Protective Effect And Mechanism Of Effective Components Of Aconitum Flavescens On Vascular Endothelial Injury

Objective:1.A variety of vascular endothelial injury models?hydrogen peroxide,high glucose,high fat induction?were established in vitro.The effective components of Aconitum flavescens for repairing vascular endothelial injury were screened by these models,and its mechanism was discussed.2.The rat vascular endothelial injury model induced by ISO was established in vivo to observe the effect of the effective components of Astragalus membranaceus on vascular endothelium and myocardial tissue in rats with myocardial fibrosis.Methods:1.Establishment of three models of vascular endothelial injury?1?Different concentrations of hydrogen peroxide?1,2,3 m M?were incubated for 10,20,and 30 min,respectively,and the optimal modeling concentration and modeling time were determined by examining the degree of vasodilation.?2?Different concentrations of glucose?22,44,88 m M?were incubated for 0.5,1,2 h,respectively.The optimal modeling concentration and modeling time were determined by examining the degree of vasodilation.?3?Different concentrations of OX-LDL?25,50,100?g/m L?were incubated for 15,30 and 60 min respectively.The optimal model concentration and modeling time were determined by examining the degree of vasodilation.2.Astragalus polysaccharide?ACP?and Astragalus aconitine?ACA?improve vascular endothelial injuryDifferent concentrations of ACP and ACA were incubated with optimal modeling conditions of hydrogen peroxide,high glucose and high fat,respectively.According to the degree of vasodilation compared with the model group,the repairing effect and mechanism of ACA and ACP on vascular endothelial injury were explored.3.ACP and ACA improve the vascular endothelial injury in rats?1?In vivo experiment,200 g SD rats were divided into 7 groups,8 in each group:the blank group,model group,positive group,ACP group?200 mg/kg?,ACA group?100mg/kg?,combined high-dose group?200 mg/kg ACP+100 mg/kg ACA?,combined with low-dose group?100 mg/kg ACP+50 mg/kg ACA?.?2?The effects of ACP and ACA on the expression of inflammatory related indexes in left ventricular tissue of rats were detected by Western blot.?3?The effect of vasodilation in each group was measured by vivo vascular ring diastolic test.?4?The morphological changes of the thoracic aorta and secondary mesenteric artery were examined by HE staining.Results:1.Incubate the vascular rings with different concentrations of hydrogen peroxide.When the concentration of hydrogen peroxide is 2 m M for 20 min,the degree of vascular injury was about 37%.Considering the changes in vasoconstriction of the blood vessels,incubating with 2 m M H₂O₂for 20 min was the best vascular endothelium damage by H₂O₂mold conditions.2.Incubate the vascular rings with 44 m M glucose for 2 h,the vasodilatation was about 40%.Considering the change of vasoconstriction of the blood vessels,the vessels were incubated with 44 m M glucose for 2 h for high glucose damage.Optimal modeling conditions for vascular endothelium.3.Incubate the vascular rings with different concentrations of OX-LDL at concentrations of 50?g/m L for 30 min,the vasodilatation was about 60%.Considering the change of vasoconstriction of the blood vessels,used 50?g/m L OX-LDL for 30min is the optimal modeling condition.4.The results of ex vivo vascular ring diastolic test showed that 2m M H₂O₂was incubated with 100,200,400?g/m L ACP for 20 min in vitro,and the optimal vasoconstriction was about 70%at 200?g/m L.ACP has a good protective effect on H₂O₂damaged blood vessels.2 m M H₂O₂was incubated with 100,200,400?g/m L ACA for 20 min.The optimal vasoconstriction at 100?g/m L was about 50%,indicating that ACA has less effect on H₂O₂injury than ACP.5.44 mM glucose was incubated with 100,200,400?g/m L ACP for 2 h,and the optimal vasoconstriction was about 70%at 200?g/m L,indicating that ACP has a good protective effect on high glucose injured blood vessels.44 m M glucose was incubated with 100,200,400?g/m L ACA for 2 h.The optimal vasoconstriction at 100?g/m L was about 80%,indicating that ACA has a good protective effect on high glucose-damaged blood vessels.6.50?g/mL OX-LDL was incubated with 100,200,400?g/m L ACP for 30 min.It was found that ACP had obvious effect on vascular endothelial repair of OX-LDL injury.As the concentration increases,the repairing effect was gradually enhanced.50?g/m L OX-LDL was incubated with 100,200,400?g/m L ACA for 30 min,and it was found that ACA had no obvious effect on vascular endothelial repair of OX-LDL injury.7.The results of:Western blot in vivo showed that The results of tissue protein expression in rats with vascular endothelial injury induced by ISO showed that ACP,ACA and high dose groups could significantly promote the phosphorylation of Akt,e NOS,I-?B and the protein expression of IL-10?IL-10?.ACP,ACA and high dose groups could also significantly reduce the protein expression of TLR-4,i NOS and NF-?B pathway.The results of HE staining showed that in the blank group,the smooth muscle cells of the thoracic aorta and the secondary mesenteric artery were arranged neatly,the internal elastic layer was closely connected,the blood vessels were clear and the endothelium was complete.In the model group,the endothelium was incomplete,the arrangement of smooth muscle cells was loose and disordered,the vascular structure was not clear,the elastic membrane was ruptured,the endothelial cells were enlarged and deformed,some of them fell off and died,and the intima was infiltrated by inflammatory cells.Compared with the model group,the inflammatory reaction was alleviated,the number of endothelial cells and smooth muscle cells were significantly increased,the number of endothelial cells fell off,and the degree of inflammatory cell infiltration decreased.Conclusions:2 mM H₂O₂ incubated blood vessels for 20 min,44 m M glucose for 2 hours,and50?g/m L OX-LDL for 30 min were best conditions for modeling blood vessels.ACP and ACA can improve the above three vascular endothelial injury models.ACP and ACA can also improve the inflammation and endothelial injury induced by ISO in rats.