Protective Effect Of Breviscapine On Renal Ischemia/Reperfusion Injury In Aged Rats

Objective: To study the protective effect and mechanism of breviscapine on renal ischemia/reperfusion injury in aged rats.Methods:forty Sprague-Dawley rats 25 months aged were randomly divided into five groups (n=8 in each),sham operation group, ischemia/reperfusion group, low-dose breviscapine group, middle-dose breviscapine group and high-dose breviscapine group. The rat's RIRI model was established by resecting the right renal firstly, followed by blocking the blood flow of left renal for 45 min, at last reperfusion 24 hours. All groups need to establish RIRI model except the sham operation group which was established by cutting the right renal and then closing the incision after 45 minutes.Each group was dealt as follow:The sham operation group were injected the same mount of physiological saline(NS) through caudal vein five days before operation and administration on schedual after operation.The ischemia/reperfusion group was dealt the same as sham operation group.All breviscapine groups were injected with bresiacapine injection(low-dose 5mg/kg,middle-dose 10 mg/kg,high-dose 20mg/kg) through caudal vein three days before operation and administration on schedual after operation. All S-D rats in different groups were injected with penicillin injection(10000u/100g)before operation deprived water from 12h to 24 hours after operation and then were killed. Renal function was assessed by measuring serum creatinine and blood urea.The MDA content were measured by Thibabituric Acid technique and SOD activity were measured xanthine oxidase technique.The apoptosis rate in renal tissue were measured by TUNEL method. The protein expression of Caspase-3,Bax and Bcl-2 was measured by immunohistochemistry. The active protein fragment expression of p53 was measured by western-blotting.Results :⑴The serum creatinine and blood urea of ischemia /reperfusion group were markedly higher than Sham operation group (P<0.05).Compaired with ischemia/reperfusion group,the serum creatinine and blood urea of the three treatment group were markedly lower(P<0.05). High-dose breviscapine group was more effective than the other low dose group.Among the three breviscapine group,blood urea had no statistical difference (P>0.05).⑵Sham operation group were found the change of glomerulosclerosis, basement membrane shrinkage, and part of the renal tubular epithelial cells atrophy.Renal tubular expansion, Cellular edema and hypochromatosis of nephric tubule in ischemia/reperfusion group were more obvious than Sham operation group.Compaired with ischemia/reperfusion group,The pathology change of the three treatment group were markedly decreased.⑶The MDA content in ischemia/reperfusion group was markedly higher than Sham operation group,the SOD activity in ischemia/reperfusion group was markedly lower than Sham operation group(P<0.05).Compaired with the ischemia/reperfusion group,the MDA content were markedly decreased(P<0.05),and the SOD activity was significantly increased in the three breviscapine group(P<0.05).The MDA content in high-dose breviscapine group was obviously lower than the other two breviscapine groups(P<0.05),but the SOD activity was obviously higher than the other two treatment groups(P<0.05).The MDA content and the SOD activity had no statistical difference between the low-dose and middle-dose breviscapine group.⑷The apoptosis ration of ischemia reperfusion group was obviously higher than Sham operation group(P<0.05).The apoptosis ration of the three treatment groups was markedly lower than ischemia/reperfusion group(P<0.05).And among the three treatment groups, high-dose breviscapine group was obviously lower than the other two treatment groups(P<0.05).But there was no statistical difference between the low-dose and middle-dose breviscapine group(P>0.05).⑸The protein expression of Caspase-3, Bcl-2 and Bax in ischemia/reperfusion group was markedly higher than Sham operation group(P<0.05).Compaired with the ischemia/reperfusion group,the protein expression of Caspase-3 and Bax were markedly decreased(P<0.05), and the protein expression of Bcl-2 was significantly increased in the three treatment group(P<0.05).The protein expression of Caspase-3 and Bax in high-dose breviscapine group was obviously lower than the other two breviscapine groups(P<0.05),but the Bcl-2 protein expression was obviously higher than the other two treatment groups(P<0.05).The protein expression of Caspase-3,Bcl-2 and Bax had no statistical difference between the low-dose and middle-dose breviscapine group (P>0.05).⑹The protein expression of p53 in ischemia/reperfusion group was markedly higher than Sham operation group(P<0.05). Compaired with the ischemia/reperfusion group,the protein expression of p53 was significantly decreased in the three treatment group(P<0.05).The protein expression of p53 in low-dose breviscapine group was obviously higher than the other two treatment groups(P<0.05).The protein expression of p53 had no statistical difference between the high-dose and middle-dose breviscapine group (P>0.05).Conclusion:The aging change of renal morphology in the aged rat,which were easy to induced renal ischemia reperfusion injury.The serum creatinine,blood urea,MDA content,apoptosis ration,Caspase-3,Bcl-2,Bax and p53 protein expression were increased in the ischemia/reperfusion injury.The Caspase-3,Bax and p53 protein expression,serum creatinine, blood urea and apoptosis ration were significantly deceased after treated with breviacapine injection. However the SOD activity and Bcl-2 protein expression were significantly increased after treated with breviacapine injection.Taken together, these results show that breviacapine injection significantly reduces the renal dysfunction caused by the ischemia/reperfusion injury.And the protective effect of breviacapine injection may be related to the clearance of reactive oxygen series and the inhibition of apoptosis,Caspase-3, Bax and p53 protein expression, and increasing of Bcl-2 protein expression.