The Effect And Mechanism Of Emblic Leafflower Fruit On Vascular Dysfunction In Hyperglycemia Rats

Object: To Study the effect and mechanism of traditional Chinese medicine emblic leafflower fruit(ELF)on vascular dysfunction of hyperglycemia rat and reveal its potential material basis for further clinical application.Methods: 1.The effect and mechanism of ELF on vascular dysfunction of hyperglycemia rat.1.1 The effect of ELF on vascular dysfunction of hyperglycemia rat Fifty male SD rats were randomly divided into two groups after one week of adaptive feeding: the experimental group with 42 rats and the control group with 8 rats.The rats were fasted but with water for 12 h,the experimental group was induced by a single dose of STZ(60 mg/kg,i.v.)injection as previously described,and the normal control rats(NCR)was injected with an equal volume of sodium citrate buffer solution.The development of hyperglycemia in rats was identified by the plasma glucose assessment of blood samples collected from the tail vein 72 h after STZ exposure.STZ-injected animals with blood glucose levels > 15.0 m M were considered diabetic.Blood glucose values were collected every week.Four days after STZ administration,the successful rats were randomly divided into four groups(1)hyperglycemic rats(HGR);(2)hyperglycemic rats treated with low dose of ELF(LD,ELF 25 g/kg);(3)hyperglycemic rats treated with mediandose(MD,ELF 50 g/kg);(4)hyperglycemic rats treated with high dose(HD,ELF 75 g/kg).The ELF consumption in rats was through the juice intragastric administration once a day for five weeks.After five weeks,rats were weighed,and glucose values in rats were measured.Then diabetic rats were anesthetized with 0.3 % pentobarbital sodium(30 mg/kg)and sacrificed by decapitation.The serum was obtained from the whole blood by centrifugation(12000 ×g,5 min,4 ?).The supernatant was used for total nitric oxide(NO)assessment.The aorta was assessed by isometric tension vasomotor and residual aorta tissue was preserved-80 ?.1.2 The mechanism of ELF on vascular dysfunction of hyperglycemia rat Real-Time q-PCR was used to detect the effect of ELF on the m RNA expression of c-Myc and Cyclin D1 in the thoracic aorta of hyperglycemic rats.,and Western Blotting was used to detect the effect of ELF on the protein expression of P-Akt(308)in the thoracic aorta of hyperglycemic rats.2.Analysis of metabolites of ELF in rats Fasted for 12 hours before the last administration,then collected 0-12 h urine,preserved at-80 ?.HPLC-MS/MS method was used to analyze the metabolites of ELF in rats.3.The effect and mechanism of Urolithin A(Uro A)on proliferation of A7r5 cells induced by high glucose The CCK8 assay was used to evaluate the effect of different concentrations of Uro A on the proliferation of A7r5 cells induced by high glucose;Real-Time q-PCR and Western Blotting were applied to research the mechanism of Uro A on the proliferation of A7r5 cells induced by high glucose.For studying the effects of Uro A on Wnt signaling,the Topflash/Fopflash luciferase assay and Western Blotting were used to detect the effect of Uro A on Wnt/?-catenin pathway in A7r5 cells induced by Licl.Result: 1.(1)ELF can increase the body weight of hyperglycemic rats and decrease the fasting blood glucose of hyperglycemia rats,but there was no significant difference.(2)ELF can reduce the postprandial blood glucose of hyperglycemic rats(P<0.05,P<0.01).(3)ELF can elevate the content of serum NO and improve endotheliumdependent vasodilation of aortic ring of hyperglycemic rats(P<0.05,P<0.01).(4)ELF inhibited the protein expression of P-Akt(308)and the m RNA expression of c-Myc and Cyclin D1 in the arterial tissue(P<0.05,P<0.01).2.Uro A is one of the metabolites of ELF in rats 3.(1)Uro A inhibited the abnormal proliferation and suppressed the protein expression of P-Akt(308)and ?-catenin,down-regulared the m RNA expression of c-Myc and Cyclin D1 of A7r5 cells induced by high glucose.(2)Uro A Ameliorates Li Cl-induced activation of Wnt/?-Catenin Signaling in A7r5 cells.Conclusions: Emblic Leafflower Fruits ameliorate vascular smooth muscle Cell Dysfunction by Akt/?-catenin in Hyperglycemia: an underlying mechanism involved in ellagitannin metabolite UroA.