Study On The Pharmacodynamic Mechanism Of The Effective Parts Of Astragalus Interfering With Diabetic Model Rats

Purpose:This study was designed to evaluate the influence on blood glucose of Astragali Radix active ingredients and the differences of descending blood glucose level in streptozotocin （STZ）–induced diabetes rats. We aimed to analyze the mechanism of astragli radix active ingredients at regulating blood glucose by assaying adiponectin in serum, livers and skeletal muscles and studying mRNA and protein levels of adiponectin receptor1（AdipoR1） and AMP-activated protein kinase（AMPK） in livers and skeletal muscles of diabetic rats.Material and methods:SD rats （n=168） were divided into 12 groups of 14 animals each group. Group1: control rats （C group） treated with nothing; Group2: STZ-induced rats （STZ group） treated with water ig; Group3: STZ-induced rats treated with traditional Chinese medicine （TCM group）; Group4: STZ-induced rats treated with western medicine （WM group）; Group5: STZ-induced rats treated with astragalix radix （AS group）; Group6: STZ-induced rats treated with astragalus flavonolids （ASF group）; Group7: STZ-induced rats treated with astragalus polysaccharides （ASP group）; Group 8: STZ-induced rats treated with astragalosides （ASS group）; Group9: astragalus flavonolids and polysaccharides （ASF+ASP group）; Group10: astragalux flavonolids and astragalosides （ASF+ASS group）; Group11: astragalux polysaccharides and astragalosides （ASP+ASS group）; Group12: STZ-induced rats treated with astragalus flavonolids and polysaccharides and astragalosides （ASF+ASP+ASS group）. Diabetic rats were induced by STZ (52 mg·kg^-1, peritioneal injection, one time) in group2^-12. After STZ treatment （7 day）, blood glucose levels of all rats were measured in group2^-12, and diabetes was confirmed ( Glucose level >16.7 mmol·L^-1). Diabetic rats were irrigated with experimental drugs for 30 days from STZ pj day.The dose of astragali radix and its active ingredients were equal to 6.3 mg·kg^-1astragali radix dried medicinal herbs. The dose of TCM and WM group to the introduction.The model rats were irrigated with distilled water at the level of AS. We measured blood glucose weekly by glucosemeter. We defected blood glucose was tested by glucosemeter. In serum, cholesterol （CHOL） , triglyceride（TG） , low density lipoprotein cholesterin（LDL_C） and high density lipoprotein cholesterol（HDL_C） in serum by automatic biochemistry analyzer. The pancreas tissues were observed using hematoxylin-eosin staining （HE）. We assayed insulin in serum and adiponectin in serum, livers and skeletal muscles by enzyme-linked immunosorbent assay （ELISA）. We examined mRNA levels of AdipoR1 and AMPK in livers and skeletal muscles of diabetic rats by real-time PCR. We also assayed ptotein levels of adipoR1 and AMPK in livers and skeletal muscles by western blotting. The date were analyzed by SPSS sofeware.Results:1 Effect on pharmacodynamics index of astragalus active ingredients in diabetic rat1.1 Effect on blood sugar We hadn’t found statistical differences of blood glucose among group1-12. The 7th day after STZ pj, the blood sugar elevated in STZ group than in control group（P<0.01）.The differences of blood sugar between STZ group and others were obvious. Blood sugar in WM and ASF+ASP group were decreased significantly （P<0.01）, and blood sugar of ASS, ASP+ASS and ASF+ASP+ASS group were also descended distinctly （P<0.05）. The blood sugar in ASF+ASP group was more significantly than other astragalus active ingredients （P<0.05）. Then at the 30th day, blood sugar in STZ- induced diabetes rats was better elevator than control group （P<0.01）. Compared blood sugar of STZ group with others groups, blood sugar were decreased in WM and ASF+ASS and ASF+ASP+ASS groups significantly （P<0.01）. Blood sugar in AS, ASF and ASF+ASP groups were also decreased （P<0.05）.1.2 Effect on blood lipidCHOL, TG, LDL_C and HDL_C in serum were more elevated than control group in STZ-induced diabetes rats （P<0.01）. CHOL and HDL_C in each STZ treatment group was diminished. CHOL was more decreased in AS and ASF groups than effective medicinal control groups （P<0.05）. HDL_C was decreased remarkably in AS, ASF and ASF+ASS groups （P<0.01）. In WM, AS, ASF, ASS, ASF+ASS, ASP+ASS and ASF+ASP+ASS groups, TG level was decreased in serum （P<0.01）. LDL_C level was decreased in ASS group （P<0.05）. 