| Objective: To observe the changes of blood vessel morphology under light microscope by HE staining to verify the success of the model;to detect the levels of serum nitric oxide(NO),endothelin-1(ET-1),and measure the changes of m RNA expressions,such as vascular endothelial growth factor(VEGF),platelet-derived growth factor(PDGF),basic fibroblast growth factor(b FGF)and other factors,to observe the effects of Guantong Formula and its disassembled prescriptions on restenosis(RS)in multiple perspectives,in order to illustrate its underlying mechanism,and to provide experimental evidences for effective TCM formulas for RS prevention and treatment.Methods:No.1 experiment: A total of 144 healthy male SD rats were randomly assigned into six groups,which were normal group,model group,sham-operated group,Guantong Formula group,disassembled prescription group with removing blood stasis,and disassembled prescriptions group with phlegm elimination,and were further divided into three subgroups according to time points of 1,7,and28 d,with 8 rats in each subgroup.The normal group was given no treatment,the sham-operated group was not given balloon injury,and the other groups conducted balloon injuries on right arteria carotis communis.The drug groups was given drug gavage for 10 ml per kilogram daily,ranging from preoperative 4 days to postoperative 28 days,while equal amount of normal saline gavage was conducted for rats in sham-operated group and normal group.All the rats were sacrificed at different postoperative time points,and the vascular segment of right arteria carotis communis was collected to observe vascular condition by HE staining.Serum was separated from blood in aorta abdominalis to detect the level of nitric oxide(NO)and endothelin 1(ET-1)by nitric reductases and ELISA,respectively.No.2 experiment: A total of 60 healthy male SD rats were randomly divided into normal group,model group,sham-operated group and Guantong Formula group,and each group were assigned into three subgroups at 1,7 and 28 d,with 5 rats in each subgroup.The processing was the same as experiment 1,and the rats were sacrificed at different time points.Eventually,m RNA expression of vascular endothelial growth factor(VEGF),platelet-derived growth factor(PDGF),basic fibroblast growth factor(b FGF)in arteria carotis communis were detected by real time PCR(RT-PCR).Results:1.HE staining results showed that Guantong Formula and its disassembled prescriptions could obviously improve the postoperative restenosis(RS)of balloon injury in arteria carotis communis,but Guantong Formula had the best therapeutic effect since it improved the severity of vascular stenosis in the greatest degree.2.Compared with normal group,the NO value was reduced in sham-operated group,and no significant differences were found(P>0.05),but was significantly increased than the other groups(P<0.05).The model group had the lowest NO value,with significant differences(P<0.01).Guantong Formula group was lower in NO value than normal group and sham-operated group,but significantly higher than model group and the two disassembled prescription groups(P<0.01).The disassembled prescription group with blood stasis activation was slightly lower than Guantong Formula group,but significantly higher than disassembled prescription group with phlegm elimination and model group(P<0.01).The serum NO content was decreased at 1,7,28 d in all groups,and there were intra-group significant differences in all groups at three time points(P<0.01),except for model group at 1,7 d and sham-operated group at all time points(P>0.05).3.Compared with the normal group,the sham operation group had increased ET-1 levels at different time points,significant differences were found at 1,7 d,but no difference at 28 d(P>0.05).The model group had the highest level of ET-1,which showed significant differences when compared with the other groups(P<0.01);Guantong Formula group had higher level of ET-1 than the normal group and sham-operated group,but lower than the model group and two disassembled prescriptions groups(P<0.01).The disassembled prescription group with activating blood and removing stasis was higher than Guantong Formula group,and lower than disassembled prescription group with sputum elimination and model group(P<0.01).Disassembled prescription group with sputum elimination had higher content of ET-1 than other groups except for model group(P<0.01),but showed no significant differences when compared to disassembled prescription group with blood stasis removing group at 28 d(P>0.05).The levels of ET-1 were decreased in all groups,which showed significant between-group differences at different time points except for at 1,7,and 7,28 d when compared with the sham-operated group(P<0.01).4.The m RNA expressions were similar in growth factors including VEGF,PBGF,b FGF.There were significant differences in reduction of m RNA expressions in each group at 1,7,28 d,and no significant difference was observed in normal group,and sham-operated group,significant differences were found in model group and Guantong Formula group.Conclusions:Guantong Formula can inhabit proliferation of vascular smooth muscle cells after balloon injury of arteria carotis communis in rats,and reduce the recurrence rate of postoperative stenosis,and its mechanism may correlated with enhancing serum level of NO and inhibiting the serum level of ET-1,and m RNA expression of VEGF,PDGF and b FGF to prevents RS.The results reveals that Guantong Formula has the best efficiency,and disassembled prescriptions with activating blood and removing stasis has better clinical outcomes than other disassembled prescriptions,so better clinical outcomes of Guantong Formula is originated from blood-activating and stasis-eliminating drugs. |
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