| Objective:Previous studies have found that breviscapine can improve type 2 diabetes and hyperlipidemia significantly. This experiment fed rats with high-fat and high-sugar to copy insulin resistance(IR) model. By observing the fasting blood glucose(FBG), the fasting serum insulin(FINS) and the insulin resistance index(IRI), we can evaluate whether the model is established. After using breviscapine for 2 weeks, then we detected the content of free fatty acid(FFA), triglyceride(TG) in serum and skeletal muscle.Observe and determine the variability of lipid deposition in skeletal muscle. Besides, by measuring the levels of long chain fatty acyl-coenzyme A(Lcco As) in skeletal muscle,the difference of fatty acid transposition(FAT/CD36) and the carnitine palmityl transferase-1(CPT-1) in protein and Mrna, we can investigate the improvement and mechanism of breviscapine on IR, lipid deposition and fatty acid metabolism.Methods:55 healthy adult male SD rats were adaptably fed for 7 days, and after fasting 12 h their FBG were measured. Choose 50 rats with normal FBG, and they were randomly divided into the normal control group(10 rats) and the high-carbohydrate group(40 rats),and they were always fed with the normal diets and the high-fat, high-carbohydrate diets respectively. After 12 weeks, all rats were tested glucose tolerance by intraperitric(50%C6H12O6, 2g/kg). 30 rats were screened, and they were randomly divided into the model group, the dose of breviscapine group(200mg/kg,ip) and the low dose of breviscapine group(100mg/kg,ip). With 10 rats in each group, these rats were respectively given drugs for 2 weeks. The normal group and the model group received the same dose of physiological saline. At the end of the 14 th week, stop feeding for 12 hours and the FBG of rats were tested and anesthetized with 10% chloral hyclrate. Take the blood from the abdominal aorta of the rats for 15 ml, remove the adipose tissue of the epididymis and the kidney and acquire the quadriceps femoris tissue. Separate the serum with a centrifuge, detect the FINS of serum and evaluate the insulin resistance index.Determine the changes of TG and FFA in serum and tissue. Stain the frozen section with Oil red O and observe the lipid deposition of skeletal muscle. Detect the content of serum Lcco As with enzyme linked immunosorbent assay. Observe the expression level of FAT/CD36 and CPT-1 in quadriceps muscle with Western blott. Detect the difference of FAT/CD36 and CPT-1 in each group with conventional PCR.Results:(1)The general situation of rats: After 14 weeks, the body weight, abdominal fat index, FBG, FINS, IRI in the model group were significantly higher than the normal group(P<0.01). The levels of high dose of breviscapine group and low dose of breviscapine group were lower than the model group(P<0.01) and the high dose of breviscapine group reduced more significantly(P<0.01). It indicates that the high-fat and high-sugar diets led to hyperglycemia and hyperisulinema and resulted in IR.Breviscapine can reduce FBG and FINS and improve IR obviously.(2)Oil red O staining results: a lot of lipid deposition of the model group rats appeared significantly(P<0.01); the situations of the high and low dose of breviscapine groups improved differently(P<0.01), and the improvement of the high dose of breviscapine was more significant(P<0.01). It indicates that the breviscapine can improve lipid deposition. The high-fat and high-sugar diets increased the fat of rats in skeletal muscle and the lipid deposition appeared significantly. The symptoms improved after the treatment of Breviscapine for 2 weeks.(3)The results of FFA and TG in tissue and serum: The content of FFA and TG in serum and skeletal muscle increased significantly(P<0.01); compared with the model group, the content in the low dose of breviscapine group FFA reduced significantly(P<0.01); compared with the low dose group, TG in the high dose group decreased significantly(P <0.01). After feeding rats for 14 weeks, TG and FFA increased obviously;breviscapine can weaken hyperlipidemia significantly.(4)The results of Lcco As in skeletal muscle: Compared with the normal group, the level of Lcco As in the model group decreased more significantly(P<0.01); after medication, compared with the model group, the level of Lcco As in the high and low dose group of rats increased more significantly(P<0.01). Lcco As is the active form of long chain fatty acids in cell. IR rats, the content of Lcco As increased, which indiated that long chain fatty acids intake cell raised. breviscapine can improve the activity of Lcco As to inhibitor intake.(5)The protein and m RNA of FAT/CD36: As for FAT/CD36 in quadriceps, the expression of the model group increased more than the normal group(P<0.01); compared with the model group, the high and low dose groups had different changes(P<0.05), and the level of the high dose group changed most obviously(P<0.05). FAT/CD36 helped fatty acids enter into cells. The content of FAT/CD36 in rats raised, which indicated that the level of fatty acids increased and breviscapine can inhibit expressing.(6)The protein and m RNA of CPT-1: Compared with the normal group, the content of CPT-1 protein and m RNA decreased more significantly(P<0.01); compared with the model group, the content of CPT-1 and m RNA(P<0.05) both increased more in the high and low dose groups. CPT-1 is a key enzyme in mitochondrin used to regulate fatty acids oxidation. The protein and m RNA in IR rats decreased significantly and the fatty acids oxidation decreased. The treating of breviscapine helped to increase the expression of CPT-1 and the fatty acids oxidation increased so that the disorder of glucose and lipid metabolism can be improved.Conclusions:(1)High-fat and high-sugar diet can replicate insulin resistance rats model.(2)FFA and TG in serum and skeletal muscle of IR rats increased significantly and lipid accumulation appeared obviously in quadriceps.(3)In the IR rats model, the content of Lcco As and FAT/CD36 associated with fatty acid intake raised, and the expression of CPT-1 linked to oxidation decreased.(4)Breviscapine can reduce FBG and FINS to improve IR and make TG and FFA reduce to improve skeletal muscle lipid accumulation. The mechanism may be related to the reduction of Lcco As and FAT/CD36 and the rising of CPT-1. |
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