Experimental Study Of Astragalus Polysaccharide Modified-collagen Type I On Angiogenesis

Background:There are many theories such as blood related with QI, therefore, the Chinese medicine which has the role of qi and blood may have a role in promoting angiogenesis. This angiogenesis effect of Chinese herbal medicine has laid a theoretical foundation. In Traditional Chinese medicine, the astragalus was thinked as the "magic medicine in treated sore", with the effect of tonifying Qi, hemogenesis and alleviate water retention.A large number of clinical applications and prescriptions show that astragalus promote tissue healing; and the promote angiogenesis effect of astragalus was discovred in several experiments. Our previous study also found that astragalus would promote angiogenesis, and the mechanism of which may be related to the promotion of vascular endothelial growth factor and other growth factors that increase the content of cell hemoglobin and hydroxyproline acid content. This project will study the angiogenesis function using the main constituents of astragalus APS load type 1 collagen in vitro, in Chick chorioallantoic membrane model and rat damage model, and discuss its mechanisms. This experiment is divided into two parts.Part I:The Synergy Effects of Astragalus Polysaccharide and Collagen I to Promote Angiogenesis on cellular levelObject:To observe the promote angiogenesis effect of the astragalus polysaccharide and collagen type I, and to discuss the synergistic effect between the astragalus polysaccharide and collagen. Methods:Firstly, to determinate the proliferation effect on human umbilical vein endothelial cells (HUVEC) of different concentrations of astragalus polysaccharides and collagen I; secondly, to survery the cell migration rate by experiment of cell migration. Then, to verificate the expression of angiopoitin I, integrinαvβ3 and VEGF. All the data were showed as mean±standard deviation and were statistical analysis of one-way ANOVA by SPSS 11.0 statistical software. P<0.05 was considered to has statistically significant. Results:The most significant effect of promote cell growth of astragalus polysaccharide was determined on the concentration of 20ug/ml.the role of promoting growth was gradually decreased as the concentration of astragalus polysaccharide increased.while the synergistic effect was discovered when combination of astragalus and collagen I, the promoting growth effect was significantly higher than 20ug/ml collagen group.(P=0.0139).The value of OD and migration rate of HUVEC was significantly higher than other groups(P<0.05).The expression of angiopoietin I.VEGF and integrinαvβ3 was increased as the increasing of the concentration of astragalus polysaccharide and collagen I, The peak value was discovered when the 20ug/ml astragalus and 20ug/ml collagen I was combinated. Conclusions:Astragalus polysaccharide can promote HUVEC proliferation and migration, and increases the expression of angiopoietin-Ⅰ, VEGF and the Integrinαvβ3.The results suggested the role of angiogenesis by astragalus polysaccharide's treatment. And the synergistic effect was discovered when the astragalus polysaccharide and collagen I was combinated.Part II:Experimental Study of Astragalus Polysaccharide Modified-collagen on AngiogenesisObject:To study the promote angiogenesis effect of Astragalus polysaccharide modified-collagen on CAM and SD rat model respectively, and approach its mechanism of synergistic effect preliminarily.Methods:Firstly, the type-I collagen was modified by astragalus polysaccharide,then the effect of promote angiogenesis was observed in the chick embryo chorioallantoic membrane (chick chorioallantoic membrance, CAM) model and damage model of SD rats.Then the new microvessel density, content of hemoglobin, prolyl hydroxyl acid content in granulation tissue and CD34 antigen staining were used to verify its role in promoting angiogenesis, and its optimal dose were observed. The expression of VEGF and of integrinαvβ3 protein in CAM was verified by Western blot to explore the role of promoting angiogenesis mechanism.The expression of VEGF and intrgrin in the granulation tissue was verified by RT-PCR. All data were showed as mean±standard deviation (±s),and was analysised by SPSS 11.0 statistical software for statistical analysis of one-way ANOVA.P<0.05 was thinked to be has statistically significant.Results:In the chick embryo CAM model, the promote angiogenesis effect of the 20ug/ml collagen group of astragalus polysaccharide+collagen was significantly higher than other groups (p<0.05), with statistical significance,and the 20ug/ml was the best dose. The content of hemoglobin and hydroxyproline in the SD rat injury model in rats wound collagen content and granulation tissue was significantly higher (p<0.01)than the group of collagen only; CD34 positive cells was observed more intensive in the granulation tissue of group of astragalus polysaccharide+collagen as compared with the control group were significantly different; in wound tissue, the expression of VEGF and integrinαvβ3 was verified by RT-PCR, that suggesting its role in promoting angiogenesis accounts for an important position. Conclusion:The moderate dose (20ug/ml) astragalus polysaccharides modified-collagen can significantly promote angiogenesis, and there was a synergistic effect between them, its mechanism was associated with the expression of VEGF and integrinαvβ3.All the results would suggest that Chinese medicine in promoting the application of angiogenesis have good prospects.