The Empirical Study Of Pinocembrin On Atherosclerosis In Apoe^-/- Mice

ObjectiveTo study the depressant mechanism of pinocembrin on apolipoprotein E gene knockout (apolipoproteinE-defieient, ApoE-/-) mice with atherosclerosis (Atherosclerosis, AS) and to attempt to provide the theory for its anti-AS effect.MethodsThe pinocembrin identified by the means of melting point determination and mass spectrometry was purified by the methods of silica gel column chromatography and semi-preparative liquid chromatography separation. Using 6-8 weeks old ApoE-/- mice, a model of atherosclerosis was established.40 mice were randomly divided into 5 groups, n=8. Groups and the treatment for each group were as follows:①control group(CGgroup):honey 5mLkg-1.d-1;②simvas-tatin group (SMgroup):simvastatin dissolved with honey 10mg.kg-1. d-1;③low-dose pinocembrin group (LPgroup):pinocembrin dissolved with honey 10 mg.kg-1.d-1;④high-dose pinocembrin group(HPgroup):pinocembrin dissolved with honey20 mg.kg-1.d-1;⑤combination of simvastatin and high-dose pinocembrin group (SPgroup):pinocembrin 20mg.Kg-1.d-1 and simvastatin 10mg.kg-1.d-1 dissolved with honey. After 14 weeks, blood samples were collected and centrifuged to obtain serum. Using automatic biochemical analyzer, serum triglycerides (TG), cholesterol (TC), high density lipoprotein (HDL), low density lipoprotein (LDL) and very low density lipoprotein (VLDL) concentration were detected; Nitrate reductase assay was used to detect the serum concentration of nitric oxide (NO) and nitric oxide synthase (NOS); Serum endothelin(ET)and vascular endothelial growth factor(VEGF) concentration were detected by the means of ELISA assay; Atherosclerotic plaque in the root of aorta was observed by the means of oil red O staining Aorta from aortic arch to the iliac artery bifurcation was stained by the means of oil red O staining, the size of atherosclerotic plaque was analyzed by the image analysis system; The expression of VEGF in mice aorta AS plaque was determined by the means of immunohistochemistry.Results①The concentration of serum TG and LDL of SM group, LP and HP group decreased compared with the CG group, the differences were statistically significant (P<0.05);②The concentration of serum NO of SM group,HPgroup,LPand the CG group decreased in turn, the differences between each other were statistically significant compared with the CG group; The difference of serum concentration of NO between LPgroup and CG group was not statistically significant (P> 0.05);③The total amount of nitric oxide synthase (TNOS) and inducible nitric oxide synthase (iNOS) of CGgroup were significantly higher than SMgroup and HP group, the difference was statistically significant (P <0.05); the effect of pinocembrin is dose dependant;④Serum concentrations of endothelin and vascular endothelial growth factor in CGgroup were higher than the SM group and the HPgroup, the difference were statistically significant (P <0.05); the effect of pinocembrin is dose dependant;⑤The area of atherosc-lerotic plaque in CG group, SM group and the HP group decreased in turn, the difference between each other was statistically significant (P<0.05); LPgroup and the HPgroup, with the increased dose of pinocembrin, area of atherosclerosis plaque showed a downward trend, the effect of pinocembrin is dose dependant;⑥VEGF expression in atherosclerotic plaque of CG group was significantly increased, VEGF expression in atherosclerotic plaque of CGgroup and SMgroup was significantly reduced,compared with the CG group, the difference was statistically significant (P<0.05); the effect of pinocembrin is dose dependant;⑦The combined treatment of simvastatin and pinocembrin had an interactive effect on blood lipids,NOS,serum ET and VEGF atherosclerotic plaque area of ApoE-/-mouse, the difference was statistically significant (P<0.05), and the interactive effect was synergetic.Conclusion①Pinocembrin could reduce serum concentration of TG, TC, LDL and VLDL,increase serum concentration of NO, and decreased serum concentrations of iNOS and ET, thereby it could reducing vascular tension, having a protect- tive effect on endothelial cells;②Pinocembrin could reduce the area of atherosclerotic plaque and the level of VEGF in atherosclerotic plaque,inhibit the formation of atherosclerotic plaque, to prevent the rupture of plaque and play a stabilizing effect on atheros-clerotic plaque;③Combined with application of simvastatin, pinocembrin could great inhibit the formation of atherosclerotic lesions; Simvastatin and pinocembrin had a synergistic protective effect on heart and vascular.