets.2.GQD can improve the reduction of ACC and FASN enzyme activities in epididymal adipose tissue caused by high-fat diets to a certain extent,and its mechanism may be related to the regulation of Acc1 and Fasn methylation.The+506 and+508 CpG sites of Fasn gene may be the main targets of GQD.3.The high-fat diet significantly reduces the mRNA levels of Acc1,Fasn and Scd1,which may be closely related to the decrease in the expression of Srebp1 and Chrebp;GQD cannot effectively reverse the decrease in the mRNA levels of Acc1,Fasn,Scd1,Srebp1 and Chrebp caused by the high-fat diets.Therefore,GQD may not play a role by improving the transcription of DNL-related genes.4.GQD may not play a role in regulating blood TG by acting on ANGPTLs-LPL in epididymal adipose tissue.