1.3 Effect on pancreas pathological morphologyPancreatic islet was larger in volume, which boundary with the acinar surrounding islet was clear and full of well-distributed pancreatic cells in pancreatic islet. Pancreatic islet had been observed accidentally with shrinked and irregular islet, which boundary wasn’t distinct. Adiministration of islet morphology in each group have improvement at some degrees.2 Effect on pharmacodynamics mechanism of astragalus active ingredient in diabetic rats2.1 Effect on serum insulinInsulin was lower significantly in STZ-induced diabetic rats than control rats （P<0.01）. Insulin was increased in TCM and AS groups compared with STZ-induced diabetic rats（P<0.01）. Between the experimental drugs treatment groups and STZ groups, insulin was elevated more significantly in AS and ASF groups, while insulin in ASP and ASF+ASP+ASS groups were more remarkable than ASF+ASS group（P<0.05）.2.2 Effect on adiponectin in serum, liver and skeletal muscleAdiponectin level in serum, liver and skeletal muscle were decreased significantly in STZ-induced diabetic rats （P<0.01）.Compared with STZ group , all adiponectin in serum, liver and skeletal muscle were ascended in AS , ASF+ASS, ASF+ASP+ASS groups, which adiponectin levels were higher than other treated groups（P<0.01）. In other groups, adiponectin level in serum, livers and skeletal muscles were higher obviously than in ASP group, next to which was ASF+ASP group（P<0.05）.2.3 Effect on mRNA expression of AdipoR1 in liver and skeletal muscleIn liver and skeletal muscle, the mRNA level of adipoR1 was decreased significantly in diabetic rats （P<0.01）. AdipoR1 mRNA expression was elevated obviously in diabetic rats treated with astragalus radix and its every active ingredients （P<0.05）, and level of adipoR1 mRNA expression in liver and skeletal muscle were ascended compared ASF+ASS groups with effective drugs control group（sP<0.05）. Among each experimental medicinal treatment groups, the mRNA levels of AdipoR1 in livers were elevated significantly in AS, ASF, ASF+ASS and ASP+ASS groups（P<0.05）,while the mRNA levels of adipoR1 in skeletal muscles were elevated significantly in ASF+ASP and ASF+ASS groups（P<0.05）. The adipoR1 protein expression in livers and skeletal muscles of diabetic rats was decreased obviously than control rats, while the protein expression of adipoR1 was ascended in AS, ASF, ASF+ASP, ASF+ASS and ASP+ASS groups. 2.4 Effect on mRNA expression of AMPK in liver and skeletal muscle In liver and skeletal muscles, the mRNA levels of AMPK was decreased significantly in diabetic rats （P<0.01）. Compared with STZ group, all of AS, ASF, ASF+ASP and ASF+ASP+ASS elevated AMPK m RNA expression in liver and skeletal muscle of diabetic rat （P<0.05）, and ASS and ASF+ASS were also elevated AMPK mRNA expression in skeletal musclse. （P<0.05）. Among each experimental drugs treatment groups, the AMPK mRNA levels in livers were elevated significantly in AS, ASF, ASF+ ASP+ASS groups（P<0.05）,while the mRNA levels of adipoR1 in skeletal muscles were elevated significantly in AS, ASS and ASF+ASS groups（P<0.05）.The AMPK protein expression in liver and skeletal muscle of diabetic rats was decreased obviously than control rats, while the protein expression of AMPK was ascended in AS, ASF, ASF+ASP, ASF+ASS and ASP+ASS groups.Conclusion:1 The results indicate that astragalus radix and ite active ingredients could reduce blood sugar of diabetic rats in varying degrees. Targets of their action have different emphasis.2 The results indicate that astragalus radix active ingredients ASF and ASF+ASS could reduce blood sugar, blood lipid .We propose that the mechanisms of ASF and ASF+ASS regulating blood sugar of diabetic rats may be related to adiponectin activate AMPK pathway.3 The study suggests that astragalus flavonolids may be the best channel to apply diabetic rats in each astragalus active ingredients .