Effects Of Fenofibrate And Pioglitazone On Vaspin Expression In 3T3-L1 Adipocytes And High-fat Diet Induced Obese Rats

Objective To investigate the effects of fenofibrate and pioglitazone on vaspin expression in 3T3-L1 adipocytes and high-fat diet induced SD obese rats.Methods(1) Preadipocytes were differentiated by MIX,dexamethasone and insulin, and turn into mature adipocytes after 8 days.During the differentiation,3T3-L1 cells were incubated with pioglitazone or fenofibrate in the concentration of 100μmol/L for 2, 4,6 and 8 days.Total RNA or protein was extracted from the adipocytes in various time of the differentiation,and the supernatant of adipocytes was harvested.Every condition was repeated three times.(2) 80 7-week-old SD male rats were randomly divided into four groups.They were normal control group(NC),high-fat diet group(HF), fenofibrate-treated group(F) and pioglitazone-treated group(P),20 rats in each group. Normal control group rats were fed with standard food and the others with high-fat diet. After 12 weeks,models of obese rats were established.Then fenofibrate and pioglitazone groups were treated with 30mg/kg fenofibrate,10mg/kg pioglitazone intragastric daily for 4 weeks respectively,and HF and NC groups with homologous solvent.The volumn of drug or solvent was 2ml/kg daily.Body weight was measured regularly.Fasting blood glucose(FBG),fasting insulin(FINS),triglyceride(TG),total cholesterol(TC),free fatty acids(FFA) were detected after 12 weeks and 16 weeks. Last epididymal adipose tissue were weighed,and harvested along with subcutaneous adipose tissue for protein extraction.(3) Semi-quantitative RT-PCR and western blot analysis were used to detect vaspin expression in the adipocytes,the supematant,the adipose tissue and sera of rats. Results(1) Vaspin mRNA and protein were expressed during the process of differentiation.Vaspin mRNA elevated to the peak at day 4(P＜0.05),but there was no further development at day 6 and day 8(P＞0.05).There were no statistical significance in vaspin protein among them during the entire period of differentiation(all P＜0.05).(2) Fenofibrate up-regulated vaspin mRNA at the 2,4,6 and 8 days of the differentiation by 6.3%,19.2%,26.9%,33.9%.Accordingly,pioglitazone increased by 5.1%,10.7%, 16.5%,20.9%.All of them were higher than control adipocytes(all P＜0.05).The levels of vaspin mRNA in fenofibrate treated adipocytes were higher than those of pioglitazone at the 4,6,8 clays of differentiation(all P＜0.05).With time passed,the effects of drugs were changed stronger.(3) Fenofibrate increased vaspin protein expression by 7.7%,20.8%,35.6%,56.5%at the 2,4,6 and 8 days of differentiation, and pioglitazone increased by 5.5%,10.9%,20.3%,39.1%.The protein levels of vaspin in the supernatant were in accord with the corresponding adipocytes.Fenofibrate increased vaspin protein expression more than pioglitazone significantly in 3T3-L1 cells and the supernatant at the 4,6,8 days of differentiation(all P＜0.05).(4) Compared with NC group,body weight,Lee's index,weight of epididymal adipose tissue,TG,FFA, FBG,and FINS in rats of HF group elevated significantly(all P＜0.05).Compared with HF group,TG,FFA,FINS decreased markedly in fenofibrate and pioglitazone treated rats(all P＜0.05).Body weight,Lee's index,weight of epididymal adipose tissue and fasting plasma glucose of fenofibrate-treated rats reduced obviously(all P＜0.05),but there was no statistical difference in these index between pioglitazone and HF groups (P＞0.05).The levels of TC were similar among all groups(P＞0.05).(5)Vaspin protein expression in epididymal adipose tissue were higher than those in subcutaneous fat (P＜0.05).High-fat diet decreased vaspin by 14.3%in epididymal fat(P＜0.05),but there was no change in subcutaneous vaspin level(P＞0.05).Compared with NC group and HF group,both epididymal and subcutaneous vaspin protein were increased by fenofibrate treatment statistically(P＜0.05).Pioglitazone elevated vaspin in epididymal adipose tissue,but inhibited the expression of vaspin in subcutaneous fat,which were lower than the level of vaspin in NC group(all P＜0.05).(6) Vaspin was detected in sera as two bands of a 45kDa and a 90kDa.High-fat diet markedly reduced the amount of 45kDa plus 90kDa by 15.7%(P＜0.05).Fenofibrate and pioglitazone reversed them to normal levels(all P＞0.05).Conclusion:(1) The peak of vaspin mRNA is observed at day 4 of differentiation.The expression of vaspin protein doesn't change during the differentiation.Fenofibrate and pioglitazone can up-regulate vaspin mRNA and protein expression in 3T3-L1 cells.(2) High-fat diet can induce obesity in rats and metabolic disturbance of glucose and lipid. Fenofibrate and pioglitazone can reduce lipid profiles and increase insulin sensitivity. Meanwhile,fenofibrate can reduce adiposity.(3) Vaspin is downregulated in sera and epididymal adipose tissue of high-fat diet rats.Fenofibrate and pioglitazone can reverse them to normal levels.Simultaneously,Fenofibrate can increase vaspin in subcutaneous adipose tissue,but the effect of pioglitazone is contrary.The structure and function of the different molecular weight vaspin in sera await further investigation.